Donor-derived stem-cells and epithelial mesenchymal transition in squamous cell carcinoma in transplant recipients.

PubMed

Verneuil, Laurence; Leboeuf, Christophe; Bousquet, Guilhem; Brugiere, Charlotte; Elbouchtaoui, Morad; Plassa, Louis-François; Peraldi, Marie-Noelle; Lebbé, Celeste; Ratajczak, Philippe; Janin, Anne

2015-12-08

Skin squamous-cell-carcinoma (SCC), is the main complication in long-term kidney-transplant recipients, and it can include donor-derived cells. Preclinical models demonstrated the involvement of epithelial mesenchymal transition (EMT) in the progression of skin SCC, and the role of Snail, an EMT transcription factor, in cancer stem-cell survival and expansion.Here, we studied stem-cells and EMT expression in SCCs and concomitant actinic keratoses (AK) in kidney-transplant recipients. In SCC and AK in 3 female recipients of male kidney-transplants, donor-derived Y chromosome in epidermal stem cells was assessed using combined XY-FISH/CD133 immunostaining, and digital-droplet-PCR on laser-microdissected CD133 expressing epidermal cells.For EMT study, double immunostainings of CD133 with vimentin or snail and slug, electron microscopy and immunostainings of keratinocytes junctions were performed. Digital droplet PCR was used to check CDH1 (E-cadherin) expression level in laser-microdissected cells co-expressing CD133 and vimentin or snail and slug.The numbers of Y-chromosome were assessed using digital droplet PCR in laser-microdissected cells co-expressing CD133 and vimentin, or snail and slug, and in CD133 positive cells not expressing any EMT maker. We identified donor-derived stem-cells in basal layers and invasive areas in all skin SCCs and in concomitant AKs, but not in surrounding normal skin.The donor-derived stem-cells expressed the EMT markers, vimentin, snail and slug in SCCs but not in AKs. The expression of the EMT transcription factor, SNAI1, was higher in stem-cells when they expressed vimentin. They were located in invasive areas of SCCs. In these areas, the expressions of claudin-1 and desmoglein 1 were reduced or absent, and within the basal layer there were features of basal membrane disappearance.Donor-derived stem cells were in larger numbers in stem cells co-expressing vimentin or snail and slug than in stem cells not expressing any EMT marker. We identified here donor-derived stem cells within skin SCC in kidney-transplant recipients. They were located in invasive areas of SCC and had EMT characteristics.

Nickel-induced Epithelial-Mesenchymal Transition by Reactive Oxygen Species Generation and E-cadherin Promoter Hypermethylation*

PubMed Central

Wu, Chih-Hsien; Tang, Sheau-Chung; Wang, Po-Hui; Lee, Huei; Ko, Jiunn-Liang

2012-01-01

Epithelial-mesenchymal transition (EMT) is considered a critical event in the pathogenesis of lung fibrosis and tumor metastasis. During EMT, the expression of differentiation markers switches from cell-cell junction proteins such as E-cadherin to mesenchymal markers such as fibronectin. Although nickel-containing compounds have been shown to be associated with lung carcinogenesis, the role of nickel in the EMT process in bronchial epithelial cells is not clear. The aim of this study was to examine whether nickel contributes to EMT in human bronchial epithelial cells. We also attempted to clarify the mechanisms involved in NiCl2-induced EMT. Our results showed that NiCl2 induced EMT phenotype marker alterations such as up-regulation of fibronectin and down-regulation of E-cadherin. In addition, the potent antioxidant N-acetylcysteine blocked EMT and expression of HIF-1α induced by NiCl2, whereas the DNA methyltransferase inhibitor 5-aza-2′-deoxycytidine restored the down-regulation of E-cadherin induced by NiCl2. Promoter hypermethylation of E-cadherin, determined by quantitative real time methyl-specific PCR and bisulfate sequencing, was also induced by NiCl2. These results shed new light on the contribution of NiCl2 to carcinogenesis. Specifically, NiCl2 induces down-regulation of E-cadherin by reactive oxygen species generation and promoter hypermethylation. This study demonstrates for the first time that nickel induces EMT in bronchial epithelial cells. PMID:22648416

Markers of epithelial-to-mesenchymal transition reflect tumor biology according to patient age and Gleason score in prostate cancer

PubMed Central

Jędroszka, Dorota; Hamouz, Raneem; Górniak, Karolina; Bednarek, Andrzej K.

2017-01-01

Introduction Prostate carcinoma (PRAD) is one of the most frequently diagnosed malignancies amongst men worldwide. It is well-known that androgen receptor (AR) plays a pivotal role in a vast majority of prostate tumors. However, recent evidence emerged stating that estrogen receptors (ERs) may also contribute to prostate tumor development. Moreover, progression and aggressiveness of prostate cancer may be associated with differential expression genes of epithelial-to-mesenchymal transition (EMT). Therefore we aimed to assess the significance of receptors status as well as EMT marker genes expression among PRAD patients in accordance to their age and Gleason score. Materials and methods We analyzed TCGA gene expression profiles of 497 prostate tumor samples according to 43 genes involved in EMT and 3 hormone receptor genes (AR, ESR1, ESR2) as well as clinical characteristic of cancer patients. Then patients were divided into four groups according to their age and 5 groups according to Gleason score. Next, we evaluated PRAD samples according to relationship between the set of variables in different combinations and compared differential expression in subsequent groups of patients. The analysis was applied using R packages: FactoMineR, gplots, RColorBrewer and NMF. Results MFA analysis resulted in distinct grouping of PRAD patients into four age categories according to expression level of AR, ESR1 and ESR2 with the most distinct group of age less than 50 years old. Further investigations indicated opposite expression profiles of EMT markers between different age groups as well as strong association of EMT gene expression with Gleason score. We found that depending on age of prostate cancer patients and Gleason score EMT genes with distinctly altered expression are: KRT18, KRT19, MUC1 and COL4A1, CTNNB1, SNAI2, ZEB1 and MMP3. Conclusions Our major observation is that prostate cancer from patients under 50 years old compared to older ones has entirely different EMT gene expression profiles showing potentially more aggressive invasive phenotype, despite Gleason score classification. PMID:29206234

Markers of epithelial-to-mesenchymal transition reflect tumor biology according to patient age and Gleason score in prostate cancer.

PubMed

Jędroszka, Dorota; Orzechowska, Magdalena; Hamouz, Raneem; Górniak, Karolina; Bednarek, Andrzej K

2017-01-01

Prostate carcinoma (PRAD) is one of the most frequently diagnosed malignancies amongst men worldwide. It is well-known that androgen receptor (AR) plays a pivotal role in a vast majority of prostate tumors. However, recent evidence emerged stating that estrogen receptors (ERs) may also contribute to prostate tumor development. Moreover, progression and aggressiveness of prostate cancer may be associated with differential expression genes of epithelial-to-mesenchymal transition (EMT). Therefore we aimed to assess the significance of receptors status as well as EMT marker genes expression among PRAD patients in accordance to their age and Gleason score. We analyzed TCGA gene expression profiles of 497 prostate tumor samples according to 43 genes involved in EMT and 3 hormone receptor genes (AR, ESR1, ESR2) as well as clinical characteristic of cancer patients. Then patients were divided into four groups according to their age and 5 groups according to Gleason score. Next, we evaluated PRAD samples according to relationship between the set of variables in different combinations and compared differential expression in subsequent groups of patients. The analysis was applied using R packages: FactoMineR, gplots, RColorBrewer and NMF. MFA analysis resulted in distinct grouping of PRAD patients into four age categories according to expression level of AR, ESR1 and ESR2 with the most distinct group of age less than 50 years old. Further investigations indicated opposite expression profiles of EMT markers between different age groups as well as strong association of EMT gene expression with Gleason score. We found that depending on age of prostate cancer patients and Gleason score EMT genes with distinctly altered expression are: KRT18, KRT19, MUC1 and COL4A1, CTNNB1, SNAI2, ZEB1 and MMP3. Our major observation is that prostate cancer from patients under 50 years old compared to older ones has entirely different EMT gene expression profiles showing potentially more aggressive invasive phenotype, despite Gleason score classification.

Osthole inhibits insulin-like growth factor-1-induced epithelial to mesenchymal transition via the inhibition of PI3K/Akt signaling pathway in human brain cancer cells.

PubMed

Lin, Ying-Chao; Lin, Jia-Ching; Hung, Chao-Ming; Chen, Yeh; Liu, Liang-Chih; Chang, Tin-Chang; Kao, Jung-Yie; Ho, Chi-Tang; Way, Tzong-Der

2014-06-04

Glioblastoma multiforme (GBM) is one of the most lethal types of tumors and highly metastatic and invasive. The epithelial-to-mesenchymal transition (EMT) is the crucial step for cancer cells to initiate the metastasis and could be induced by many growth factors. In this study, we found that GBM8401 cells were converted to fibroblastic phenotype and the space between the cells became expanded in response to insulin-like growth factor-1 (IGF-1) treatment. Epithelial markers were downregulated and mesenchymal markers were upregulated simultaneously after IGF-1 treatment. Our results illustrate that IGF-1 was able to induce EMT in GBM8401 cells. Osthole would reverse IGF-1-induced morphological changes, upregulated the expression of epithelial markers, and downregulated the expression of mesenchymal markers. Moreover, wound-healing assay also showed that osthole could inhibit IGF-1-induced migration of GBM8401 cells. By using dual-luciferase reporter assay and real-time PCR, we demonstrated that osthole inhibited IGF-1-induced EMT at the transcriptional level. Our study found that osthole decreased the phosphorylation of Akt and GSK3β and recovered the GSK3β bioactivity in inhibiting EMT transcription factor Snail and Twist expression. These results showed that osthole inhibited IGF-1-induced EMT by blocking PI3K/Akt pathway. We hope that osthole can be used in anticancer therapy and be a new therapeutic medicine for GBM in the future.

Chemical Chaperone of Endoplasmic Reticulum Stress Inhibits Epithelial-Mesenchymal Transition Induced by TGF-β1 in Airway Epithelium via the c-Src Pathway

PubMed Central

Lee, Heung-Man; Kang, Ju-Hyung; Shin, Jae-Min; Lee, Seoung-Ae

2017-01-01

Epithelial-mesenchymal transition (EMT) is a biological process that allows epithelial cells to assume a mesenchymal cell phenotype. EMT is considered as a therapeutic target for several persistent inflammatory airway diseases related to tissue remodeling. Herein, we investigated the role of endoplasmic reticulum (ER) stress and c-Src in TGF-β1-induced EMT. A549 cells, primary nasal epithelial cells (PNECs), and inferior nasal turbinate organ cultures were exposed to 4-phenylbutylic acid (4PBA) or PP2 and then stimulated with TGF-β1. We found that E-cadherin, vimentin, fibronectin, and α-SMA expression was increased in nasal polyps compared to inferior turbinates. TGF-β1 increased the expression of EMT markers such as E-cadherin, fibronectin, vimentin, and α-SMA and ER stress markers (XBP-1s and GRP78), an effect that was blocked by PBA or PP2 treatment. 4-PBA and PP2 also blocked the effect of TGF-β1 on migration of A549 cells and suppressed TGF-β1-induced expression of EMT markers in PNECs and organ cultures of inferior turbinate. In conclusion, we demonstrated that 4PBA inhibits TGF-β1-induced EMT via the c-Src pathway in A549 cells, PNECs, and inferior turbinate organ cultures. These results suggest an important role for ER stress and a diverse role for TGF-β1 in upper airway chronic inflammatory disease such as CRS. PMID:28804222

Chemical Chaperone of Endoplasmic Reticulum Stress Inhibits Epithelial-Mesenchymal Transition Induced by TGF-β1 in Airway Epithelium via the c-Src Pathway.

PubMed

Lee, Heung-Man; Kang, Ju-Hyung; Shin, Jae-Min; Lee, Seoung-Ae; Park, Il-Ho

2017-01-01

Epithelial-mesenchymal transition (EMT) is a biological process that allows epithelial cells to assume a mesenchymal cell phenotype. EMT is considered as a therapeutic target for several persistent inflammatory airway diseases related to tissue remodeling. Herein, we investigated the role of endoplasmic reticulum (ER) stress and c-Src in TGF- β 1-induced EMT. A549 cells, primary nasal epithelial cells (PNECs), and inferior nasal turbinate organ cultures were exposed to 4-phenylbutylic acid (4PBA) or PP2 and then stimulated with TGF- β 1. We found that E-cadherin, vimentin, fibronectin, and α -SMA expression was increased in nasal polyps compared to inferior turbinates. TGF- β 1 increased the expression of EMT markers such as E-cadherin, fibronectin, vimentin, and α -SMA and ER stress markers (XBP-1s and GRP78), an effect that was blocked by PBA or PP2 treatment. 4-PBA and PP2 also blocked the effect of TGF- β 1 on migration of A549 cells and suppressed TGF- β 1-induced expression of EMT markers in PNECs and organ cultures of inferior turbinate. In conclusion, we demonstrated that 4PBA inhibits TGF- β 1-induced EMT via the c-Src pathway in A549 cells, PNECs, and inferior turbinate organ cultures. These results suggest an important role for ER stress and a diverse role for TGF- β 1 in upper airway chronic inflammatory disease such as CRS.

Low molecular weight fucoidan and its fractions inhibit renal epithelial mesenchymal transition induced by TGF-β1 or FGF-2.

PubMed

Li, Xinpeng; Li, Xiaohong; Zhang, Quanbin; Zhao, Tingting

2017-12-01

We investigated the renal protective effects of low molecular weight fucoidan (LMWF) and its two fractions (F0.5 and F1.0), which were extracted from Laminaria japonica, on the epithelial-mesenchymal transition (EMT) induced by transforming growth factor beta 1 (TGF-β1) and fibroblast growth factor 2 (FGF-2) in HK-2 human renal proximal tubular cells. Cell morphology and EMT markers (fibronectin and alpha-smooth muscle actin) demonstrated that cells treated with TGF-β1 or FGF-2 developed EMT to a significant extent. Treatment with LMWF or its fractions markedly attenuated the EMT and decreased expression of the EMT markers. The F1.0 fraction, the sulfated fucan fraction, was found to be the main active component of LMWF, and heparanase (HPSE) was a key factor in renal tubular epithelial trans-differentiation. The F1.0 fraction inhibited elevated HPSE and matrix metallopeptidase 9 expression, thereby attenuating the progress of EMT. Copyright © 2017 Elsevier B.V. All rights reserved.

Reversion of malignant phenotypes of human glioblastoma cells by β-elemene through β-catenin-mediated regulation of stemness-, differentiation- and epithelial-to-mesenchymal transition-related molecules.

PubMed

Zhu, Tingzhun; Li, Xiaoming; Luo, Lihan; Wang, Xiaogang; Li, Zhiqing; Xie, Peng; Gao, Xu; Song, Zhenquan; Su, Jingyuan; Liang, Guobiao

2015-11-12

Glioblastoma is the most common and lethal type of primary brain tumor. β-Elemene, a natural plant drug extracted from Curcuma wenyujin, has shown strong anti-tumor effects in various tumors with low toxicity. However, the effects of β-elemene on malignant phenotypes of human glioblastoma cells remain to be elucidated. Here we evaluated the effects of β-elemene on cell proliferation, survival, stemness, differentiation and the epithelial-to-mesenchymal transition (EMT) in vitro and in vivo, and investigated the mechanisms underlying these effects. Human primary and U87 glioblastoma cells were treated with β-elemene, cell viability was measured using a cell counting kit-8 assay, and treated cells were evaluated by flow cytometry. Western blot analysis was carried out to determine the expression levels of stemness markers, differentiation-related molecules and EMT-related effectors. Transwell assays were performed to further determine EMT of glioblastoma cells. To evaluate the effect of β-elemene on glioblastoma in vivo, we subcutaneously injected glioblastoma cells into the flank of nude mice and then intraperitoneally injected NaCl or β-elemene. The tumor xenograft volumes were measured every 3 days and the expression of stemness-, differentiation- and EMT-related effectors was determined by Western blot assays in xenografts. β-Elemene inhibited proliferation, promoted apoptosis, impaired invasiveness in glioblastoma cells and suppressed the growth of animal xenografts. The expression levels of the stemness markers CD133 and ATP-binding cassette subfamily G member 2 as well as the mesenchymal markers N-cadherin and β-catenin were significantly downregulated, whereas the expression levels of the differentiation-related effectors glial fibrillary acidic protein, Notch1, and sonic hedgehog as well as the epithelial marker E-cadherin were upregulated by β-elemene in vitro and in vivo. Interestingly, the expression of vimentin was increased by β-elemene in vitro; this result was opposite that for the in vivo procedure. Inhibiting β-catenin enhanced the anti-proliferative, EMT-inhibitory and specific marker expression-regulatory effects of β-elemene. β-Elemene reversed malignant phenotypes of human glioblastoma cells through β-catenin-involved regulation of stemness-, differentiation- and EMT-related molecules. β-Elemene represents a potentially valuable agent for glioblastoma therapy.

SiRNA-HMGA2 weakened AGEs-induced epithelial-to-mesenchymal transition in tubular epithelial cells.

PubMed

Bai, Yi-Hua; Wang, Jia-Ping; Yang, Min; Zeng, Yi; Jiang, Hong-Ying

2015-02-20

Diabetic nephropathy as the most common cause of end-stage renal disease accounts for a significant increase in morbidity and mortality in patients. Epithelial to mesenchymal transition (EMT) of tubular cells is associated with diabetic nephropathy. Advanced glycation end products (AGEs) are thought to be involved in the pathogenesis of diabetic nephropathy via multifactorial mechanisms. However, whether AGEs could induce EMT in Tubular epithelial cells is still unknown. In this study, we found that AGEs induced EMT and accompanied by reduced expression of the epithelial markers E-cadherin and enhanced expression of the mesenchymal markers vimentin and alpha-smooth muscle actin. Furthermore, the expression of HMGA2 was upregulated by AGEs. Far more interesting, its knockdown by short interfering RNA (siRNA) effectively reversed AGEs-induced EMT. Meanwhile, we also found that knockdown of HMGA2 inhibited high AGEs-induced generation of reactive oxygen species (ROS) and the activation of p38 MAPK. Collectively, these studies suggest that HMGA2 plays a important role in EMT during Diabetic nephropathy and more study toward HMGA2 should be played in renal pathogenesis. Copyright © 2015 Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Tsai, Jie-Heng; Hsu, Li-Sung; Clinical Laboratory, Chung Shan Medical University Hospital, Taichung 402, Taiwan, ROC

The molecular basis of epithelial–mesenchymal transition (EMT) functions as a potential therapeutic target for breast cancer because EMT may endow breast tumor-initiating cells with stem-like characteristics and enable the dissemination of breast cancer cells. We have recently verified the antitumor activity of 3,5,4′-trimethoxystilbene (MR-3), a naturally methoxylated derivative of resveratrol, in colorectal cancer xenografts via an induction of apoptosis. The effect of MR-3 on EMT and the invasiveness of human MCF-7 breast adenocarcinoma cell line were also explored. We found that MR-3 significantly increased epithelial marker E-cadherin expression and triggered a cobblestone-like morphology of MCF-7 cells, while reciprocally decreasing themore » expression of mesenchymal markers, such as snail, slug, and vimentin. In parallel with EMT reversal, MR-3 downregulated the invasion and migration of MCF-7 cells. Exploring the action mechanism of MR-3 on the suppression of EMT and invasion indicates that MR-3 markedly reduced the expression and nuclear translocation of β-catenin, accompanied with the downregulation of β-catenin target genes and the increment of membrane-bound β-catenin. These results suggest the involvement of Wnt/β-catenin signaling in the MR-3-induced EMT reversion of MCF-7 cells. Notably, MR-3 restored glycogen synthase kinase-3β activity by inhibiting the phosphorylation of Akt, the event required for β-catenin destruction via a proteasome-mediated system. Overall, these findings indicate that the anti-invasive activity of MR-3 on MCF-7 cells may result from the suppression of EMT via down-regulating phosphatidylinositol 3-kinase (PI3K)/AKT signaling, and consequently, β-catenin nuclear translocation. These occurrences ultimately lead to the blockage of EMT and the invasion of breast cancer cells. - Highlights: • MR-3 blocked MCF-7 cell invasion by inducing a reversal of EMT. • Wnt/β-catenin signaling is involved in MR-3-induced EMT reversion of MCF-7 cells. • Knockdown of β-catenin was sufficient to restore epithelial marker E-cadherin levels. • MR-3 recovered the function of GSK-3β that inhibits β-catenin nuclear translocation.« less

ZEB1 promotes the progression and metastasis of cervical squamous cell carcinoma via the promotion of epithelial-mesenchymal transition

PubMed Central

Ma, Yihui; Zheng, Xiangyu; Zhou, Jun; Zhang, Ying; Chen, Kuisheng

2015-01-01

Objective: The process of epithelial-mesenchymal transition (EMT) clearly contributes to cancer metastasis. The aim of this study was to investigate the expression of the EMT-related transcription repressor ZEB1 and the expression of EMT-associated markers (E-cadherin, β-catenin and N-cadherin) in cervical squamous cell carcinoma. In addition, the role of ZEB1 and these EMT-associated markers in the progression and metastasis of cervical squamous cell carcinoma was explored. Methods: The expression of ZEB1, E-cadherin, β-catenin and N-cadherin was evaluated in 81 specimens of cervical squamous cell carcinoma by immunohistochemistry; the clinicopathological significance of these markers was then analyzed. Results: 1) Of the 81 samples, 37 cases (45.7%) were positive for ZEB1, and nuclear expression of ZEB1 in tumor cells was positively associated with the differentiation status of the tumor tissue (P < 0.05), vascular invasion (P < 0.05) and lymph node metastasis (P < 0.05). 2) The loss of E-cadherin and β-catenin expression in tumor cells and the acquisition of N-cadherin expression were positively associated with the differentiation status of the tumor tissue (P < 0.05) and with the occurrence of vascular invasion (P < 0.05). 3) A significant negative correlation was observed between ZEB1 and E-cadherin expression (Spearman = -0.636, P < 0.05) and between ZEB1 and β-catenin expression (Spearman = -0.417, P < 0.05). Moreover, a significant positive correlation was observed between ZEB1 and N-cadherin expression (Spearman = 0.557, P < 0.05). Conclusions: These results emphasize the role of EMT in cervical squamous cell carcinoma. The upregulation of ZEB1 is associated with the abnormal expression of E-cadherin, β-catenin and N-cadherin, which might promote the progression and metastasis of cervical squamous cell carcinoma. PMID:26617850

Targeting Androgen Receptor to Suppress Macrophage-induced EMT and Benign Prostatic Hyperplasia (BPH) Development

PubMed Central

Lu, Tianjing; Lin, Wen-Jye; Izumi, Kouji; Wang, Xiaohai; Xu, Defeng; Fang, Lei-Ya; Li, Lei; Jiang, Qi

2012-01-01

Early studies suggested macrophages might play roles in inflammation-associated benign prostatic hyperplasia (BPH) development, yet the underlying mechanisms remain unclear. Here we first showed that CD68+ macrophages were identified in both epithelium and the stromal area of human BPH tissues. We then established an in vitro co-culture model with prostate epithelial and macrophage cell lines to study the potential impacts of infiltrating macrophages in the BPH development and found that co-culturing prostate epithelial cells with macrophages promoted migration of macrophages. In a three-dimensional culture system, the sphere diameter of BPH-1 prostate cells was significantly increased during coculture with THP-1 macrophage cells. Mechanism dissection suggested that expression levels of epithelial-mesenchymal transition (EMT) markers, such as N-cadherin, Snail, and TGF-β2, were increased, and administration of anti-TGF-β2 neutralizing antibody during co-culture suppressed the EMT and THP-1-mediated growth of BPH-1 cells, suggesting THP-1 might go through EMT to influence the BPH development and progression. Importantly, we found that modulation of androgen receptor (AR) in BPH-1 and mPrE cells significantly increased THP-1 and RAW264.7 cell migration, respectively, and enhanced expression levels of EMT markers, suggesting that AR in prostate epithelial cells might play a role in promoting macrophage-mediated EMT in prostate epithelial cells. Silencing AR function via an AR degradation enhancer, ASC-J9, decreased the macrophage migration to BPH-1 cells and suppressed EMT marker expression. Together, these results provide the first evidence to demonstrate that prostate epithelial AR function is important for macrophage-mediated EMT and proliferation of prostate epithelial cells, which represents a previously unrecognized role of AR in the cross-talk between macrophages and prostate epithelial cells. These results may provide new insights for a new therapeutic approach to battle BPH via targeting AR and AR-mediated inflammatory signaling pathways. PMID:22915828

Targeting androgen receptor to suppress macrophage-induced EMT and benign prostatic hyperplasia (BPH) development.

PubMed

Lu, Tianjing; Lin, Wen-Jye; Izumi, Kouji; Wang, Xiaohai; Xu, Defeng; Fang, Lei-Ya; Li, Lei; Jiang, Qi; Jin, Jie; Chang, Chawnshang

2012-10-01

Early studies suggested macrophages might play roles in inflammation-associated benign prostatic hyperplasia (BPH) development, yet the underlying mechanisms remain unclear. Here we first showed that CD68(+) macrophages were identified in both epithelium and the stromal area of human BPH tissues. We then established an in vitro co-culture model with prostate epithelial and macrophage cell lines to study the potential impacts of infiltrating macrophages in the BPH development and found that co-culturing prostate epithelial cells with macrophages promoted migration of macrophages. In a three-dimensional culture system, the sphere diameter of BPH-1 prostate cells was significantly increased during coculture with THP-1 macrophage cells. Mechanism dissection suggested that expression levels of epithelial-mesenchymal transition (EMT) markers, such as N-cadherin, Snail, and TGF-β2, were increased, and administration of anti-TGF-β2 neutralizing antibody during co-culture suppressed the EMT and THP-1-mediated growth of BPH-1 cells, suggesting THP-1 might go through EMT to influence the BPH development and progression. Importantly, we found that modulation of androgen receptor (AR) in BPH-1 and mPrE cells significantly increased THP-1 and RAW264.7 cell migration, respectively, and enhanced expression levels of EMT markers, suggesting that AR in prostate epithelial cells might play a role in promoting macrophage-mediated EMT in prostate epithelial cells. Silencing AR function via an AR degradation enhancer, ASC-J9, decreased the macrophage migration to BPH-1 cells and suppressed EMT marker expression. Together, these results provide the first evidence to demonstrate that prostate epithelial AR function is important for macrophage-mediated EMT and proliferation of prostate epithelial cells, which represents a previously unrecognized role of AR in the cross-talk between macrophages and prostate epithelial cells. These results may provide new insights for a new therapeutic approach to battle BPH via targeting AR and AR-mediated inflammatory signaling pathways.

Aggressive Phenotype of Cells Disseminated via Hematogenous and Lymphatic Route in Breast Cancer Patients.

PubMed

Markiewicz, Aleksandra; Nagel, Anna; Szade, Jolanta; Majewska, Hanna; Skokowski, Jaroslaw; Seroczynska, Barbara; Stokowy, Tomasz; Welnicka-Jaskiewicz, Marzena; Zaczek, Anna J

2018-06-01

Intratumoral heterogeneity of breast cancer remains a major challenge in successful treatment. Failure of cancer therapies can also be accredited to inability to systemically eradicate cancer stem cells (CSCs). Recent evidence points to the role of epithelial-mesenchymal transition (EMT) in expanding the pool of tumor cells with CSCs features. Thus, we assessed expression level as well as heterogeneity of CSCs markers in primary tumors (PT), lymph node metastasis (LNM), and circulating tumor cells (CTCs)-enriched blood fractions in order to correlate them with signs of EMT activation as well as clinicopathological data of breast cancer patients. Level of CSCs markers (ALDH1, CD44, CD133, OCT-4, NANOG) and EMT markers was quantified in PT (N=107), LNM (N=56), and CTCs-enriched blood fractions (N=85). Heterogeneity of CSCs markers expression within each PT and LNM was assessed by calculating Gini Index. Percentage of ALDH1-positive cells was elevated in PT in comparison to LNM (P = .005). However, heterogeneity of the four CSCs markers: ALDH1 (P = .019), CD133 (P = .009), OCT-4 (P = .027), and CD44 (P < .001) was decreased in LNM. Samples classified as mesenchymal (post-EMT) showed elevated expression of CSCs markers (OCT-4 and CD44 in PT; OCT-4 in LNM; ALDH1, OCT-4, NANOG, CD44 in CTCs). Patients with mesenchymal-like CTCs had worse prognosis than patients with epithelial-like or no CTCs (P = .0025). CSCs markers are enriched in PT, LNM, and CTCs with mesenchymal features, but their heterogeneity is decreased in metastatic lymph nodes. Mesenchymal CTCs phenotype correlates with poor prognosis of the patients. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

Overexpression of Human Papillomavirus Type 16 Oncoproteins Enhances Epithelial-Mesenchymal Transition via STAT3 Signaling Pathway in Non-Small Cell Lung Cancer Cells.

PubMed

Zhang, Wenzhang; Wu, Xin; Hu, Liang; Ma, Yuefan; Xiu, Zihan; Huang, Bingyu; Feng, Yun; Tang, Xudong

2017-05-24

The human papillomavirus (HPV) infection may be associated with the development and progression of non-small cell lung cancer (NSCLC). However, the role of HPV-16 oncoproteins in the development and progression of NSCLC is not completely clear. Epithelial-mesenchymal transition (EMT), a crucial step for invasion and metastasis, plays a key role in the development and progression of NSCLC. Here we explored the effect of HPV-16 oncoproteins on EMT and the underlying mechanisms. NSCLC cell lines, A549 and NCI-H460, were transiently transfected with the EGFP-N1-HPV-16 E6 or E7 plasmid. Real-time PCR and Western blot analysis were performed to analyze the expression of EMT markers. A protein microarray was used to screen the involved signaling pathway. Our results showed that overexpression of HPV-16 E6 and E7 oncoproteins in NSCLC cells significantly promoted EMT-like morphologic changes, downregulated the mRNA and protein levels of EMT epithelial markers (E-cadherin and ZO-1), and upregulated the mRNA and protein levels of EMT mesenchymal markers (N-cadherin and vimentin) and transcription factors (ZEB-1 and Snail-1). Furthermore, the HPV-16 E6 oncoprotein promoted STAT3 activation. Moreover, WP1066, a specific signal transducer and activator of transcription 3 (STAT3) inhibitor, reversed the effect of HPV-16 E6 on the expression of ZO-1, vimentin, and ZEB-1 in transfected NSCLC cells. Taken together, our results suggest that overexpression of HPV-16 E6 and E7 oncoproteins enhances EMT, and the STAT3 signaling pathway may be involved in HPV-16 E6-induced EMT in NSCLC cells.

NANOG regulates epithelial-mesenchymal transition and chemoresistance in ovarian cancer.

PubMed

Qin, Shan; Li, Yanfang; Cao, Xuexia; Du, Jiexian; Huang, Xianghua

2017-02-28

A key transcription factor associated with poor prognosis and resistance to chemotherapy in ovarian cancer is NANOG. However, the mechanism by which NANOG functions remains undefined. It has been suggested that epithelial-to-mesenchymal transition (EMT) also contributes to development of drug resistance in different cancers. We thus determined whether NANOG expression was associated with EMT and chemoresistance in epithelial ovarian cancer cells. NANOG expression was increased in epithelial ovarian cancer cell lines compared with its expression in normal epithelial ovarian cell lines. NANOG expression in SKOV-3 or OV2008 cells directly correlated with high expression of mesenchymal cell markers and inversely with low expression of epithelial cell marker. RNAi-mediated silencing of NANOG in SKOV-3 reversed the expression of mesenchymal cell markers and restored expression of E-cadherin. Reversibly, stable overexpression of NANOG in Moody cells increased expression of N-cadherin whereas down-regulating expression of E-cadherin, cumulatively indicating that NANOG plays an important role in maintaining the mesenchymal cell markers. Modulating NANOG expression did not have any effect on proliferation or colony formation. Susceptibility to cisplatin increased in SKOV-3 cells on down-regulating NANOG and reversible results were obtained in Moody cells post-overexpression of NANOG. NANOG silencing in SKOV-3 and OV2008 robustly attenuated in vitro migration and invasion. NANOG expression exhibited a biphasic pattern in patients with ovarian cancer and expression was directly correlated to chemoresistance retrospectively. Cumulatively, our data demonstrate that NANOG expression modulates chemosensitivity and EMT resistance in ovarian cancer. © 2017 The Author(s).

Over-expression of FoxM1 leads to epithelial-mesenchymal transition and cancer stem cell phenotype in pancreatic cancer cells

PubMed Central

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

2011-01-01

FoxM1 is known to play important role in the development and progression of many malignancies including pancreatic cancer. Studies have shown that the acquisition of Epithelial-to-mesenchymal transition (EMT) phenotype and induction of cancer stem cell (CSC) or cancer stem-like cell phenotypes are highly inter-related, and contributes to drug resistance, tumor recurrence and metastasis. The molecular mechanism(s) by which FoxM1 contributes to the acquisition of EMT phenotype and induction of CSC self-renewal capacity is poorly understood. Therefore, we established FoxM1 over-expressing pancreatic cancer (AsPC-1) cells, which showed increased cell growth, clonogenicity and cell migration. Moreover, over-expression of FoxM1 led to the acquisition of EMT phenotype by activation of mesenchymal cell markers, ZEB1, ZEB2, Snail2, E-cadherin, and vimentin, which is consistent with increased sphere-forming (pancreatospheres) capacity and expression of CSC surface markers (CD44 and EpCAM). We also found that over-expression of FoxM1 led to decreased expression of miRNAs (let-7a, let-7b, let-7c, miR-200b and miR-200c); however, re-expression of miR-200b inhibited the expression of ZEB1, ZEB2, vimentin as well as FoxM1, and induced the expression of E-cadherin, leading to the reversal of EMT phenotype. Finally, we found that genistein, a natural chemo-preventive agent, inhibited cell growth, clonogenicity, cell migration and invasion, EMT phenotype, and formation of pancreatospheres consistent with reduced expression of CD44 and EpCAM. These results suggest, for the first time, that FoxM1 over-expression is responsible for the acquisition of EMT and CSC phenotype, which is in part mediated through the regulation of miR-200b and these processes, could be easily attenuated by genistein. PMID:21503965

Suppression of the epidermal growth factor receptor inhibits epithelial-mesenchymal transition in human pancreatic cancer PANC-1 cells.

PubMed

Chang, Zhi-Gang; Wei, Jun-Min; Qin, Chang-Fu; Hao, Kun; Tian, Xiao-Dong; Xie, Kun; Xie, Xue-Hai; Yang, Yin-Mo

2012-05-01

Aberrant expression of epidermal growth factor receptor (EGFR) has been detected in pancreatic cancer; however, the mechanisms of EGFR in inducing pancreatic cancer development have not been adequately elucidated. The objective of this study was to determine the role of EGFR in mediating epithelial-mesenchymal transition (EMT) in pancreatic cancer cells. Pancreatic cancer cell line PANC-1 was transfected with small interfering RNA of EGFR by use of a lentiviral expression vector to establish an EGFR-knockdown cell line (si-PANC-1). PANC-1 cells transfected with lentiviral vector expressing negative control sequence were used as negative control (NC-PANC-1). Scratch assay and transwell study were used to analyze cell migration and invasion. Real-time PCR and Western blotting were used to detect the expression of EMT markers E-cadherin, N-cadherin, vimentin, and fibronectin and transcription factors snail, slug, twist1, and sip1 in PANC-1, NC-PANC-1, and si-PANC-1 cells. Immunofluorescent staining with these antibodies and confocal microscopy were used to observe their cellular location and morphologic changes. After RNA interference of EGFR, the migration and invasion ability of si-PANC-1 cells decreased significantly. The expression of epithelial phenotype marker E-cadherin increased and the expression of mesenchymal phenotype markers N-cadherin, vimentin, and fibronectin decreased, indicating reversion of EMT. We also observed intracellular translocation of E-cadherin. Expression of transcription factors snail and slug in si-PANC-1 cells decreased significantly. Suppression of EGFR expression can significantly inhibit EMT of pancreatic cancer PANC-1 cells. The mechanism may be related with the down-regulation of the expression of transcription factors snail and slug.

Suppression of the Epidermal Growth Factor-like Domain 7 and Inhibition of Migration and Epithelial-Mesenchymal Transition in Human Pancreatic Cancer PANC-1 Cells.

PubMed

Wang, Yun-Liang; Dong, Feng-Lin; Yang, Jian; Li, Zhi; Zhi, Qiao-Ming; Zhao, Xin; Yang, Yong; Li, De-Chun; Shen, Xiao-Chun; Zhou, Jin

2015-01-01

Epidermal growth factor-like domain multiple 7 (EGFL7), a secreted protein specifically expressed by endothelial cells during embryogenesis, recently was identified as a critical gene in tumor metastasis. Epithelial-mesenchymal transition (EMT) was found to be closely related with tumor progression. Accordingly, it is important to investigate the migration and EMT change after knock-down of EGFL7 gene expression in human pancreatic cancer cells. EGFL7 expression was firstly testified in 4 pancreatic cancer cell lines by real-time polymerase chain reaction (Real-time PCR) and western blot, and the highest expression of EGFL7 was found in PANC-1 cell line. Then, PANC-1 cells transfected with small interference RNA (siRNA) of EGFL7 using plasmid vector were named si-PANC-1, while transfected with negative control plasmid vector were called NC-PANC-1. Transwell assay was used to analyze the migration of PANC-1 cells. Real-time PCR and western blotting were used to detect the expression change of EGFL7 gene, EMT markers like E-Cadherin, N-Cadherin, Vimentin, Fibronectin and transcription factors like snail, slug in PANC-1, NC- PANC-1, and si-PANC-1 cells, respectively. After successful plasmid transfection, EGFL7 gene were dramatically knock-down by RNA interference in si-PANC-1 group. Meanwhile, migration ability decreased significantly, compared with PANC-1 and NC-PANC-1 group. Meanwhile, the expression of epithelial phenotype marker E-Cadherin increased and that of mesenchymal phenotype markers N-Cadherin, Vimentin, Fibronectin dramatically decreased in si-PANC-1 group, indicating a reversion of EMT. Also, transcription factors snail and slug decreased significantly after RNA interference. Current study suggested that highly-expressed EGFL7 promotes migration of PANC-1 cells and acts through transcription factors snail and slug to induce EMT, and further study is needed to confirm this issue.

Cigarette smoke induced urocystic epithelial mesenchymal transition via MAPK pathways.

PubMed

Yu, Dexin; Geng, Hao; Liu, Zhiqi; Zhao, Li; Liang, Zhaofeng; Zhang, Zhiqiang; Xie, Dongdong; Wang, Yi; Zhang, Tao; Min, Jie; Zhong, Caiyun

2017-01-31

Cigarette smoke has been shown to be a major risk factor for bladder cancer. Epithelial-mesenchymal transition (EMT) is a crucial process in cancer development. The role of MAPK pathways in regulating cigarette smoke-triggered urocystic EMT remains to be elucidated. Human normal urothelial cells and BALB/c mice were used as in vitro and in vivo cigarette smoke exposure models. Exposure of human normal urothelial cells to cigarette smoke induced morphological change, enhanced migratory and invasive capacities, reduced epithelial marker expression and increased mesenchymal marker expression, along with the activation of MAPK pathways. Moreover, we revealed that ERK1/2 and p38 inhibitors, but rather JNK inhibitor, effectively attenuated cigarette smoke-induced urocystic EMT. Importantly, the regulatory function of ERK1/2 and p38 pathways in cigarette smoke-triggered urocystic EMT was further confirmed in mice exposed to CS for 12 weeks. These findings could provide new insight into the molecular mechanisms of cigarette smoke-associated bladder cancer development as well as its potential intervention.

Epithelial-to-mesenchymal transition leads to loss of EpCAM and different physical properties in circulating tumor cells from metastatic breast cancer.

PubMed

Hyun, Kyung-A; Koo, Gi-Bang; Han, Hyunju; Sohn, Joohyuk; Choi, Wonshik; Kim, Seung-Il; Jung, Hyo-Il; Kim, You-Sun

2016-04-26

The dissemination of circulating tumor cells (CTCs) requires the Epithelial-to-Mesenchymal transition (EMT), in which cells lose their epithelial characteristics and acquire more mesenchymal-like phenotypes. Current isolation of CTCs relies on affinity-based approaches reliant on the expression of Epithelial Cell Adhesion Molecule (EpCAM). Here we show EMT-induced breast cancer cells maintained in prolonged mammosphere culture conditions possess increased EMT markers and cancer stem cell markers, as well as reduced cell mass and size by quantitative phase microscopy; however, EpCAM expression is dramatically decreased in these cells. Moreover, CTCs isolated from breast cancer patients using a label-free microfluidic flow fractionation device had differing expression patterns of EpCAM, indicating that affinity approaches reliant on EpCAM expression may underestimate CTC number and potentially miss critical subpopulations. Further characterization of CTCs, including low-EpCAM populations, using this technology may improve detection techniques and cancer diagnosis, ultimately improving cancer treatment.

Osteo-/odontogenic differentiation of induced mesenchymal stem cells generated through epithelial-mesenchyme transition of cultured human keratinocytes.

PubMed

Yi, Jin-Kyu; Mehrazarin, Shebli; Oh, Ju-Eun; Bhalla, Anu; Oo, Jenessa; Chen, Wei; Lee, Min; Kim, Reuben H; Shin, Ki-Hyuk; Park, No-Hee; Kang, Mo K

2014-11-01

Revascularization of necrotic pulp has been successful in the resolution of periradicular inflammation; yet, several case studies suggest the need for cell-based therapies using mesenchymal stem cells (MSCs) as an alternative for de novo pulp regeneration. Because the availability of MSCs may be limited, especially in an aged population, the current study reports an alternative approach in generating MSCs from epidermal keratinocytes through a process called epithelial-mesenchymal transition (EMT). We induced EMT in primary normal human epidermal keratinocytes (NHEKs) by transient transfection of small interfering RNA targeting the p63 gene. The resulting cells were assayed for their mesenchymal marker expression, proliferation capacities as a monolayer and in a 3-dimensional collagen scaffold, and differentiation capacities. Transient transfection of p63 small-interfering RNA successfully abolished the expression of endogenous p63 in NHEKs and induced the expression of mesenchymal markers (eg, vimentin and fibronectin), whereas epithelial markers (eg, E-cadherin and involucrin) were lost. The NHEKs exhibiting the EMT phenotype acquired extended replicative potential and an increased telomere length compared with the control cells. Similar to the established MSCs, the NHEKs with p63 knockdown showed attachment onto the 3-dimensional collagen scaffold and underwent progressive proliferation and differentiation. Upon differentiation, these EMT cells expressed alkaline phosphatase activity, osteocalcin, and osteonectin and readily formed mineralized nodules detected by alizarin S red staining, showing osteo-/odontogenic differentiation. The induction of EMT in primary NHEKs by means of transient p63 knockdown allows the generation of induced MSCs from autologous sources. These cells may be used for tissues engineering purposes, including that of dental pulp. Copyright © 2014 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

A Patient-Derived, Pan-Cancer EMT Signature Identifies Global Molecular Alterations and Immune Target Enrichment Following Epithelial-to-Mesenchymal Transition.

PubMed

Mak, Milena P; Tong, Pan; Diao, Lixia; Cardnell, Robert J; Gibbons, Don L; William, William N; Skoulidis, Ferdinandos; Parra, Edwin R; Rodriguez-Canales, Jaime; Wistuba, Ignacio I; Heymach, John V; Weinstein, John N; Coombes, Kevin R; Wang, Jing; Byers, Lauren Averett

2016-02-01

We previously demonstrated the association between epithelial-to-mesenchymal transition (EMT) and drug response in lung cancer using an EMT signature derived in cancer cell lines. Given the contribution of tumor microenvironments to EMT, we extended our investigation of EMT to patient tumors from 11 cancer types to develop a pan-cancer EMT signature. Using the pan-cancer EMT signature, we conducted an integrated, global analysis of genomic and proteomic profiles associated with EMT across 1,934 tumors including breast, lung, colon, ovarian, and bladder cancers. Differences in outcome and in vitro drug response corresponding to expression of the pan-cancer EMT signature were also investigated. Compared with the lung cancer EMT signature, the patient-derived, pan-cancer EMT signature encompasses a set of core EMT genes that correlate even more strongly with known EMT markers across diverse tumor types and identifies differences in drug sensitivity and global molecular alterations at the DNA, RNA, and protein levels. Among those changes associated with EMT, pathway analysis revealed a strong correlation between EMT and immune activation. Further supervised analysis demonstrated high expression of immune checkpoints and other druggable immune targets, such as PD1, PD-L1, CTLA4, OX40L, and PD-L2, in tumors with the most mesenchymal EMT scores. Elevated PD-L1 protein expression in mesenchymal tumors was confirmed by IHC in an independent lung cancer cohort. This new signature provides a novel, patient-based, histology-independent tool for the investigation of EMT and offers insights into potential novel therapeutic targets for mesenchymal tumors, independent of cancer type, including immune checkpoints. ©2015 American Association for Cancer Research.

The Epigenetic Factor KDM2B Regulates EMT and Small GTPases in Colon Tumor Cells.

PubMed

Zacharopoulou, Nefeli; Tsapara, Anna; Kallergi, Galatea; Schmid, Evi; Alkahtani, Saad; Alarifi, Saud; Tsichlis, Philip N; Kampranis, Sotirios C; Stournaras, Christos

2018-05-14

The epigenetic factor KDM2B is a histone demethylase expressed in various tumors. Recently, we have shown that KDM2B regulates actin cytoskeleton organization, small Rho GTPases signaling, cell-cell adhesion and migration of prostate tumor cells. In the present study, we addressed its role in regulating EMT and small GTPases expression in colon tumor cells. We used RT-PCR for the transcriptional analysis of various genes, Western blotting for the assessment of protein expression and immunofluorescence microscopy for visualization of fluorescently labeled proteins. We report here that KDM2B regulates EZH2 and BMI1 in HCT116 colon tumor cells. Knockdown of this epigenetic factor induced potent up-regulation of the protein levels of the epithelial markers E-cadherin and ZO-1, while the mesenchymal marker N-cadherin was downregulated. On the other hand, KDM2B overexpression downregulated the levels of both epithelial markers and upregulated the mesenchymal marker, suggesting control of EMT by KDM2B. In addition, RhoA, RhoB and RhoC protein levels diminished upon KDM2B-knockdown, while all three small GTPases became upregulated in KDM2B-overexpressing HCT116 cell clones. Interestingly, Rac1 GTPase level increased upon KDM2B-knockdown and diminished in KDM2B-overexpressing HCT116 colon tumor- and DU-145 prostate cancer cells. These results establish a clear functional role of the epigenetic factor KDM2B in the regulation of EMT and small-GTPases expression in colon tumor cells and further support the recently postulated oncogenic role of this histone demethylase in various tumors. © 2018 The Author(s). Published by S. Karger AG, Basel.

GSK-3β and vitamin D receptor are involved in β-catenin and snail signaling in high glucose-induced epithelial-mesenchymal transition of mouse podocytes.

PubMed

Guo, Jia; Xia, Nannan; Yang, Lili; Zhou, Sijie; Zhang, Qian; Qiao, Yingjin; Liu, Zhangsuo

2014-01-01

Epithelial-mesenchymal transition (EMT) is recognized to play an important role in diabetic nephropathy (DN). To analyze the roles of glycogen synthase kinase 3β (GSK-3β), β-catenin and Snail signaling in high glucose (HG)-induced mouse podocytes EMT. Differentiated podocytes were divided into: the normal glucose group (NG: glucose 5.6mM), the HG groups (12.5HG: 12.5mM; 25HG: 25mM; and 50HG: 50mM of glucose), and the osmotic control group (NG+M: glucose 5.6mM and mannitol 44.4mM). GSK-3β, β-catenin and Snail were assessed using semi-quantitative RT-PCR, western blot and immunofluorescence. β-catenin and Snail pathways were assessed after down-regulating GSK-3β expression using an inhibitor (LiCl) or a small-interfering RNA (siRNA). HG increased GSK-3β, β-catenin and Snail expressions, and promoted EMT, as shown by decreased nephrin expression (epithelial marker), and increased α-SMA expression (mesenchymal marker). GSK-3β inhibitor and GSK-3β siRNA decreased β-catenin and Snail expressions, and reversed HG-induced EMT. Immunofluorescence showed that GSK-3β and β-catenin did not completely overlap; β-catenin was transferred to the nucleus in the 25HG group. VDR seems to be involved in HG-induced β-catenin nuclear translocation. Down-regulating GSK-3β expression decreased β-catenin and Snail expression and reversed HG-induced podocytes EMT. Thus, modulating GSK-3β might be a target to slow or prevent DN. © 2014 S. Karger AG, Basel. © 2014 S. Karger AG, Basel.

Up-regulation of miR-200 and let-7 by natural agents leads to the reversal of epithelial-mesenchymal transition in gemcitabine-resistant pancreatic cancer cells

PubMed Central

Li, Yiwei; VandenBoom, Timothy G.; Kong, Dejuan; Wang, Zhiwei; Ali, Shadan; Philip, Philip A.; Sarkar, Fazlul H.

2009-01-01

Pancreatic cancer (PC) is the fourth most common cause of cancer death in the United States and the aggressiveness of PC is in part due to its intrinsic and extrinsic drug resistance characteristics, which is also associated with the acquisition of epithelial-to-mesenchymal transition (EMT). Emerging evidence also suggest that the processes of EMT is regulated by the expression status of many microRNAs (miRNAs), which are believed to function as key regulators of various biological and pathological processes during tumor development and progression. In the present study, we compared the expression of miRNAs between gemcitabine-sensitive and gemcitabine-resistant PC cells, and investigated whether the treatment of cells with “natural agents” [3,3′-diinodolylmethane (DIM) or isoflavone] could affect the expression of miRNAs. We found that the expression of miR-200b, miR-200c, let-7b, let-7c, let-7d, and let-7e was significantly down-regulated in gemcitabine-resistant cells that showed EMT characteristics such as elongated fibroblastoid morphology, lower expression of epithelial marker E-cadherin, and higher expression of mesenchymal markers such as vimentin and ZEB1. Moreover, we found that re-expression of miR-200 by transfection studies or treatment of gemcitabine-resistant cells with either DIM or isoflavone resulted in the down-regulation of ZEB1, slug, and vimentin, which was consistent with morphological reversal of EMT phenotype leading to epithelial morphology. These results provide experimental evidence, for the first time, that DIM and isoflavone could function as miRNA regulators leading to the reversal of EMT phenotype, which is likely to be important for designing novel therapies for PC. PMID:19654291

The transcription factor LEF-1 induces an epithelial–mesenchymal transition in MDCK cells independent of β-catenin

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kobayashi, Wakako; Ozawa, Masayuki, E-mail: mozawa@m.kufm.kagoshima-u.ac.jp

2013-12-06

Highlights: •The transcription factor LEF-1 induces an EMT in MDCK cells. •A mutant LEF-1 that cannot interact with β-catenin retained the ability. •The nuclear function of β-catenin was not necessary for the LEF-1-induced EMT. •The mRNA levels of Slug, ZEB1, and ZEB2 increased significantly in these cells. -- Abstract: 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. LEF-1 is a member of the lymphoid enhancer-binding factor/T-cell factor (LEF/TCF) family of DNA-binding transcription factors, which interactmore » with nuclear β-catenin and act as central transcriptional mediators of Wnt signaling. To investigate the role of LEF-1 in EMT, we generated stable LEF-1 transfectants using MDCK cells. The transfectants had a spindle-shaped mesenchymal morphology, and enhanced migration and invasiveness relative to control cells. These EMT changes were accompanied by the downregulation of an epithelial marker protein, E-cadherin, and the upregulation of mesenchymal marker proteins, vimentin and N-cadherin. Consistent with these observations, the mRNA levels of Slug, ZEB1, and ZEB2—EMT-related transcription factors—increased significantly. Although the N-terminally deleted mutant LEF-1 cannot interact with β-catenin, it retained the ability to induce EMT. Consistent with these observations, neither the expression of a dominant negative β-catenin/engrailed chimera, nor the expression of a cytoplasmic domain of E-cadherin that sequesters β-catenin from binding to LEF/TCF, reversed LEF-1-induced EMT. Together, these data indicated that the nuclear function of β-catenin was not necessary for the induction of Slug, ZEB1, and ZEB2 expression leading to EMT.« less

Doxycycline inhibits the cancer stem cell phenotype and epithelial-to-mesenchymal transition in breast cancer.

PubMed

Zhang, Le; Xu, Liang; Zhang, Fengchun; Vlashi, Erina

2017-04-18

Experimental evidence suggest that breast tumors originate from breast cancer stem cells (BCSCs), and that mitochondrial biogenesis is essential for the anchorage-independent clonal expansion and survival of CSCs, thus rendering mitochondria a significant target for novel treatment approaches. One of the recognized side effects of the FDA-approved drug, doxycycline is the inhibition of mitochondrial biogenesis. Here we investigate the mechanism by which doxycycline exerts its inhibitory effects on the properties of breast cancer cells and BCSCs, such as mammosphere forming efficiency, invasion, migration, apoptosis, the expression of stem cell markers and epithelial-to-mesenchymal transition (EMT) related markers of breast cancer cells. In addition, we explored whether autophagy plays a role in the inhibitory effect of doxycycline on breast cancer cells. We find that doxycyline can inhibit the viability and proliferation of breast cancer cells and BCSCs, decrease mammosphere forming efficiency, migration and invasion, and EMT of breast cancer cells. Expression of stem cell factors Oct4, Sox2, Nanog and CD44 were also significantly downregulated after doxycycline treatment. Moreover, doxycycline could down-regulate the expression of the autophagy marker LC-3BI and LC-3BII, suggesting that inhibiting autophagy may be responsible in part for the observed effects on proliferation, EMT and stem cell markers. The potent inhibition of EMT and cancer stem-like characteristics in breast cancer cells by doxycycline treatment suggests that this drug can be repurposed as an anti-cancer drug in the treatment of breast cancer patients in the clinic.

Doxycycline inhibits the cancer stem cell phenotype and epithelial-to-mesenchymal transition in breast cancer

PubMed Central

Xu, Liang; Zhang, Fengchun; Vlashi, Erina

2017-01-01

ABSTRACT Experimental evidence suggest that breast tumors originate from breast cancer stem cells (BCSCs), and that mitochondrial biogenesis is essential for the anchorage-independent clonal expansion and survival of CSCs, thus rendering mitochondria a significant target for novel treatment approaches. One of the recognized side effects of the FDA-approved drug, doxycycline is the inhibition of mitochondrial biogenesis. Here we investigate the mechanism by which doxycycline exerts its inhibitory effects on the properties of breast cancer cells and BCSCs, such as mammosphere forming efficiency, invasion, migration, apoptosis, the expression of stem cell markers and epithelial-to-mesenchymal transition (EMT) related markers of breast cancer cells. In addition, we explored whether autophagy plays a role in the inhibitory effect of doxycycline on breast cancer cells. We find that doxycyline can inhibit the viability and proliferation of breast cancer cells and BCSCs, decrease mammosphere forming efficiency, migration and invasion, and EMT of breast cancer cells. Expression of stem cell factors Oct4, Sox2, Nanog and CD44 were also significantly downregulated after doxycycline treatment. Moreover, doxycycline could down-regulate the expression of the autophagy marker LC-3BI and LC-3BII, suggesting that inhibiting autophagy may be responsible in part for the observed effects on proliferation, EMT and stem cell markers. The potent inhibition of EMT and cancer stem-like characteristics in breast cancer cells by doxycycline treatment suggests that this drug can be repurposed as an anti-cancer drug in the treatment of breast cancer patients in the clinic. PMID:27753527

MDA-9/Syntenin (SDCBP) modulates small GTPases RhoA and Cdc42 via transforming growth factor β1 to enhance epithelial-mesenchymal transition in breast cancer.

PubMed

Menezes, Mitchell E; Shen, Xue-Ning; Das, Swadesh K; Emdad, Luni; Sarkar, Devanand; Fisher, Paul B

2016-12-06

Epithelial-mesenchymal transition (EMT) is one of the decisive steps regulating cancer invasion and metastasis. However, the molecular mechanisms underlying this transition require further clarification. MDA-9/syntenin (SDCBP) expression is elevated in breast cancer patient samples as well as cultured breast cancer cells. Silencing expression of MDA-9 in mesenchymal metastatic breast cancer cells triggered a change in cell morphology in both 2D- and 3D-cultures to a more epithelial-like phenotype, along with changes in EMT markers, cytoskeletal rearrangement and decreased invasion. Conversely, over expressing MDA-9 in epithelial non-metastatic breast cancer cells instigated a change in morphology to a more mesenchymal phenotype with corresponding changes in EMT markers, cytoskeletal rearrangement and an increase in invasion. We also found that MDA-9 upregulated active levels of known modulators of EMT, the small GTPases RhoA and Cdc42, via TGFβ1. Reintroducing TGFβ1 in MDA-9 silenced cells restored active RhoA and cdc42 levels, modulated cytoskeletal rearrangement and increased invasion. We further determined that MDA-9 interacts with TGFβ1 via its PDZ1 domain. Finally, in vivo studies demonstrated that silencing the expression of MDA-9 resulted in decreased lung metastasis and TGFβ1 re-expression partially restored lung metastases. Our findings provide evidence for the relevance of MDA-9 in mediating EMT in breast cancer and support the potential of MDA-9 as a therapeutic target against metastatic disease.

Na, K-ATPase subunits as markers for epithelial-mesenchymal transition in cancer and fibrosis

PubMed Central

Rajasekaran, Sigrid A.; Huynh, Thu P.; Wolle, Daniel G.; Espineda, Cromwell E.; Inge, Landon J.; Skay, Anna; Lassman, Charles; Nicholas, Susanne B.; Harper, Jeffrey F.; Reeves, Anna E.; Ahmed, Mansoor M.; Leatherman, James M; Mullin, James M.; Rajasekaran, Ayyappan K.

2010-01-01

Epithelial-to-mesenchymal transition (EMT) is an important developmental process, participates in tissue repair and occurs during pathological processes of tumor invasiveness, metastasis and tissue fibrosis. The molecular mechanisms leading to EMT are poorly understood. While it is well documented that transforming growth factor (TGF)-β plays a central role in the induction of EMT, the targets of TGF-β signaling are poorly defined. We have shown earlier that Na,K-ATPase β1-subunit levels are highly reduced in poorly differentiated kidney carcinoma cells in culture and in patients’ tumor samples. In this study, we provide evidence that Na,K-ATPase is a new target of TGF-β1-mediated EMT in renal epithelial cells, a model system used in studies of both cancer progression and fibrosis. We show that following treatment with TGF-β1 the surface expression of the β1-subunit of Na,K-ATPase is reduced, prior to well-characterized EMT markers and is associated with the acquisition of a mesenchymal phenotype. RNAi mediated knockdown confirmed the specific involvement of the Na,K-ATPase β1-subunit in the loss of the epithelial phenotype and exogenous over-expression of the Na,K-ATPase β1-subunit attenuated TGF-β1-mediated EMT. We further show that both Na,K-ATPase α- and β-subunit levels are highly reduced in renal fibrotic tissues. These findings for the first time reveal that Na,K-ATPase is a target of TGF-β1-mediated EMT and is associated with the progression of EMT in both cancer and fibrosis. PMID:20501797

Overexpression of Snail in retinal pigment epithelial triggered epithelial–mesenchymal transition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Hui; Li, Min; Xu, Ding

2014-03-28

Highlights: • First reported overexpression of Snail in RPE cells could directly trigger EMT. • Further confirmed the regulator role of Snail in RPE cells EMT in vitro. • Snail may be a potential therapeutic target to prevent the fibrosis of PVR. - Abstract: Snail transcription factor has been implicated as an important regulator in epithelial–mesenchymal transition (EMT) during tumourigenesis and fibrogenesis. Our previous work showed that Snail transcription factor was activated in transforming growth factor β1 (TGF-β1) induced EMT in retinal pigment epithelial (RPE) cells and may contribute to the development of retinal fibrotic disease such as proliferative vitreoretinopathymore » (PVR). However, whether Snail alone has a direct role on retinal pigment epithelial–mesenchymal transition has not been investigated. Here, we analyzed the capacity of Snail to drive EMT in human RPE cells. A vector encoding Snail gene or an empty vector were transfected into human RPE cell lines ARPE-19 respectively. Snail overexpression in ARPE-19 cells resulted in EMT, which was characterized by the expected phenotypic transition from a typical epithelial morphology to mesenchymal spindle-shaped. The expression of epithelial markers E-cadherin and Zona occludin-1 (ZO-1) were down-regulated, whereas mesenchymal markers a-smooth muscle actin (a-SMA) and fibronectin were up-regulated in Snail expression vector transfected cells. In addition, ectopic expression of Snail significantly enhanced ARPE-19 cell motility and migration. The present data suggest that overexpression of Snail in ARPE-19 cells could directly trigger EMT. These results may provide novel insight into understanding the regulator role of Snail in the development of retinal pigment epithelial–mesenchymal transition.« less

Quantitative analysis of injury-induced anterior subcapsular cataract in the mouse: a model of lens epithelial cells proliferation and epithelial-mesenchymal transition.

PubMed

Xiao, Wei; Chen, Xiaoyun; Li, Weihua; Ye, Shaobi; Wang, Wencong; Luo, Lixia; Liu, Yizhi

2015-02-10

The mouse lens capsular injury model has been widely used in investigating the mechanisms of anterior subcapsular cataract (ASC) and posterior capsule opacification (PCO), and evaluating the efficacy of antifibrotic compounds. Nevertheless, there is no available protocol to quantitatively assess the treatment outcomes. Our aim is to describe a new method that can successfully quantify the wound and epithelial-mesenchymal transition (EMT) markers expression in vivo. In this model, lens anterior capsule was punctured with a hypodermic needle, which triggered lens epithelial cells (LECs) proliferation and EMT rapidly. Immunofluorescent staining of injured lens anterior capsule whole-mounts revealed the formation of ASC and high expression of EMT markers in the subcapsular plaques. A series of sectional images of lens capsule were acquired from laser scanning confocal microscopy (LSCM) three-dimensional (3D) scanning. Using LSCM Image Browser software, we can not only obtain high resolution stereo images to present the spatial structures of ASC, but also quantify the subcapsular plaques and EMT markers distribution successfully. Moreover, we also demonstrated that histone deacetylases (HDACs) inhibitor TSA significantly prevented injury-induced ASC using this method. Therefore, the present research provides a useful tool to study ASC and PCO biology as well as the efficacy of new therapies.

Notch-1 induces Epithelial-mesenchymal transition consistent with cancer stem cell phenotype in pancreatic cancer cells

PubMed Central

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

2011-01-01

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. PMID:21463919

Shenqi detoxification granule combined with P311 inhibits epithelial-mesenchymal transition in renal fibrosis via TGF-β1-Smad-ILK pathway.

PubMed

Cai, Pingping; Liu, Xiang; Xu, Yuan; Qi, Fanghua; Si, Guomin

2017-01-01

Shenqi detoxification granule (SDG), a traditional Chinese herbal formula, has been shown to have nephroprotective and anti-fibrotic activities in patients with chronic kidney disease (CKD). However, its mechanisms in renal fibrosis and the progression of CKD remain largely unknown. P311, a highly conserved 8-kDa intracellular protein, plays a key role in renal fibrosis by regulating epithelial-mesenchymal transition (EMT). Previously, we found P311 might be involved in the pathogenesis of renal fibrosis by inhibiting EMT via the TGF-β1-Smad-ILK pathway. We also found SDG combined with P311 could ameliorate renal fibrosis by regulating the expression of EMT markers. Here we further examined the effect and mechanism of SDG combined with P311 on TGF-β1-mediated EMT in a rat model of unilateral ureteral occlusion (UUO) renal fibrosis. After establishment of the UUO model successfully, the rats were gavaged with SDG daily and/or injected with recombinant adenovirus p311 (also called Ad-P311) through the tail vein each week for 4 weeks. Serum creatinine (Cr), blood urea nitrogen (BUN) and albumin (ALB) levels were tested to observe renal function, and hematoxylin eosin (HE) and Masson staining were performed to observe kidney histopathology. Furthermore, the expression of EMT markers (E-cadherin and α-smooth muscle actin (α-SMA)) and EMT-related molecules TGF-β1, pSmad2/3, Smad7 and ILK were observed using immunohistochemical staining and Western blot analysis. Treatment with SDG and P311 improved renal function and histopathological abnormalities, as well as reversing the changes of EMT markers and EMT-related molecules, which indicated SDG combined with P311 could attenuate renal fibrosis in UUO rats, and the underlying mechanism might involve TGF-β1-mediated EMT and the TGF-β1-Smad-ILK signaling pathway. Therefore, SDG might be a novel alternative therapy for treating renal fibrosis and delaying the progression of CKD. Furthermore, SDG combined with P311 might have a synergistic effect on attenuating renal fibrosis.

Specificity protein 1 (Sp1) maintains basal epithelial expression of the miR-200 family: implications for epithelial-mesenchymal transition.

PubMed

Kolesnikoff, Natasha; Attema, Joanne L; Roslan, Suraya; Bert, Andrew G; Schwarz, Quenten P; Gregory, Philip A; Goodall, Gregory J

2014-04-18

Epithelial-mesenchymal transition (EMT) is required for the specification of tissues during embryonic development and is recapitulated during the metastatic progression of tumors. The miR-200 family plays a critical role in enforcing the epithelial state with their expression lost in cells undergoing EMT. EMT can be mediated by activation of the ZEB1 and ZEB2 (ZEB) transcription factors, which repress miR-200 expression via a self-reinforcing double negative feedback loop to promote the mesenchymal state. However, it remains unclear what factors drive and maintain epithelial-specific expression of miR-200 in the absence of EMT-inducing factors. Here, we show that the transcription factor Specificity Protein 1 (Sp1) binds to the miR-200b∼200a∼429 proximal promoter and activates miR-200 expression in epithelial cells. In mesenchymal cells, Sp1 expression is maintained, but its ability to activate the miR-200 promoter is perturbed by ZEB-mediated repression. Reduction of Sp1 expression caused changes in EMT-associated markers in epithelial cells. Furthermore, we observed co-expression of Sp1 and miR-200 during mouse embryonic development wherein miR-200 expression was only lost in regions with high ZEB expression. Together, these findings indicate that miR-200 family members require Sp1 to drive basal expression and to maintain an epithelial state.

The bile acid receptor GPBAR1 (TGR5) is expressed in human gastric cancers and promotes epithelial-mesenchymal transition in gastric cancer cell lines

PubMed Central

Cipriani, Sabrina; Marchianò, Silvia; Marino, Elisabetta; Zampella, Angela; Rende, Mario; Mosci, Paolo; Distrutti, Eleonora; Donini, Annibale; Fiorucci, Stefano

2016-01-01

GPBAR1 (also known as TGR5) is a bile acid activated receptor expressed in several adenocarcinomas and its activation by secondary bile acids increases intestinal cell proliferation. Here, we have examined the expression of GPBAR1 in human gastric adenocarcinomas and investigated whether its activation promotes the acquisition of a pro-metastatic phenotype. By immunohistochemistry and RT-PCR analysis we found that expression of GPBAR1 associates with advanced gastric cancers (Stage III-IV). GPBAR1 expression in tumors correlates with the expression of N-cadherin, a markers of epithelial-mesenchymal transition (EMT) (r=0.52; P<0.01). Expression of GPBAR1, mRNA and protein, was detected in cancer cell lines, with MKN 45 having the higher expression. Exposure of MKN45 cells to GPBAR1 ligands, TLCA, oleanolic acid or 6-ECDCA (a dual FXR and GPBAR1 ligand) increased the expression of genes associated with EMT including KDKN2A, HRAS, IGB3, MMP10 and MMP13 and downregulated the expression of CD44 and FAT1 (P<0.01 versus control cells). GPBAR1 activation in MKN45 cells associated with EGF-R and ERK1 phosphorylation. These effects were inhibited by DFN406, a GPBAR1 antagonist, and cetuximab. GPBAR1 ligands increase MKN45 migration, adhesion to peritoneum and wound healing. Pretreating MKN45 cells with TLCA increased propensity toward peritoneal dissemination in vivo. These effects were abrogated by cetuximab. In summary, we report that GPBAR1 is expressed in advanced gastric cancers and its expression correlates with markers of EMT. GPBAR1 activation in MKN45 cells promotes EMT. These data suggest that GPBAR1 antagonist might have utility in the treatment of gastric cancers. PMID:27409173

EMT blockage strategies: Targeting Akt dependent mechanisms for breast cancer metastatic behaviour modulation.

PubMed

Rafael, D; Doktorovová, S; Florindo, H F; Gener, P; Abasolo, I; Schwartz, S; Videira, M A

2015-01-01

Epithelial Mesenchymal Transition (EMT) is an event where epithelial cells acquire mesenchymal-like phenotype. EMT can occur as a physiological phenomenon during tissue development and wound healing, but most importantly, EMT can confer highly invasive properties to epithelial carcinoma cells. The impairment of E-cadherin expression, an essential cell-cell adhesion protein, together with an increase in the expression of mesenchymal markers, such as N-cadherin, vimentin, and fibronectin, characterize the EMT process and are usually correlated with tumor migration, and metastization. A wide range of micro-environmental and intracellular factors regulate tumor development and progression. The dynamic cross-talk between the adhesion-related proteins such as E-cadherin and the EMT-related transcription factors, with special focus on TWIST, will be discussed here, with the aim of finding a suitable biological pathway to be used as potential target for cancer therapy. Emerging concepts such as the role of the PI3K/AKT/TWIST pathway in the regulation of the E-cadherin expression will be highlighted, since it seems to be consistently involved in cells EMT. The well-known efficacy of the RNA interference as a tool to silence the expression of specific proteins has come into focus as a strategy to control different tumor sub-populations. Despite the oligonucleotides enormous sensitivity and low in vivo stability, new (nano)technological solutions are expected to enable RNAi clinical application in cancer therapy.

Epithelial-to-mesenchymal transition and estrogen receptor α mediated epithelial dedifferentiation mark the development of benign prostatic hyperplasia.

PubMed

Shao, Rui; Shi, Jiandang; Liu, Haitao; Shi, Xiaoyu; Du, Xiaoling; Klocker, Helmut; Lee, Chung; Zhu, Yan; Zhang, Ju

2014-06-01

Epithelial-to-mesenchymal transition (EMT) has been reported involved in the pathogenesis of fibrotic disorders and associated with stemness characteristics. Recent studies demonstrated that human benign prostatic hyperplasia (BPH) development involves accumulation of mesenchymal-like cells derived from the prostatic epithelium. However, the inductive factors of EMT in the adult prostate and the cause-and-effect relationship between EMT and stemness characteristics are not yet resolved. EMT expression patterns were immunohistochemically identified in the human epithelia of normal/BPH prostate tissue and in a rat BPH model induced by estrogen/androgen (E2/T, ratio 1:100) alone or in the presence of the ER antagonist raloxifene. Gene expression profiles were analyzed in micro-dissected prostatic epithelia of rat stimulated by E2/T for 3 days. Two main morphological features both accompanied with EMT were observed in the epithelia of human BPH. Luminal cells undergoing EMT dedifferentiated from a cytokeratin (CK) CK18(+) /CK8(+) /CK19(+) to a CK18(-) /CK8(+) /CK19(-) phenotype and CK14 expression increased in basal epithelial cells. ERα expression was closely related to these dedifferentiated cells and the expression of EMT markers. A similar pattern of EMT events was observed in the E2/T induced rat model of BPH in comparison to the prostates of untreated rats, which could be prevented by raloxifene. Epithelial and mesenchymal phenotype switching is an important mechanism in the etiology of BPH. ERα mediated enhanced estrogenic effect is a crucial inductive factor of epithelial dedifferentiation giving rise to activation of an EMT program in prostate epithelium. © 2014 Wiley Periodicals, Inc.

Induction of hepatic haematopoiesis with fibronectin expression by EMT stromal cells during the second trimester of development.

PubMed

Lambropoulou, M; Tamiolakis, D; Venizelos, I; Alexiadis, G; Anastasopoulos, G; Limberis, V; Galazios, G; Tsikouras, P; Simopoulou, M; Nikolaidou, S; Petrakis, G; Papadopoulos, N

2007-09-01

In an initial period of vertebrate phylogeny (bone marrow-less vertebrates), lymphohaematopoiesis takes place in numerous organs containing a suitable microenvironment. Among other organs (i.e., gonads, kidney and spleen), the liver is apparently the most appropriate site for homing and differentiation of haematopoietic cell precursors. Interaction between haematopoietic cells and stromal cells is important for regulation of haematopoiesis. Numerous soluble and membrane-bound factors directly regulating haematopoiesis have been documented, but little is known about the effect of the foetal hepatic epithelial-to-mesenchymal transition (EMT) stromal cells' activity and their product-fibronectin, on foetal hepatic haematopoiesis. The binding of late-stage erythroid cells to FN has been well characterised and is believed to be critical for the terminal stages of erythroid differentiation. The intention of this article is to provide a quantitative overview of FN, produced by hepatic EMT stromal cells, in foetal hepatic haematopoiesis during the first and second trimester of development. Paraffin-embedded specimens from the liver of 30 human embryos in the first and second trimesters of gestation were investigated by conventional histology and immunohistology for the presence of FN and specific haematopoietic cell types. The staining intensity, and localisation of FN and haematopoietic markers in sequential sections were examined. Furthermore, double immunohistochemical staining was performed to assess simultaneous detection of FN and haematopoietic markers. FN was expressed in the EMT stromal cells of the hepatic portal triads more strongly during the second trimester than the first. Furthermore, an intense immunostaining for haematopoietic lineages, and especially for erythropoiesis, was observed in the second trimester compared to the first. The results of the double immunostaining disclosed an intimate co-expression of the FN and CD haematopoietic markers. Foetal hepatic EMT stromal cells provide a unique microenvironment that supports the emergence, expansion and maintenance of human foetal haematopoietic development during the mid-gestational stage. FN produced by the EMT stromal cells follows a time course parallel to that of haematopoiesis. We suggest that in foetal liver, phenotypic modifications of EMT stromal cells expressing FN concerning the cell adhesion capacity of the protein are associated with proliferation and differentiation of specific haematopoietic cell lineages during the second trimester of gestation, probably reflecting the increasing demand of the growing foetus for mature erythroid and myeloid cells.

Potential Therapeutic Targets of Epithelial-Mesenchymal Transition in Melanoma

PubMed Central

Pearlman, Ross L.; de Oca, Mary Katherine Montes; Pal, Harish Chandra; Afaq, Farrukh

2017-01-01

Melanoma is a cutaneous neoplastic growth of melanocytes with great potential to invade and metastasize, especially when not treated early and effectively. Epithelial-mesenchymal transition (EMT) is the process by which melanocytes lose their epithelial characteristics and acquire mesenchymal phenotypes. Mesenchymal protein expression increases the motility, invasiveness, and metastatic potential of melanoma. Many pathways play a role in promotion of mesenchymal protein expression including RAS/RAF/MEK/ERK, PI3K/AKT/mTOR, Wnt/β-catenin, and several others. Downstream effectors of these pathways induce expression of EMT transcription factors including Snail, Slug, Twist, and Zeb that promote repression of epithelial and induction of mesenchymal character. Emerging research has demonstrated that a variety of small molecule inhibitors as well as phytochemicals can influence the progression of EMT and may even reverse the process, inducing re-expression of epithelial markers. Phytochemicals are of particular interest as supplementary treatment options because of their relatively low toxicities and anti-EMT properties. Modulation of EMT signaling pathways using synthetic small molecules and phytochemicals is a potential therapeutic strategy for reducing the aggressive progression of metastatic melanoma. In this review, we discuss the emerging pathways and transcription factor targets that regulate EMT and evaluate potential synthetic small molecules and naturally occurring compounds that may reduce metastatic melanoma progression. PMID:28131904

TESTIN was commonly hypermethylated and involved in the epithelial-mesenchymal transition of endometrial cancer.

PubMed

Dong, Ruofan; Pu, Hong; Wang, Yuan; Yu, Jinjin; Lian, Kuixian; Mao, Caiping

2015-05-01

We previously reported frequent loss of TESTIN in human endometrial carcinoma, which significantly suppressed tumor proliferation and invasion. Herein, we further explored the mechanisms underlying TESTIN loss and its roles in the epithelial-mesenchymal transition (EMT, a key step for tumor spreading). Methylation-specific PCR was performed to investigate the promoter status of TESTIN in a panel of endometrial cancer and normal endometrium tissues. The expression of TESTIN mRNA was determined by real-time PCR. Up- and down-regulation of TESTIN were achieved by transient transfection with pcDNA3.1-TESTIN and shRNA-TESTIN plasmids, respectively. The EMT alterations were observed under the optical microscope and EMT-related markers were detected by real-time PCR and western blot. Compared to the control (3.6%), TESTIN was hypermethylated in 43.7% endometrial cancer tissues (p < 0.001). Moreover, TESTIN hypermethylation was significantly correlated with advanced tumor stage, deep myometrial invasion and lymphatic node metastasis. In vitro, the demethylating agent dramatically restored the expression of TESTIN. In addition, up-regulation of TESTIN significantly suppressed the EMT procedure; whereas down-regulation of TESTIN enhanced EMT. In conclusion, we demonstrated that loss of TESTIN was mainly caused by hypermethylation, which might be a potent prognostic marker. Furthermore, we proved that TESTIN significantly suppressed the EMT procedure, proposing restoration of TESTIN to be a novel therapeutic strategy for endometrial carcinoma. © 2015 APMIS. Published by John Wiley & Sons Ltd.

Epithelial-mesenchymal transition (EMT) is not sufficient for spontaneous murine breast cancer metastasis.

PubMed

Lou, Yuanmei; Preobrazhenska, Olena; auf dem Keller, Ulrich; Sutcliffe, Margaret; Barclay, Lorena; McDonald, Paul C; Roskelley, Calvin; Overall, Christopher M; Dedhar, Shoukat

2008-10-01

Epithelial-mesenchymal transition (EMT) has been linked to metastatic propensity. The 4T1 tumor is a clinically relevant model of spontaneous breast cancer metastasis. Here we characterize 4T1-derived cell lines for EMT, in vitro invasiveness and in vivo metastatic ability. Contrary to expectations, 67NR cells, which form primary tumors but fail to metastasize, express vimentin and N-cadherin, but not E-cadherin. 4T1 cells express E-cadherin and ZO-1, but are migratory, invasive, and metastasize to multiple sites. 66cl4 cells form lung metastases and display a mixed phenotype, but are not as migratory or invasive as 67NR cells. These findings demonstrate that the metastatic ability of breast cancer cells does not strictly correlate with genotypic and phenotypic properties of EMT per se, and suggest that other processes may govern metastatic capability. Gene expression analysis of primary tumors did not identify differences in EMT markers, but did reveal candidate genes that may influence metastatic ability. Copyright (c) 2008 Wiley-Liss, Inc.

Molecular Detection of EMT Markers in Circulating Tumor Cells from Metastatic Non-Small Cell Lung Cancer Patients: Potential Role in Clinical Practice

PubMed Central

Milano, Annalisa; Mazzetta, Francesca; Valente, Sabatino; Ranieri, Danilo; Leone, Laura; Botticelli, Andrea; Lauro, Salvatore; Torrisi, Maria Rosaria; Marchetti, Paolo

2018-01-01

Background Non-small cell lung cancer (NSCLC) is the most common cause of cancer-related mortality; nevertheless, there are few data regarding detection of circulating tumor cells (CTCs) in NSCLC, compared to other kinds of cancers in which their prognostic roles have already been defined. This difference is likely due to detection methods based on the epithelial marker expression which ignore CTCs undergoing epithelial-mesenchymal transition (CTCsEMT). Methods After optimization of the test with spiking experiments of A549 cells undergoing TGF-β1-induced EMT (A549EMT), the CTCsEMT were enriched by immunomagnetic depletion of leukocytes and then characterized by a RT-PCR assay based on the retrieval of epithelial and EMT-related genes. Blood samples from ten metastatic NSCLC patients before starting treatment and during chemotherapy were used to test this approach by longitudinal monitoring. Ten age- and sex-matched healthy subjects were also enrolled as controls. Results Recovery experiments of spiked A549EMT cells showed that the RT-PCR assay is a reliable method for detection of CTCsEMT. CTCsEMT were detected in three patients at baseline and in six patients after four cycles of cysplatin-based chemotherapy. Longitudinal monitoring of three patients showed that the CTCsEMT detection is related to poor therapeutic response. Conclusions The RT-PCR-based approach for the evaluation of CTCsEMT phenotype could be a promising and inexpensive tool to predict the prognosis and the therapeutic response in NSCLC patients. PMID:29682444

Altered expression of epithelial mesenchymal transition and pluripotent associated markers by sex steroid hormones in human embryonic stem cells.

PubMed

Jeon, So-Ye; Hwang, Kyung-A; Kim, Cho-Won; Jeung, Eui-Bae; Choi, Kyung-Chul

2017-07-01

Embryonic stem (ES) cells are pluripotent stem cells derived from a developmental stage of pre‑implanted embryos. The present study investigated the effect of female sex steroid hormones on the characteristics of human ES cells by using a feeder‑free culture protocol. In a feeder‑free condition without sex hormones, human ES cells assumed the form of tightly packed cells that grow in a monolayer. The cells had clean and defined edges with no evidence of differentiation and expressed several markers specific for undifferentiated ES cells including POU class 5 homeobox 1 (POU5F1), sex determining region Y‑box 2 (SOX2) and NANOG homeobox (NANOG). It was then investigated if female sex steroid hormones including 17β‑estradiol (E2) and progesterone (P4) altered the protein expression of epithelial-mesenchymal transition (EMT) associated markers in addition to pluripotency markers including POU5F1, SOX2 and NANOG in human ES cells. The protein expression levels of N‑cadherin, Snail and Slug were increased while E‑cadherin expression was decreased by treatment of E2 or P4, and the expression levels of POU5F1, SOX2 and NANOG were decreased by the treatment of E2 or P4. When E2 and P4 were treated in combination with an estrogen receptor inhibitor (ICI 182,780) and progesterone receptor inhibitor (RU486) respectively, their effects on EMT and pluripotency of ES cells were restored to control levels. The results suggested that E2 and P4 may regulate EMT and pluripotency of human ES cells by mediating their receptors. The present study may aid in the understanding of the role of sex steroid hormones in the cellular biology of human ES cells.

Menadione (Vitamin K3) induces apoptosis of human oral cancer cells and reduces their metastatic potential by modulating the expression of epithelial to mesenchymal transition markers and inhibiting migration.

PubMed

Suresh, Shruthy; Raghu, Dinesh; Karunagaran, Devarajan

2013-01-01

Oral cancer is one of the most commonly occurring cancers worldwide, decreasing the patient's survival rate due to tumor recurrence and metastasis. Menadione (Vitamin K3) is known to exhibit cytotoxicity in various cancer cells but the present study focused on its effects on viability, apoptosis, epithelial to mesenchymal transition (EMT), anchorage independent growth and migration of oral cancer cells. The results show that menadione is more cytotoxic to SAS (oral squamous carcinoma) cells but not to non-tumorigenic HEK293 and HaCaT cells. Menadione treatment increased the expression of pro-apoptotic proteins, Bax and p53, with a concurrent decrease in anti-apoptotic proteins, Bcl-2 and p65. Menadione induced the expression of E-cadherin but reduced the expression of EMT markers, vimentin and fibronectin. Menadione also inhibited anchorage independent growth and migration in SAS cells. These findings reveal and confirm that menadione is a potential candidate in oral cancer therapy as it exhibits cytotoxic, antineoplastic and antimigratory effects besides effectively blocking EMT in oral cancer cells.

Hepatocyte growth factor upregulation promotes carcinogenesis and epithelial-mesenchymal transition in hepatocellular carcinoma via Akt and COX-2 pathways

PubMed Central

Ogunwobi, Olorunseun O.

2013-01-01

Advanced hepatocellular carcinoma (HCC) is an important cause of cancer mortality. Epithelial-mesenchymal transition (EMT) has been shown to be an important biological process in cancer progression and metastasis. We have focused on elucidating factors that induce EMT to promote carcinogenesis and subsequent metastasis in HCC using the BNL CL.2 (BNL) and BNL 1ME A. 7R.1 (1MEA) cell lines. BNL cells are normal hepatocytes whereas the 1MEA cells are HCC cells derived from chemical transformation of the BNL cells. Their morphological characteristics were examined. Expression levels of hepatocyte growth factor (HGF), markers of EMT and mediators of HGF signaling were determined and functional characteristics were compared. BNL cells were treated with HGF and effects on EMT-marker and mediators of HGF signaling were analyzed. BNL cells display characteristic epithelial morphology whereas 1MEA cells display mesenchymal characteristics. 1MEA cells express and secrete more HGF than BNL cells. There was significantly decreased expression of E-cadherin, albumin, AAT and increased expression of fibronectin, collagen-1, vimentin, snail and slug in 1MEA cells. There was also increased expression of cyclooxygenase-2 (COX-2), Akt and phosphorylated Akt (pAkt) in 1MEA cells. Moreover, 1MEA cells had increased migratory capacity inhibited by inhibition of COX-2 and Akt but not extracellular signal regulated kinase (ERK). Molecular mesenchymal characteristics of 1MEA cells were reversed by inhibition of COX-2, Akt and ERK. Treatment of BNL cells with HGF led to decreased expression of E-cadherin and increased expression of fibronectin, vimentin, snail, slug, COX-2, Akt, pAkt and increased migration, invasiveness and clonogenicity. We conclude that development of HCC is associated with upregulation of HGF which promotes EMT and carcinogenesis via upregulation of COX-2 and Akt. Consequently, HGF signaling may be targeted for therapy in advanced and metastatic HCC. PMID:21744257

Hepatocyte growth factor upregulation promotes carcinogenesis and epithelial-mesenchymal transition in hepatocellular carcinoma via Akt and COX-2 pathways.

PubMed

Ogunwobi, Olorunseun O; Liu, Chen

2011-12-01

Advanced hepatocellular carcinoma (HCC) is an important cause of cancer mortality. Epithelial-mesenchymal transition (EMT) has been shown to be an important biological process in cancer progression and metastasis. We have focused on elucidating factors that induce EMT to promote carcinogenesis and subsequent metastasis in HCC using the BNL CL.2 (BNL) and BNL 1ME A. 7R.1 (1MEA) cell lines. BNL cells are normal hepatocytes whereas the 1MEA cells are HCC cells derived from chemical transformation of the BNL cells. Their morphological characteristics were examined. Expression levels of hepatocyte growth factor (HGF), markers of EMT and mediators of HGF signaling were determined and functional characteristics were compared. BNL cells were treated with HGF and effects on EMT-marker and mediators of HGF signaling were analyzed. BNL cells display characteristic epithelial morphology whereas 1MEA cells display mesenchymal characteristics. 1MEA cells express and secrete more HGF than BNL cells. There was significantly decreased expression of E-cadherin, albumin, AAT and increased expression of fibronectin, collagen-1, vimentin, snail and slug in 1MEA cells. There was also increased expression of cyclooxygenase-2 (COX-2), Akt and phosphorylated Akt (pAkt) in 1MEA cells. Moreover, 1MEA cells had increased migratory capacity inhibited by inhibition of COX-2 and Akt but not extracellular signal regulated kinase (ERK). Molecular mesenchymal characteristics of 1MEA cells were reversed by inhibition of COX-2, Akt and ERK. Treatment of BNL cells with HGF led to decreased expression of E-cadherin and increased expression of fibronectin, vimentin, snail, slug, COX-2, Akt, pAkt and increased migration, invasiveness and clonogenicity. We conclude that development of HCC is associated with upregulation of HGF which promotes EMT and carcinogenesis via upregulation of COX-2 and Akt. Consequently, HGF signaling may be targeted for therapy in advanced and metastatic HCC.

Keratin 8 and 18 Loss in Epithelial Cancer Cells Increases Collective Cell Migration and Cisplatin Sensitivity through Claudin1 Up-regulation*

PubMed Central

Fortier, Anne-Marie; Asselin, Eric; Cadrin, Monique

2013-01-01

Keratins 8 and 18 (K8/18) are simple epithelial cell-specific intermediate filament proteins. Keratins are essential for tissue integrity and are involved in intracellular signaling pathways that regulate cell response to injuries, cell growth, and death. K8/18 expression is maintained during tumorigenesis; hence, they are used as a diagnostic marker in tumor pathology. In recent years, studies have provided evidence that keratins should be considered not only as markers but also as regulators of cancer cell signaling. The loss of K8/18 expression during epithelial-mesenchymal transition (EMT) is associated with metastasis and chemoresistance. In the present study, we investigated whether K8/18 expression plays an active role in EMT. We show that K8/18 stable knockdown using shRNA increased collective migration and invasiveness of epithelial cancer cells without modulating EMT markers. K8/18-depleted cells showed PI3K/Akt/NF-κB hyperactivation and increased MMP2 and MMP9 expression. K8/18 deletion also increased cisplatin-induced apoptosis. Increased Fas receptor membrane targeting suggests that apoptosis is enhanced via the extrinsic pathway. Interestingly, we identified the tight junction protein claudin1 as a regulator of these processes. This is the first indication that modulation of K8/18 expression can influence the phenotype of epithelial cancer cells at a transcriptional level and supports the hypothesis that keratins play an active role in cancer progression. PMID:23449973

Endothelial induced EMT in breast epithelial cells with stem cell properties.

PubMed

Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J R; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A; Petersen, Ole William; Magnusson, Magnus K; Gudjonsson, Thorarinn

2011-01-01

Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M) derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad) to N-Cadherin (N-Cad) and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44(high)/CD24(low) ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer.

Endothelial Induced EMT in Breast Epithelial Cells with Stem Cell Properties

PubMed Central

Sigurdsson, Valgardur; Hilmarsdottir, Bylgja; Sigmundsdottir, Hekla; Fridriksdottir, Agla J. R.; Ringnér, Markus; Villadsen, Rene; Borg, Ake; Agnarsson, Bjarni A.; Petersen, Ole William; Magnusson, Magnus K.; Gudjonsson, Thorarinn

2011-01-01

Epithelial to mesenchymal transition (EMT) is a critical event in cancer progression and is closely linked to the breast epithelial cancer stem cell phenotype. Given the close interaction between the vascular endothelium and cancer cells, especially at the invasive front, we asked whether endothelial cells might play a role in EMT. Using a 3D culture model we demonstrate that endothelial cells are potent inducers of EMT in D492 an immortalized breast epithelial cell line with stem cell properties. Endothelial induced mesenchymal-like cells (D492M) derived from D492, show reduced expression of keratins, a switch from E-Cadherin (E-Cad) to N-Cadherin (N-Cad) and enhanced migration. Acquisition of cancer stem cell associated characteristics like increased CD44high/CD24low ratio, resistance to apoptosis and anchorage independent growth was also seen in D492M cells. Endothelial induced EMT in D492 was partially blocked by inhibition of HGF signaling. Basal-like breast cancer, a vascular rich cancer with stem cell properties and adverse prognosis has been linked with EMT. We immunostained several basal-like breast cancer samples for endothelial and EMT markers. Cancer cells close to the vascular rich areas show no or decreased expression of E-Cad and increased N-Cad expression suggesting EMT. Collectively, we have shown in a 3D culture model that endothelial cells are potent inducers of EMT in breast epithelial cells with stem cell properties. Furthermore, we demonstrate that basal-like breast cancer contains cells with an EMT phenotype, most prominently close to vascular rich areas of these tumors. We conclude that endothelial cells are potent inducers of EMT and may play a role in progression of basal-like breast cancer. PMID:21915264

Epoxyeicosatrienoic Acid Analog Decreases Renal Fibrosis by Reducing Epithelial-to-Mesenchymal Transition

PubMed Central

Skibba, Melissa; Hye Khan, Md. Abdul; Kolb, Lauren L.; Yeboah, Michael M.; Falck, John R.; Amaradhi, Radhika; Imig, John D.

2017-01-01

Renal fibrosis, which is a critical pathophysiological event in chronic kidney diseases, is associated with renal epithelial-to-mesenchymal transition (EMT). Epoxyeicosatrienoic acids (EETs) are Cyp epoxygenase arachidonic acid metabolites that demonstrate biological actions that result in kidney protection. Herein, we investigated the ability of 14,15-EET and its synthetic analog, EET-A, to reduce kidney fibrosis induced by unilateral ureter obstruction (UUO). C57/BL6 male mice underwent sham or UUO surgical procedures and were treated with 14,15-EET or EET-A in osmotic pump (i.p.) for 10 days following UUO surgery. UUO mice demonstrated renal fibrosis with an 80% higher kidney-collagen positive area and 70% higher α-smooth muscle actin (SMA) positive renal areas compared to the sham group. As a measure of collagen content, kidney hydroxyproline content was also higher in UUO (6.4 ± 0.5 μg/10 mg) compared to sham group (2.5 ± 0.1 μg/10 mg). Along with marked renal fibrosis, UUO mice had reduced renal expression of EET producing Cyp epoxygenase enzymes. Endogenous 14,15-EET or EET-A demonstrated anti-fibrotic action in UUO by reducing kidney-collagen positive area (50–60%), hydroxyproline content (50%), and renal α-SMA positive area (85%). In UUO mice, renal expression of EMT inducers, Snail1 and ZEB1 were higher compared to sham group. Accordingly, renal epithelial marker E-cadherin expression was reduced and mesenchymal marker expression was elevated in the UUO compared to sham mice. Interestingly, EET-A reduced EMT in UUO mice by deceasing renal Snail1 and ZEB1 expression. EET-A treatment also opposed the decrease in renal E-cadherin expression and markedly reduced several prominent renal mesenchymal/myofibroblast markers in UUO mice. Overall, our results demonstrate that EET-A is a novel anti-fibrotic agent that reduces renal fibrosis by decreasing renal EMT. PMID:28713267

Glomerular parietal epithelial cells of adult murine kidney undergo EMT to generate cells with traits of renal progenitors

PubMed Central

G, Swetha; Chandra, Vikash; Phadnis, Smruti; Bhonde, Ramesh

2011-01-01

Abstract Glomerular parietal epithelial cells (GPECs) are known to revert to embryonic phenotype in response to renal injury. However, the mechanism of de-differentiation in GPECs and the underlying cellular processes are not fully understood. In the present study, we show that cultured GPECs of adult murine kidney undergo epithelial-mesenchymal transition (EMT) to generate cells, which express CD24, CD44 and CD29 surface antigens. Characterization by qRT-PCR and immunostaining of these clonogenic cells demonstrate that they exhibit metastable phenotype with co-expression of both epithelial (cytokeratin-18) and mesenchymal (vimentin) markers. Transcript analysis by qRT-PCR revealed high expression of metanephric mesenchymal (Pax-2, WT-1, Six-1, Eya-1, GDNF) and uteric bud (Hoxb-7, C-Ret) genes in these cells, indicating their bipotent progenitor status. Incubation of GPECs with EMT blocker Prostaglandin E2, resulted in low expression of renal progenitor markers reflecting the correlation between EMT and acquired stemness in these cells. Additional in vitro renal commitment assays confirmed their functional staminality. When injected into E13.5 kidney rudiments, the cells incorporated into the developing kidney primordia and co-culture with E13.5 spinal cord resulted in branching and tubulogenesis in these cells. When implanted under renal capsule of unilaterally nephrectomized mice, these cells differentiated into immature glomeruli and vascular ducts. Our study demonstrates that EMT plays a major role in imparting plasticity to terminally differentiated GPECs by producing metastable cells with traits of kidney progenitors. The present study would improve our understanding on epithelial cell plasticity, furthering our knowledge of its role in renal repair and regeneration. PMID:19840197

Systematic review of the old and new concepts in the epithelial-mesenchymal transition of colorectal cancer

PubMed Central

Gurzu, Simona; Silveanu, Camelia; Fetyko, Annamaria; Butiurca, Vlad; Kovacs, Zsolt; Jung, Ioan

2016-01-01

Epithelial-to-mesenchymal transition (EMT) is defined as the transformation of an epithelial cell into a spindle cell with the loss of membrane E-cadherin expression and the gain of mesenchymal markers positivity. In the field of colorectal cancer (CRC), first data about EMT was published in 1995 and more than 400 papers had been written up to March 2016. Most of them are focused on the molecular pathways and experimentally-proved chemoresistance. In the present article, an update in the field of EMT in CRC based on the review of the literature and personal experience of the authors is presented. The information about the molecular and immunohistochemical (IHC) particularities of these processes and their possible role in the prognosis of CRC were also up-dated. This article focuses on the IHC quantification of the EMT, the immunoprofile of tumor buds and on the relation between EMT, angiogenesis, and stem cells activation. The EMT-induced chemoresistance vs chemotherapy- or radiotherapy-induced EMT and cellular senescence was also synthesized for both conventional and targeted therapy. As a future perspective, the EMT-angiogenesis-stemness link could be used as a possible valuable parameter for clinical follow-up and targeted therapeutic oncologic management of patients with CRC. Association of dexamethasone and angiotensin converting enzyme inhibitors combined with conventional chemotherapies could have clinical benefits in patients with CRC. The main conclusion is that, although many studies have been published, the EMT features are still incompletely elucidated and newly discovered EMT markers provide confusing data in understanding this complicated process, which might have significant clinical impact. PMID:27570416

YB-1 overexpression promotes a TGF-β1-induced epithelial–mesenchymal transition via Akt activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ha, Bin; Lee, Eun Byul; Cui, Jun

2015-03-06

The Y-box binding protein-1 (YB-1) is a transcription/translation regulatory protein, and the expression thereof is associated with cancer aggressiveness. In the present study, we explored the regulatory effects of YB-1 during the transforming growth factor-β1 (TGF-β1)-induced epithelial-to-mesenchymal transition (EMT) in lung adenocarcinoma cells. Downregulation of YB-1 increased E-cadherin promoter activity, and upregulation of YB-1 decreased promoter activity, suggesting that the YB-1 level may be correlated with the EMT. TGF-β1 induced YB-1 expression, and TGF-β1 translocated cytosolic YB-1 into the nucleus. YB-1 overexpression promoted TGF-β1-induced downregulation of epithelial markers, upregulation of mesenchymal markers, and cell migration. Moreover, YB-1 overexpression enhanced themore » expression of E-cadherin transcriptional repressors via TGF-β1-induced Akt activation. Our findings afford new insights into the role played by YB-1 in the TGF-β1 signaling pathway. - Highlights: • YB-1 regulates E-cadherin expression in A549 cells. • TGF-β1 induces upregulating and nuclear localization of YB-1. • YB-1 overexpression accelerates TGF-β1-induced EMT and cell migration. • YB-1 regulates Snail and Slug expression via Akt activation.« less

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

PubMed

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. Copyright © 2016 Elsevier Ltd. All rights reserved.

Combination Treatment with Apricoxib and IL-27 Enhances Inhibition of Epithelial-Mesenchymal Transition in Human Lung Cancer Cells through a STAT1 Dominant Pathway

PubMed Central

Lee, Mi-Heon; Kachroo, Puja; Pagano, Paul C; Yanagawa, Jane; Wang, Gerald; Walser, Tonya C; Krysan, Kostyantyn; Sharma, Sherven; John, Maie St.; Dubinett, Steven M; Lee, Jay M

2015-01-01

Background The cyclooxygenase 2 (COX-2) pathway has been implicated in the molecular pathogenesis of many malignancies, including lung cancer. Apricoxib, a selective COX-2 inhibitor, has been described to inhibit epithelial-mesenchymal transition (EMT) in human malignancies. The mechanism by which apricoxib may alter the tumor microenvironment by affecting EMT through other important signaling pathways is poorly defined. IL-27 has been shown to have anti-tumor activity and our recent study showed that IL-27 inhibited EMT through a STAT1 dominant pathway. Objective The purpose of this study is to investigate the role of apricoxib combined with IL-27 in inhibiting lung carcinogenesis by modulation of EMT through STAT signaling. Methods and Results Western blot analysis revealed that IL-27 stimulation of human non-small cell lung cancer (NSCLC) cell lines results in STAT1 and STAT3 activation, decreased Snail protein and mesenchymal markers (N-cadherin and vimentin) and a concomitant increase in expression of epithelial markers (E-cadherin, β-and γ-catenins), and inhibition of cell migration. The combination of apricoxib and IL-27 resulted in augmentation of STAT1 activation. However, IL-27 mediated STAT3 activation was decreased by the addition of apricoxib. STAT1 siRNA was used to determine the involvement of STAT1 pathway in the enhanced inhibition of EMT and cell migration by the combined IL-27 and apricoxib treatment. Pretreatment of cells with STAT1 siRNA inhibited the effect of combined IL-27 and apricoxib in the activation of STAT1 and STAT3. In addition, the augmented expression of epithelial markers, decreased expression mesenchymal markers, and inhibited cell migration by the combination treatment were also inhibited by STAT1 siRNA, suggesting that the STAT1 pathway is important in the enhanced effect from the combination treatment. Conclusion Combined apricoxib and IL-27 has an enhanced effect in inhibition of epithelial-mesenchymal transition and cell migration in human lung cancer cells through a STAT1 dominant pathway. PMID:26523208

miR-1271 inhibits migration, invasion and epithelial-mesenchymal transition by targeting ZEB1 and TWIST1 in pancreatic cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Huaize; Wang, Han; Liu, Xiaoxiao

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 whilemore » 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.« less

Prevention of posterior capsular opacification through cyclooxygenase-2 inhibition

PubMed Central

Barden, Curtis A; Lu, Ping; Kusewitt, Donna F.; Colitz, Carmen M. H.

2007-01-01

Purpose To determine if cyclooxygenase-2 (COX-2) is upregulated when lens epithelial cells (LEC) in clinical samples of cataracts and posterior capsule opacification (PCO) undergo epithelial-mesenchymal transition (EMT)-like changes. We also wanted to learn if inhibition of the enzymatic activity of COX-2 could prevent PCO formation. Methods To ensure that EMT-like changes were occurring in LEC, real-time RT-PCR was used to examine expression of EMT markers. Clinical samples of canine cataracts and PCO were examined for COX-2 expression using immunohistochemistry, western blot analysis, and real-time RT-PCR. The COX-2 inhibitors, rofecoxib and celecoxib, were used in an ex vivo model of PCO formation, and the effects on cellular migration, proliferation, and apoptosis were analyzed using immunohistochemistry and western blots. Prostaglandin E2 (PGE2) expression was examined with ELISA. Results Markers of EMT, such as lumican, Snail, Slug, and COX-2 were expressed in LEC. In clinical samples of cataracts and PCO, there was overexpression of COX-2 protein and mRNA. Both rofecoxib and celecoxib were effective at inhibiting PCO formation in our ex vivo model. Prevention of PCO with the COX-2 inhibitors appeared to work through decreased migration and proliferation, and increased apoptosis. Neither of the drugs had a toxic effect on confluent LEC and appeared to inhibit PCO through their pharmacologic action. Synthesis of PGE2 was inhibiting in the capsules treated with the COX-2 inhibiting drugs. Conclusions Extracapsular phacoemulsification cataract surgery is the most common surgical procedure performed in human and veterinary ophthalmology. The most frequent postoperative complication is PCO. The LEC that remain adhered to the lens capsule undergo EMT-like changes, proliferate, and migrate across the posterior lens capsule causing opacities. We have shown that COX-2, a protein associated with EMT, is upregulated in canine cataracts and PCO. Inhibiting the enzymatic activity effectively prevented EMT of LEC in our ex vivo model of PCO through pharmacologic action, and not acute toxicity. These findings indicate that using COX-2 inhibitors in vivo may be an effective technique in preventing PCO. PMID:17563718

Markers of fibrosis and epithelial to mesenchymal transition demonstrate field cancerization in histologically normal tissue adjacent to breast tumors

PubMed Central

Trujillo, Kristina A.; Heaphy, Christopher M.; Mai, Minh; Vargas, Keith M.; Jones, Anna C.; Vo, Phung; Butler, Kimberly S.; Joste, Nancy E.; Bisoffi, Marco; Griffith, Jeffrey K

2011-01-01

Previous studies have shown that a field of genetically altered but histologically normal tissue extends 1 cm or more from the margins of human breast tumors. The extent, composition and biological significance of this field are only partially understood, but the molecular alterations in affected cells could provide mechanisms for limitless replicative capacity, genomic instability and a microenvironment that supports tumor initiation and progression. We demonstrate by microarray, qRT-PCR and immunohistochemistry a signature of differential gene expression that discriminates between patient-matched, tumor-adjacent histologically normal breast tissues located 1 cm and 5 cm from the margins of breast adenocarcinomas (TAHN-1 and TAHN-5, respectively). The signature includes genes involved in extracellular matrix remodeling, wound healing, fibrosis and epithelial to mesenchymal transition (EMT). Myofibroblasts, which are mediators of wound healing and fibrosis, and intra-lobular fibroblasts expressing MMP2, SPARC, TGF-β3, which are inducers of EMT, were both prevalent in TAHN-1 tissues, sparse in TAHN-5 tissues, and absent in normal tissues from reduction mammoplasty. Accordingly, EMT markers S100A4 and vimentin were elevated in both luminal and myoepithelial cells, and EMT markers α-smooth muscle actin and SNAIL were elevated in luminal epithelial cells of TAHN-1 tissues. These results identify cellular processes that are differentially activated between TAHN-1 and TAHN-5 breast tissues, implicate myofibroblasts as likely mediators of these processes, provide evidence that EMT is occurring in histologically normal tissues within the affected field and identify candidate biomarkers to investigate whether or how field cancerization contributes to the development of primary or recurrent breast tumors. PMID:21105047

Novel Omega-3 Fatty Acid Epoxygenase Metabolite Reduces Kidney Fibrosis

PubMed Central

Sharma, Amit; Khan, Md. Abdul Hye; Levick, Scott P.; Lee, Kin Sing Stephen; Hammock, Bruce D.; Imig, John D.

2016-01-01

Cytochrome P450 (CYP) monooxygenases epoxidize the omega-3 polyunsaturated fatty acid (PUFA) docosahexaenoic acid into novel epoxydocosapentaenoic acids (EDPs) that have multiple biological actions. The present study determined the ability of the most abundant EDP regioisomer, 19,20-EDP to reduce kidney injury in an experimental unilateral ureteral obstruction (UUO) renal fibrosis mouse model. Mice with UUO developed kidney tubular injury and interstitial fibrosis. UUO mice had elevated kidney hydroxyproline content and five-times greater collagen positive fibrotic area than sham control mice. 19,20-EDP treatment to UUO mice for 10 days reduced renal fibrosis with a 40%–50% reduction in collagen positive area and hydroxyproline content. There was a six-fold increase in kidney α-smooth muscle actin (α-SMA) positive area in UUO mice compared to sham control mice, and 19,20-EDP treatment to UUO mice decreased α-SMA immunopositive area by 60%. UUO mice demonstrated renal epithelial-to-mesenchymal transition (EMT) with reduced expression of the epithelial marker E-cadherin and elevated expression of multiple mesenchymal markers (FSP-1, α-SMA, and desmin). Interestingly, 19,20-EDP treatment reduced renal EMT in UUO by decreasing mesenchymal and increasing epithelial marker expression. Overall, we demonstrate that a novel omega-3 fatty acid metabolite 19,20-EDP, prevents UUO-induced renal fibrosis in mice by reducing renal EMT. PMID:27213332

Systematic analyses reveal long non-coding RNA (PTAF)-mediated promotion of EMT and invasion-metastasis in serous ovarian cancer.

PubMed

Liang, Haihai; Zhao, Xiaoguang; Wang, Chengyu; Sun, Jian; Chen, Yingzhun; Wang, Guoyuan; Fang, Lei; Yang, Rui; Yu, Mengxue; Gu, Yunyan; Shan, Hongli

2018-06-21

A deeper mechanistic understanding of epithelial-to-mesenchymal transition (EMT) regulation is needed to improve current anti-metastasis strategies in ovarian cancer (OvCa). This study was designed to investigate the role of lncRNAs in EMT regulation during process of invasion-metastasis in serous OvCa to improve current anti-metastasis strategies for OvCa. We systematically analyzes high-throughput gene expression profiles of both lncRNAs and protein-coding genes in OvCa samples with integrated epithelial (iE) subtype and integrated mesenchymal (iM) subtype labels. Mouse models, cytobiology, molecular biology assays and clinical samples were performed to elucidate the function and underlying mechanisms of lncRNA PTAF-mediated promotion of EMT and invasion-metastasis in serous OvCa. We constructed a lncRNA-mediated competing endogenous RNA (ceRNA) regulatory network that affects the expression of many EMT-related protein-coding genes in mesenchymal OvCa. Using a combination of in vitro and in vivo studies, we provided evidence that the lncRNA PTAF-miR-25-SNAI2 axis controlled EMT in OvCa. Our results revealed that up-regulated PTAF induced elevated SNAI2 expression by competitively binding to miR-25, which in turn promoted OvCa cell EMT and invasion. Moreover, we found that silencing of PTAF inhibited tumor progression and metastasis in an orthotopic mouse model of OvCa. We then observed a significant correlation between PTAF expression and EMT markers in OvCa patients. The lncRNA PTAF, a mediator of TGF-β signaling, can predispose OvCa patients to metastases and may serve as a potential target for anti-metastatic therapies for mesenchymal OvCa patients.

A short-term intervention with selenium affects expression of genes implicated in the epithelial-to-mesenchymal transition in the prostate

PubMed Central

Kok, Dieuwertje E.G.; Kiemeney, Lambertus A.L.M.; Verhaegh, Gerald W.; Schalken, Jack A.; van Lin, Emile N.J.T.; Michiel Sedelaar, J.P.; Alfred Witjes, J.; Hulsbergen - van de Kaa, Christina A.; van't Veer, Pieter; Kampman, Ellen; Afman, Lydia A.

2017-01-01

In parallel with the inconsistency in observational studies and chemoprevention trials, the mechanisms by which selenium affects prostate cancer risk have not been elucidated. We conducted a randomized, placebo-controlled trial to examine the effects of a short-term intervention with selenium on gene expression in non-malignant prostate tissue. Twenty-three men received 300 μg selenium per day in the form of selenized yeast (n=12) or a placebo (n=11) during 5 weeks. Prostate biopsies collected from the transition zone before and after intervention were analysed for 15 participants (n=8 selenium, n=7 placebo). Pathway analyses revealed that the intervention with selenium was associated with down-regulated expression of genes involved in cellular migration, invasion, remodeling and immune responses. Specifically, expression of well-established epithelial markers, such as E-cadherin and epithelial cell adhesion molecule EPCAM, was up-regulated, while the mesenchymal markers vimentin and fibronectin were down-regulated after intervention with selenium. This implies an inhibitory effect of selenium on the epithelial-to-mesenchymal transition (EMT). Moreover, selenium was associated with down-regulated expression of genes involved in wound healing and inflammation; processes which are both related to EMT. In conclusion, our explorative data showed that selenium affected expression of genes implicated in EMT in the transition zone of the prostate. PMID:28076331

A short-term intervention with selenium affects expression of genes implicated in the epithelial-to-mesenchymal transition in the prostate.

PubMed

Kok, Dieuwertje E G; Kiemeney, Lambertus A L M; Verhaegh, Gerald W; Schalken, Jack A; van Lin, Emile N J T; Sedelaar, J P Michiel; Witjes, J Alfred; Hulsbergen-van de Kaa, Christina A; van 't Veer, Pieter; Kampman, Ellen; Afman, Lydia A

2017-02-07

In parallel with the inconsistency in observational studies and chemoprevention trials, the mechanisms by which selenium affects prostate cancer risk have not been elucidated. We conducted a randomized, placebo-controlled trial to examine the effects of a short-term intervention with selenium on gene expression in non-malignant prostate tissue. Twenty-three men received 300 µg selenium per day in the form of selenized yeast (n=12) or a placebo (n=11) during 5 weeks. Prostate biopsies collected from the transition zone before and after intervention were analysed for 15 participants (n=8 selenium, n=7 placebo). Pathway analyses revealed that the intervention with selenium was associated with down-regulated expression of genes involved in cellular migration, invasion, remodeling and immune responses. Specifically, expression of well-established epithelial markers, such as E-cadherin and epithelial cell adhesion molecule EPCAM, was up-regulated, while the mesenchymal markers vimentin and fibronectin were down-regulated after intervention with selenium. This implies an inhibitory effect of selenium on the epithelial-to-mesenchymal transition (EMT). Moreover, selenium was associated with down-regulated expression of genes involved in wound healing and inflammation; processes which are both related to EMT. In conclusion, our explorative data showed that selenium affected expression of genes implicated in EMT in the transition zone of the prostate.

EphA2 affects the sensitivity of oxaliplatin by inducing EMT in oxaliplatin-resistant gastric cancer cells.

PubMed

Wen, Qiaocheng; Chen, Zihua; Chen, Zhikang; Chen, Jinxiang; Wang, Ran; Huang, Changhao; Yuan, Weijie

2017-07-18

Erythropoietin-producing hepatocellular receptor A2 (EphA2) is upregulated in gastric cancer tissues and cells, which is accompanied by epithelial-mesenchymal transition (EMT). The current study was designed to establish the oxaliplatin-resistant human gastric cancer cell line SGC-7901/L-OHP, to determine if EMT in these cells could be reversed, and to determine if the susceptibility of these cells to oxaliplatin was affected by silencing EphA2 expression. We found that EphA2 expression levels were upregulated in gastric cancer and associated with chemotherapy sensitivity. EphA2 and the EMT molecular markers N-cadherin and Snail were upregulated in SGC-7901/L-OHP cells, while silencing of EphA2 using small interfering RNA had the opposite effect. Moreover, silencing of EphA2 inhibited cell migration and invasion, and significantly enhanced the sensitivity of oxaliplatin-resistant gastric cancer cells to oxaliplatin. These observations demonstrate that EphA2 affects the sensitivity to oxaliplatin by inducing EMT in oxaliplatin-resistant gastric cancer cells.

PSC-derived Galectin-1 inducing epithelial-mesenchymal transition of pancreatic ductal adenocarcinoma cells by activating the NF-κB pathway

PubMed Central

Tang, Dong; Zhang, Jingqiu; Yuan, Zhongxu; Zhang, Hongpeng; Chong, Yang; Huang, Yuqin; Wang, Jie; Xiong, Qingquan; Wang, Sen; Wu, Qi; Tian, Ying; Lu, Yongdie; Ge, Xiao; Shen, Wenjing; Wang, Daorong

2017-01-01

Galectin-1 has previously been shown to be strongly expressed in activated pancreatic stellate cells (PSCs) and promote the development and metastasis of pancreatic ductal adenocarcinoma (PDAC). However, the molecular mechanisms by which Galectin-1 promotes the malignant behavior of pancreatic cancer cells remain unclear. In this study, we examined the effects of Galectin-1 knockdown or overexpression in PSCs co-cultured with pancreatic cancer (PANC-1) cells. Immunohistochemical analysis showed expression of epithelial-mesenchymal transition (EMT) markers and MMP9 were positively associated with the expression of Galectin-1 in 66 human PDAC tissues. In addition, our in vitro studies showed PSC-derived Galectin-1 promoted the proliferation, invasion, and survival (anti-apoptotic effects) of PANC-1 cells. We also showed PSC-derived Galectin-1 induced EMT of PANC-1 cells and activated the NF-кB pathway in vitro. Our mixed (PSCs and PANC-1 cells) mouse orthotopic xenograft model indicated that overexpression of Galectin-1 in PSCs significantly promoted the proliferation, growth, invasion, and liver metastasis of the transplanted tumor. Moreover, Galectin-1 overexpression in PSCs was strongly associated with increased expression of EMT markers in both the orthotopic xenograft tumor in the pancreas and in metastatic lesions of naked mice. We conclude that PSC-derived Galectin-1 promotes the malignant behavior of PDAC by inducing EMT via activation of the NF-κB pathway. Our results suggest that targeting Galectin-1 in PSCs could represent a promising therapeutic strategy for PDAC progression and metastasis. PMID:29156810

Vasohibin 2 promotes epithelial-mesenchymal transition in human breast cancer via activation of transforming growth factor β 1 and hypoxia dependent repression of GATA-binding factor 3.

PubMed

Tu, Min; Li, Zhanjun; Liu, Xian; Lv, Nan; Xi, Chunhua; Lu, Zipeng; Wei, Jishu; Song, Guoxin; Chen, Jianmin; Guo, Feng; Jiang, Kuirong; Wang, Shui; Gao, Wentao; Miao, Yi

2017-03-01

Vasohibin 2 (VASH2) is identified as an angiogenic factor, and has been implicated in tumor angiogenesis, proliferation and epithelial-mesenchymal transition (EMT). To investigate the EMT role of VASH2 in breast cancer, we overexpressed or knocked down expression of VASH2 in human breast cancer cell lines. We observed that VASH2 induced EMT in vitro and in vivo. The transforming growth factor β1 (TGFβ1) pathway was activated by VASH2, and expression of a dominant negative TGFβ type II receptor could block VASH2-mediated EMT. In clinical breast cancer tissues VASH2 positively correlated with TGFβ1 expression, but negatively correlated with E-cadherin (a marker of EMT) expression. Under hypoxic conditions in vitro or in vivo, we found that down-regulation of estrogen receptor 1 (ESR1) in VASH2 overexpressing ESR1 positive cells suppressed E-cadherin. Correlation coefficient analysis indicated that VASH2 and ESR1 expression were negatively correlated in clinical human breast cancer tissues. Further study revealed that a transcription factor of ESR1, GATA-binding factor 3 (GATA3), was down-regulated by VASH2 under hypoxia or in vivo. These findings suggest that VASH2 drives breast cancer cells to undergo EMT by activation of the TGFβ1 pathway and hypoxia dependent repression GATA3-ESR1 pathway, leading to cancer metastasis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

PD-L1 Expression Promotes Epithelial to Mesenchymal Transition in Human Esophageal Cancer.

PubMed

Chen, Lujun; Xiong, Yuqi; Li, Jing; Zheng, Xiao; Zhou, Qi; Turner, Abbey; Wu, Changping; Lu, Binfeng; Jiang, Jingting

2017-01-01

PD-L1 (Programmed cell death 1 ligand 1, PD-L1), an essential immune checkpoint molecule in the tumor microenvironment, is an important target for cancer immunotherapy. We have previously reported that its expression in human gastric and esophageal cancer tissues is significantly associated with cancer progression and patients' postoperative prognoses. Its expression in cancer cells is well known to inhibit the T cell-mediated anti-tumor response, and this mechanism of action has been targeted for cancer immunotherapy. As of now, the autonomous effect of PD-L1 on cancer cells is not well understood, thus our present study aimed to examine the role of PD-L1 intervention in cellular biological functions, especially epithelial to mesenchymal transition (EMT), of the human esophageal cancer cell line, Eca-109 cells. Immunohistochemistry assay was used to investigate the correlation between expression of PD-L1 and EMT markers in human esophageal cancer tissues. Intervention of PD-L1 by using RNAi and over-expression methods were used to study the role of PD-L1 in regulation of biological behaviors and EMT in Eca-109 cells. Our clinical and pathological data demonstrated that tumor samples in the EMT positive subgroup had higher PD-L1 expression than those in the EMT negative subgroup. By manipulating PD-L1 expression in Eca-109 cells either through ablation or overexpression of wild type and the cytoplasmic domain-truncated mutant, we demonstrated that PD-L1 expression significantly promoted the cell viability, migration and EMT phenotype. Furthermore, our study also indicated that PD-1 fusion protein mediated stimulation of PD-L1 and the cytoplasmic domain of PD-L1 played a critical role in promoting EMT phenotype of Eca-109 cells, thereby suggesting that PD-1 receptor usually by triggering the reverse signaling can effect PD-L1 mediated regulation of esophageal cancer cell response. Our present study reveals a tumor cell-autonomous role of PD-L1 signaling in promoting EMT in human esophageal cancer. © 2017 The Author(s). Published by S. Karger AG, Basel.

Rapamycin protects against paraquat-induced pulmonary fibrosis: Activation of Nrf2 signaling pathway.

PubMed

Xu, Yiheng; Tai, Wenlin; Qu, Xiaoyuan; Wu, Wenjuan; Li, ZhenKun; Deng, Shuhao; Vongphouttha, Chanthasone; Dong, Zhaoxing

2017-08-19

Paraquat (PQ) is a widely used herbicide indeveloping countries worldwide, and pulmonary fibrosis is one of the most typical features of PQ poisoning. The molecular mechanism of PQ toxicity especially how to treat PQ-induced pulmonary fibrosis is still largely unknown. In animal model of pulmonary fibrosis, we used HE staining, western blotting assay and Real-time PCR assay to analyze the effects of rapamycin on the PQ-induced epithelial mesenchymal transition (EMT). We found that PQ induced the pulmonary fibrosis using HE staining and Masson's staining, and up-regulated the activity of HYP and the mRNA expressions of Collagen I and III (COL-1and COL-3) in pulmonary tissues. We also found that rapamycin down-regulated the mesenchymal cell marker Vimentin and up-regulated the epithelial cell marker E-cadherin both in mRNA and protein levels compared with PQ group. And the EMT associated transcription factor Snail was decreased by rapamycin treatment compared with PQ group. And PQ decreased the Nrf2 expression both in mRNA and protein levels, and rapamycin inhibited these effects of PQ. SFN, a activator of Nrf2, could inhibit the EMT and the expression of Snail. And knockdowon of Nrf2 could abolish the inhibitory effects of rapamycin of PQ-induced EMT. In conclusion, rapamycin protects against paraquat-induced pulmonary fibrosis by activation of Nrf2 signaling pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

Apelin attenuates TGF-β1-induced epithelial to mesenchymal transition via activation of PKC-ε in human renal tubular epithelial cells.

PubMed

Wang, Li-Yan; Diao, Zong-Li; Zheng, Jun-Fang; Wu, Yi-Ru; Zhang, Qi-Dong; Liu, Wen-Hu

2017-10-01

Epithelial to mesenchymal transition (EMT), a process whereby fully differentiated epithelial cells transition to a mesenchymal phenotype, has been implicated in the pathogenesis of renal fibrosis. Apelin, a bioactive peptide, has recently been recognized to protect against renal profibrotic activity, but the underlying mechanism has not yet been elucidated. In this study, we investigated the regulation of EMT in the presence of apelin-13 in vitro. Expression of the mesenchymal marker alpha-smooth muscle actin (α-SMA) and the epithelial marker E-cadherin was examined by immunofluorescence and western blotting in transforming growth factor beta 1 (TGF-β1)-stimulated human proximal tubular epithelial cells. Expression of extracellular matrix, fibronectin and collagen-I was examined by quantitative real-time PCR and ELISA. F13A, an antagonist of the apelin receptor APJ, and small interfering RNA targeting protein kinase C epsilon (PKC-ε) were used to explore the relevant signaling pathways. Apelin attenuated TGF-β1-induced EMT, and inhibited the EMT-associated increase in α-SMA, loss of E-cadherin, and secretion of extracellular matrix. Moreover, apelin activated PKC-ε in tubular epithelial cells, which in turn decreased phospho-Smad2/3 levels and increased Smad-7 levels. APJ inhibition or PKC-ε deletion diminished apelin-induced modulation of Smad signaling and suppression of tubular EMT. Our findings identify a novel PKC-ε-dependent mechanism in which apelin suppresses TGF-β1-mediated activation of Smad signaling pathways and thereby inhibits tubular EMT. These results suggest that apelin may be a new agent that can suppress renal fibrosis and retard chronic kidney disease progression. Copyright © 2017 Elsevier Inc. All rights reserved.

S1P1 receptor inhibits kidney epithelial mesenchymal transition triggered by ischemia/reperfusion injury via the PI3K/Akt pathway.

PubMed

Wang, Weina; Wang, Aimei; Luo, Guochang; Ma, Fengqiao; Wei, Xiaoming; Bi, Yongyi

2018-06-13

Ischemia/reperfusion (I/R) is a major cause of acute kidney injury (AKI), along with delayed graft function, which can trigger chronic kidney injury by stimulating epithelial to mesenchymal transition (EMT) in the kidney canaliculus. Sphingosine 1-phosphate receptor 1 (S1P1) is a G protein-coupled receptor that is indispensable for vessel homeostasis. This study aimed to investigate the influence of S1P1 on the mechanisms underlying I/R-induced EMT in the kidney using in vivo and in vitro models. Wild-type (WT) and S1P1-overexpressing kidney canaliculus cells were subject to hypoxic conditions followed by reoxygenation in the presence or absence of FTY720-P, a potent S1P1 agonist. In vivo, bilateral arteria renalis in wild-type mice and mice with silenced S1P1 were clamped for 30 min to obtain I/R models. We found that hypoxia/reoxygenation (H/R) significantly enhanced the expressions of EMT biomarkers and down-regulated S1P1 expression in wild-type canaliculus cells. In contrast, FTY720-P treatment or overexpression of S1P1 significantly suppressed EMT in wild-type canaliculus cells. Furthermore, after 48-72 h, a significant upregulation of EMT biomarker expression was triggered by I/R in mice with silenced S1P1, while the expressions of these markers did not change in wild-type mice. A kt activity was increased with H/R-induced EMT, suggesting that the protective influence of FTY720-P was due to its inhibition of PI3K/Akt. Therefore, the results of this study provide evidence that down-regulation of S1P1 expression is essential for the generation and progression of EMT triggered by I/R. S1P1 exhibits a prominent inhibitory effect on kidney I/R-induced EMT in the kidney by affecting the PI3K/Akt pathway.

Persistent Transmissible Gastroenteritis Virus Infection Enhances Enterotoxigenic Escherichia coli K88 Adhesion by Promoting Epithelial-Mesenchymal Transition in Intestinal Epithelial Cells.

PubMed

Xia, Lu; Dai, Lei; Yu, Qinghua; Yang, Qian

2017-11-01

Transmissible gastroenteritis virus (TGEV) is a coronavirus characterized by diarrhea and high morbidity rates, and the mortality rate is 100% in piglets less than 2 weeks old. Pigs infected with TGEV often suffer secondary infection by other pathogens, which aggravates the severity of diarrhea, but the mechanisms remain unknown. Here, we hypothesized that persistent TGEV infection stimulates the epithelial-mesenchymal transition (EMT), and thus enterotoxigenic Escherichia coli (ETEC) can more easily adhere to generating cells. Intestinal epithelial cells are the primary targets of TGEV and ETEC infections. We found that TGEV can persistently infect porcine intestinal columnar epithelial cells (IPEC-J2) and cause EMT, consistent with multiple changes in key cell characteristics. Infected cells display fibroblast-like shapes; exhibit increases in levels of mesenchymal markers with a corresponding loss of epithelial markers; have enhanced expression levels of interleukin-1β (IL-1β), IL-6, IL-8, transforming growth factor β (TGF-β), and tumor necrosis factor alpha (TNF-α) mRNAs; and demonstrate increases in migratory and invasive behaviors. Additional experiments showed that the activation of the phosphatidylinositol 3-kinase (PI3K)/Akt and extracellular signal-regulated kinase (ERK) signaling pathways via TGF-β is critical for the TGEV-mediated EMT process. Cellular uptake is also modified in cells that have undergone EMT. TGEV-infected cells have higher levels of integrin α5 and fibronectin and exhibit enhanced ETEC K88 adhesion. Reversal of EMT reduces ETEC K88 adhesion and inhibits the expression of integrin α5 and fibronectin. Overall, these results suggest that TGEV infection induces EMT in IPEC-J2 cells, increasing the adhesion of ETEC K88 in the intestine and facilitating dual infection. IMPORTANCE Transmissible gastroenteritis virus (TGEV) causes pig diarrhea and is often followed by secondary infection by other pathogens. In this study, we showed that persistent TGEV infection induces an EMT in porcine intestinal columnar epithelial cells (IPEC-J2) and enhances the adhesion of the secondary pathogen ETEC K88. Additional experiments suggest that integrin α5 and fibronectin play an important role in TGEV-enhanced ETEC K88 adhesion. Reversal of EMT reduces the expression of integrin α5 and fibronectin and also reduces ETEC K88 adhesion. We conclude that TGEV infection triggers EMT and facilitates dual infection. Our results provide new insights into secondary infection and suggest that targeted anti-EMT therapy may have implications for the prevention and treatment of secondary infection. Copyright © 2017 American Society for Microbiology.

Fractionated irradiation-induced EMT-like phenotype conferred radioresistance in esophageal squamous cell carcinoma

PubMed Central

Zhang, Hongfang; Luo, Honglei; Jiang, Zhenzhen; Yue, Jing; Hou, Qiang; Xie, Ruifei; Wu, Shixiu

2016-01-01

The efficacy of radiotherapy, one major treatment modality for esophageal squamous cell carcinoma (ESCC) is severely attenuated by radioresistance. Epithelial-to-mesenchymal transition (EMT) is a cellular process that determines therapy response and tumor progression. However, whether EMT is induced by ionizing radiation and involved in tumor radioresistance has been less studied in ESCC. Using multiple fractionated irradiation, the radioresistant esophageal squamous cancer cell line KYSE-150R had been established from its parental cell line KYSE-150. We found KYSE-150R displayed a significant EMT phenotype with an elongated spindle shape and down-regulated epithelial marker E-cadherin and up-regulated mesenchymal marker N-cadherin in comparison with KYSE-150. Furthermore, KYSE-150R also possessed some stemness-like properties characterized by density-dependent growth promotion and strong capability for sphere formation and tumorigenesis in NOD-SCID mice. Mechanical studies have revealed that WISP1, a secreted matricellular protein, is highly expressed in KYSE-150R and mediates EMT-associated radioresistance both in ESCC cells and in xenograft tumor models. Moreover, WISP1 has been demonstrated to be closely associated with the EMT phenotype observed in ESCC patients and to be an independent prognosis factor of ESCC patients treated with radiotherapy. Our study highlighted WISP1 as an attractive target to reverse EMT-associated radioresistance in ESCC and can be used as an independent prognostic factor of patients treated with radiotherapy. PMID:27125498

Epithelial-mesenchymal transition in colonies of rhesus monkey embryonic stem cells: a model for processes involved in gastrulation.

PubMed

Behr, Rüdiger; Heneweer, Carola; Viebahn, Christoph; Denker, Hans-Werner; Thie, Michael

2005-01-01

Rhesus monkey embryonic stem (rhES) cells were grown on mouse embryonic fibroblast (MEF) feeder layers for up to 10 days to form multilayered colonies. Within this period, stem cell colonies differentiated transiently into complex structures with a disc-like morphology. These complex colonies were characterized by morphology, immunohistochemistry, and marker mRNA expression to identify processes of epithelialization as well as epithelial-mesenchymal transition (EMT) and pattern formation. Typically, differentiated colonies were comprised of an upper and a lower ES cell layer, the former growing on top of the layer of MEF cells whereas the lower ES cell layer spread out underneath the MEF cells. Interestingly, in the central part of the colonies, a roundish pit developed. Here the feeder layer disappeared, and upper layer cells seemed to ingress and migrate through the pit downward to form the lower layer while undergoing a transition from the epithelial to the mesenchymal phenotype, which was indicated by the loss of the marker proteins E-cadherin and ZO-1 in the lower layer. In support of this, we found a concomitant 10-fold upregulation of the gene Snail2, which is a key regulator of the EMT process. Conversion of epiblast to mesoderm was also indicated by the regulated expression of the mesoderm marker Brachyury. An EMT is a characteristic process of vertebrate gastrulation. Thus, these rhES cell colonies may be an interesting model for studies on some basic processes involved in early primate embryogenesis and may open new ways to study the regulation of EMT in vitro.

Mesenchymal-Epithelial Transition and Circulating Tumor Cells in Small Cell Lung Cancer.

PubMed

Hamilton, Gerhard; Rath, Barbara

2017-01-01

Cancer patients die of metastatic disease but knowledge regarding individual steps of this complex process of intravasation, spread and extravasation leading to secondary lesions is incomplete. Subpopulations of tumor cells are supposed to undergo an epithelial-mesenchymal transition (EMT), to enter the bloodstream and eventually establish metastases in a reverse process termed mesenchymal-epithelial transition (MET). Small cell lung cancer (SCLC) represents a unique model to study metastatic spread due to early dissemination and relapse, as well as availability of a panel of circulating cancer cell (CTC) lines recently. Additionally, chemosensitive SCLC tumor cells switch to a completely resistant phenotype during cancer recurrence. In advanced disease, SCLC patients display extremely high blood counts of CTCs in contrast to other tumors, like breast, prostate and colon cancer. Local inflammatory conditions at the primary tumor site and recruitment of macrophages seem to increase the shedding of tumor cells into the circulation in processes which may proceed independently of EMT. Since millions of cells are released by tumors into the circulation per day, analysis of a limited number of CTCs at specific time points are difficult to be related to the development of metastatic lesions which may occur approximately one year later. We have obtained a panel of SCLC CTC cell line from patients with relapsing disease, which share characteristic markers of this malignancy and a primarily epithelial phenotype with unique formation of large tumorospheres, containing quiescent and hypoxic cells. Although smoking and inflammation promote EMT, partial expression of vimentin indicates a transitional state with partial EMT in these cell lines at most. The CTC lines exhibit high expression of EpCAM , absent phosphorylation of β-catenin and background levels of Snail. Provided that these tumor cells had ever undergone EMT, here in advanced disease MET seem to have occurred already in the peripheral circulation. Alternative explanations for the expression of mesenchymal markers of the CTC lines are the heterogeneity of SCLC cells, cooperative migration or altered gene expression in response to the inflammatory tumor microenvironment allowing for tumor spread without EMT/MET.

The invasive phenotype of placenta accreta extravillous trophoblasts associates with loss of E-cadherin.

PubMed

Duzyj, C M; Buhimschi, I A; Motawea, H; Laky, C A; Cozzini, G; Zhao, G; Funai, E F; Buhimschi, C S

2015-06-01

Epithelial-to-mesenchymal transition (EMT) is a process of molecular and phenotypic epithelial cell alteration promoting invasiveness. Loss of E-cadherin (E-CAD), a transmembrane protein involved in cell adhesion, is a marker of EMT. Proteolysis into N- and C-terminus fragments by ADAM10 and presenilin-1 (PSEN-1) generates soluble (sE-CAD) and transcriptionally active forms. We studied the protein expression patterns of E-CAD in the serum and placenta of women with histologically-confirmed over-invasive placentation. The patterns of expression and levels of sE-CAD were analyzed by Western blot, immunoassay, and immunoprecipitation. Tissue immunostaining for E-CAD, cytokeratin-7 (epithelial marker), vimentin (mesenchymal marker), ADAM10, PSEN-1 and β-catenin expression were investigated in parallel. N-terminus cleaved 80 kDa sE-CAD fragments were present in serum of pregnant women with gestational age regulation of the circulatory levels. Women with advanced trophoblast invasion did not display circulatory levels of sE-CAD different from those of women with normal placentation. Histologically, extravillous trophoblasts (EVT) closer to the placental-myometrial interface demonstrated less E-CAD staining than those found deeper in the myometrium. These cells expressed both vimentin and cytokeratin, an additional feature of EMT. EVT of placentas with advanced invasion displayed intracellular E-CAD C-terminus immunoreactivity predominating over that of the extracellular N-terminus, a pattern consistent with preferential PSEN-1 processing. Local processing of E-CAD may be an important molecular mechanism controlling the invasive phenotype of accreta EVT. Copyright © 2015 Elsevier Ltd. All rights reserved.

Simultaneous stimulation with tumor necrosis factor-α and transforming growth factor-β1 induces epithelial-mesenchymal transition in colon cancer cells via the NF-κB pathway.

PubMed

Li, Yuanfei; Zhu, Guoqiang; Zhai, Huihong; Jia, Junmei; Yang, Wenhui; Li, Xiaoqing; Liu, Lixin

2018-05-01

Epithelial-mesenchymal transition (EMT) is critical in the progression of numerous types of carcinoma, and endows invasive and metastatic properties upon cancer cells. The tumor microenvironment facilitates tumor metastasis to distant organs. Various signaling pathways contribute to this process. In the present study, SW480 colon adenocarcinoma cells were treated with transforming growth factor-β1 (TGF-β1; 10 ng/ml) and tumor necrosis factor-α (TNF-α; 20 ng/ml), alone or in combination, for 72 h, and EMT was assessed using immunofluorescence, western blot analysis and migration assays. The functions of p38 mitogen-activated protein kinase, extracellular signal-regulated kinase (ERK) and nuclear factor-κB (NF-κB) pathways in EMT were examined. It was demonstrated that the cooperation of TGF-β1 and TNF-α signaling promoted the morphological conversion of the SW480 cells from an epithelial to a mesenchymal phenotype. Furthermore, simultaneous exposure to TNF-α and TGF-β1 downregulated the expression of E-cadherin (an epithelial marker) and increased the expression of N-cadherin and vimentin (mesenchymal markers). Additionally, the migratory capacity of the SW480 cells increased. The inhibition of p38 and ERK signaling exhibited no effect on EMT, whereas the inhibition of inhibitor of NF-κB kinase subunit β blocked the EMT induced by TGF-β1 and TNF-α. In conclusion, the results of the present study demonstrated that TNF-α and TGF-β1 synergistically promoted EMT in SW480 cells via the NF-κB pathway, independent of p38 activation and ERK1/2 signaling. These results suggest a novel function of TGF-β1 and TNF-α during EMT in colon carcinoma and, thus, provide insights into potential therapeutic interventions.

High LET Radiation Can Enhance TGF(Beta) Induced EMT and Cross-Talk with ATM Pathways

NASA Technical Reports Server (NTRS)

Wang, Minli; Hada, Megumi; Huff, Janice; Pluth, Janice M.; Anderson, Janniffer; ONeill, Peter; Cucinotta, Francis A.

2010-01-01

The TGF(Beta) pathway has been shown to regulate or directly interact with the ATM pathway in the response to radiation in mammary epithelial cells. We investigated possible interactions between the TGF(Beta) and ATM pathways following simulated space radiation using hTERT immortalized human esophageal epithelial cells (EPC-hTERT), mink lung epithelial cells (Mv1lu), and several human fibroblast cell lines. TGF(Beta) is a key modulator of the Epithelial-Mesenchymal Transition (EMT), important in cancer progression and metastasis. The implication of EMT by radiation also has several lines of developing evidence, however is poorly understood. The identification of TGF(Beta) induced EMT can be shown in changes to morphology, related gene over expression or down regulation, which can be detected by RT-PCR, and immunostaining and western blotting. In this study, we have observed morphologic and molecular alternations consistent with EMT after Mv1lu cells were treated with TGF(Beta) High LET radiation enhanced TGF(Beta) mediated EMT with a dose as low as 0.1Gy. In order to consider the TGF(Beta) interaction with ATM we used a potent ATM inhibitor Ku55933 and investigated gene expression changes and Smad signaling kinetics. Ku559933 was observed to reverse TGF(Beta) induced EMT, while this was not observed in dual treated cells (radiation+TGF(Beta)). In EPC-hTERT cells, TGF(Beta) alone was not able to induce EMT after 3 days of application. A combined treatment with high LET, however, significantly caused the alteration of EMT markers. To study the function of p53 in the process of EMT, we knocked down P53 through RNA interference. Morphology changes associated with EMT were observed in epithelial cells with silenced p53. Our study indicates: high LET radiation can enhance TGF(Beta) induced EMT; while ATM is triggering the process of TGF(Beta)-induced EMT, p53 might be an essential repressor for EMT phenotypes.

Platelets enhance tissue factor protein and metastasis initiating cell markers, and act as chemoattractants increasing the migration of ovarian cancer cells.

PubMed

Orellana, Renan; Kato, Sumie; Erices, Rafaela; Bravo, María Loreto; Gonzalez, Pamela; Oliva, Bárbara; Cubillos, Sofía; Valdivia, Andrés; Ibañez, Carolina; Brañes, Jorge; Barriga, María Isabel; Bravo, Erasmo; Alonso, Catalina; Bustamente, Eva; Castellon, Enrique; Hidalgo, Patricia; Trigo, Cesar; Panes, Olga; Pereira, Jaime; Mezzano, Diego; Cuello, Mauricio A; Owen, Gareth I

2015-04-15

An increase in circulating platelets, or thrombocytosis, is recognized as an independent risk factor of bad prognosis and metastasis in patients with ovarian cancer; however the complex role of platelets in tumor progression has not been fully elucidated. Platelet activation has been associated with an epithelial to mesenchymal transition (EMT), while Tissue Factor (TF) protein expression by cancer cells has been shown to correlate with hypercoagulable state and metastasis. The aim of this work was to determine the effect of platelet-cancer cell interaction on TF and "Metastasis Initiating Cell (MIC)" marker levels and migration in ovarian cancer cell lines and cancer cells isolated from the ascetic fluid of ovarian cancer patients. With informed patient consent, ascitic fluid isolated ovarian cancer cells, cell lines and ovarian cancer spheres were co-cultivated with human platelets. TF, EMT and stem cell marker levels were determined by Western blotting, flow cytometry and RT-PCR. Cancer cell migration was determined by Boyden chambers and the scratch assay. The co-culture of patient-derived ovarian cancer cells with platelets causes: 1) a phenotypic change in cancer cells, 2) chemoattraction and cancer cell migration, 3) induced MIC markers (EMT/stemness), 3) increased sphere formation and 4) increased TF protein levels and activity. We present the first evidence that platelets act as chemoattractants to cancer cells. Furthermore, platelets promote the formation of ovarian cancer spheres that express MIC markers and the metastatic protein TF. Our results suggest that platelet-cancer cell interaction plays a role in the formation of metastatic foci.

Long non-coding RNA NRON is downregulated in HCC and suppresses tumour cell proliferation and metastasis.

PubMed

Yao, Zhicheng; Xiong, Zhiyong; Li, Ruixi; Liang, Hao; Jia, Changchang; Deng, Meihai

2018-05-14

Dysregulation of long non-coding RNAs is a newly identified mechanism for tumour progression. Previous studies have suggested that the nuclear factor of activated T cells (NFAT) gene plays a very important role in cancer growth and metastasis. However, lncNRON is a newly identified repressor of NFAT, and its function is largely unknown, especially in hepatocellular carcinoma (HCC). Therefore, the expression levels of lncNRON in 215 pairs of HCC tissue were evaluated by qRT-PCR, and its relationship to clinicopathological parameters, recurrence, and survival was analysed. Furthermore, stably overexpressing lncNRON cell lines were constructed and evaluated for cell phenotype. Finally, we detected epithelial-to-mesenchymal transition (EMT) proteins to determine the underlying mechanism involved in lncNRON function. We observed that lncNRON was downregulated in HCC tumour tissues; low lncNRON expression was associated with poor tumour differentiation and the presence of vascular tumour thrombus, which tended to result in poor clinical outcomes, as demonstrated by the recurrence rate and survival curves. Functional analysis showed that lncNRON overexpression impaired colony formation and cell viability and inhibited cell migration and invasion. A study using tumour-bearing mice showed that lncNRON markedly limited tumour growth and lung metastasis in vivo. Importantly, western blot analysis revealed that the expression of the EMT-related epithelial marker, E-cadherin, increased, whereas the expression of mesenchymal markers N-cadherin, snail, and vimentin was attenuated by lncNRON overexpression in HCC cells. Therefore, lower lncNRON expression indicates a poorer clinical outcome in HCC. LncNRON overexpression can suppress HCC growth and metastasis via inhibiting the EMT, and lncNRON may function as a new HCC prognostic marker. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Murine hepatocellular carcinoma derived stem cells reveal epithelial-to-mesenchymal plasticity.

PubMed

Jayachandran, Aparna; Shrestha, Ritu; Dhungel, Bijay; Huang, I-Tao; Vasconcelos, Marianna Yumi Kawashima; Morrison, Brian J; Ramlogan-Steel, Charmaine A; Steel, Jason C

2017-09-26

To establish a model to enrich and characterize stem-like cells from murine normal liver and hepatocellular carcinoma (HCC) cell lines and to further investigate stem-like cell association with epithelial-to-mesenchymal transition (EMT). In this study, we utilized a stem cell conditioned serum-free medium to enrich stem-like cells from mouse HCC and normal liver cell lines, Hepa 1-6 and AML12, respectively. We isolated the 3-dimensional spheres and assessed their stemness characteristics by evaluating the RNA levels of stemness genes and a cell surface stem cell marker by quantitative reverse transcriptase-PCR (qRT-PCR). Next, we examined the relationship between stem cells and EMT using qRT-PCR. Three-dimensional spheres were enriched by culturing murine HCC and normal hepatocyte cell lines in stem cell conditioned serum-free medium supplemented with epidermal growth factor, basic fibroblast growth factor and heparin sulfate. The 3-dimensional spheres had enhanced stemness markers such as Klf4 and Bmi1 and hepatic cancer stem cell (CSC) marker Cd44 compared to parental cells grown as adherent cultures. We report that epithelial markers E-cadherin and ZO-1 were downregulated, while mesenchymal markers Vimentin and Fibronectin were upregulated in 3-dimensional spheres. The 3-dimensional spheres also exhibited changes in expression of Snai , Zeb and Twist family of EMT transcription factors. Our novel method successfully enriched stem-like cells which possessed an EMT phenotype. The isolation and characterization of murine hepatic CSCs could establish a precise target for the development of more effective therapies for HCC.

Polyphenolic Nutrients in Cancer Chemoprevention and Metastasis: Role of the Epithelial-to-Mesenchymal (EMT) Pathway

PubMed Central

Amawi, Haneen; Ashby, Charles R.; Peraman, Ramalingam

2017-01-01

The epithelial-to-mesenchymal transition (EMT) has received significant interest as a novel target in cancer prevention, metastasis, and resistance. The conversion of cells from an epithelial, adhesive state to a mesenchymal, motile state is one of the key events in the development of cancer metastasis. Polyphenols have been reported to be efficacious in the prevention of cancer and reversing cancer progression. Recently, the antimetastatic efficacy of polyphenols has been reported, thereby expanding the potential use of these compounds beyond chemoprevention. Polyphenols may affect EMT pathways, which are involved in cancer metastasis; for example, polyphenols increase the levels of epithelial markers, but downregulate the mesenchymal markers. Polyphenols also alter the level of expression and functionality of important proteins in other signaling pathways that control cellular mesenchymal characteristics. However, the specific proteins that are directly affected by polyphenols in these signaling pathways remain to be elucidated. The aim of this review is to analyze current evidence regarding the role of polyphenols in attenuating EMT-mediated cancer progression and metastasis. We also discuss the role of the most important polyphenol subclasses and members of the polyphenols in reversing metastasis and targeting EMT. Finally, limitations and future directions to improve our understanding in this field are discussed. PMID:28825675

A novel interaction of PAK4 with PPARγ to regulate Nox1 and radiation-induced epithelial-to-mesenchymal transition in glioma.

PubMed

Kesanakurti, D; Maddirela, D; Banasavadi-Siddegowda, Y K; Lai, T-H; Qamri, Z; Jacob, N K; Sampath, D; Mohanam, S; Kaur, B; Puduvalli, V K

2017-09-14

Tumor recurrence in glioblastoma (GBM) is, in part, attributed to increased epithelial-to-mesenchymal transition (EMT) and enhanced tumor cell dissemination in adjacent brain parenchyma after ionizing radiation (IR). EMT is associated with aggressive behavior, increased stem-like characteristics and treatment resistance in malignancies; however, the underlying signaling mechanisms that regulate EMT are poorly understood. We identified grade-dependent p21-activated kinases 4 (PAK4) upregulation in gliomas and further determined its role in mesenchymal transition and radioresistance. IR treatment significantly elevated expression and nuclear localization of PAK4 in correlation with induction of reactive oxygen species (ROS) and mesenchymal transition in GBM cells. Stable PAK4 overexpression promoted mesenchymal transition by elevating EMT marker expression in these cells. Of note, transcription factor-DNA-binding arrays and chromatin immunoprecipitation experiments identified the formation of a novel nuclear PAK4/PPARγ complex which was recruited to the promoter of Nox1, a peroxisome proliferator-activated receptor gamma (PPARγ) target gene. In addition, IR further elevated PAK4/PPARγ complex co-recruitment to Nox1 promoter, and increased Nox1 expression and ROS levels associated with mesenchymal transition in these cells. Conversely, specific PAK4 downregulation decreased PPARγ-mediated Nox1 expression and suppressed EMT in IR-treated cells. In vivo orthotopic tumor experiments showed inhibition of growth and suppression of IR-induced PPARγ and Nox1 expression by PAK4 downregulation in tumors. Our results provide the first evidence of a novel role for PAK4 in IR-induced EMT and suggest potential therapeutic efficacy of targeting PAK4 to overcome radioresistance in gliomas.

Irisin suppresses the migration, proliferation, and invasion of lung cancer cells via inhibition of epithelial-to-mesenchymal transition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shao, Lei; Jinan Central Hospital Affiliated to Shandong University, Jinan, 250012; Li, Huanjie

Irisin is involved in promoting metabolism, immune regulation, and affects chronic inflammation in many systemic diseases, including gastric cancer. However, the role of irisin in lung cancer is not well characterized. To determine whether irisin has a protective effect against lung cancer, we cultured A549 and NCI-H446 lung cancer cells and treated them with irisin. We detected the proliferation by MTT assay, and assessed the migration and invasion of the cells by scratch wound healing assay and Tran-swell assay. The expression levels of epithelial-to-mesenchymal transition (EMT) markers and the related signaling pathways were detected by western blot analysis. Meanwhile, anmore » inhibitor of PI3K was used to investigate the effect of irsin. Finally, the expression of Snail was detected. We demonstrated that irisin inhibits the proliferation, migration, and invasion of lung cancer cells, and has a novel role in mediating the PI3K/AKT pathway in the cells. Irisin can reverse the activity of EMT and inhibit the expression of Snail via mediating the PI3K/AKT pathway, which is a key regulator of Snail. These results revealed that irisin inhibited EMT and reduced the invasion of lung cancer cells via the PI3K/AKT/Snail pathway. - Highlights: • Irisin inhibits the proliferation of lung cancer cells. • Irisin inhibits the migration and invasion of lung cancer cells. • Irisin affects the expression of EMT markers via inhibiting the PI3K/AKT pathway in lung cancer cells. • Irisin induces Snail downregulation via PI3K/AKT pathway activation.« less

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

PubMed Central

Ando, Seijitsu; Otani, Hitomi; Yagi, Yasuhiro; Kawai, Kenzo; Araki, Hiromasa; Fukuhara, Shirou; Inagaki, Chiyoko

2007-01-01

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 transition (EMT) as monitored by cell shapes, and epithelial or myofibroblast marker at least partly through EGFR transactivation via receptor-linked Src activation. PMID:17433115

Disruption of β-catenin/CBP signaling inhibits human airway epithelial-mesenchymal transition and repair.

PubMed

Moheimani, Fatemeh; Roth, Hollis M; Cross, Jennifer; Reid, Andrew T; Shaheen, Furquan; Warner, Stephanie M; Hirota, Jeremy A; Kicic, Anthony; Hallstrand, Teal S; Kahn, Michael; Stick, Stephen M; Hansbro, Philip M; Hackett, Tillie-Louise; Knight, Darryl A

2015-11-01

The epithelium of asthmatics is characterized by reduced expression of E-cadherin and increased expression of the basal cell markers ck-5 and p63 that is indicative of a relatively undifferentiated repairing epithelium. This phenotype correlates with increased proliferation, compromised wound healing and an enhanced capacity to undergo epithelial-mesenchymal transition (EMT). The transcription factor β-catenin plays a vital role in epithelial cell differentiation and regeneration, depending on the co-factor recruited. Transcriptional programs driven by the β-catenin/CBP axis are critical for maintaining an undifferentiated and proliferative state, whereas the β-catenin/p300 axis is associated with cell differentiation. We hypothesized that disrupting the β-catenin/CBP signaling axis would promote epithelial differentiation and inhibit EMT. We treated monolayer cultures of human airway epithelial cells with TGFβ1 in the presence or absence of the selective small molecule ICG-001 to inhibit β-catenin/CBP signaling. We used western blots to assess expression of an EMT signature, CBP, p300, β-catenin, fibronectin and ITGβ1 and scratch wound assays to assess epithelial cell migration. Snai-1 and -2 expressions were determined using q-PCR. Exposure to TGFβ1 induced EMT, characterized by reduced E-cadherin expression with increased expression of α-smooth muscle actin and EDA-fibronectin. Either co-treatment or therapeutic administration of ICG-001 completely inhibited TGFβ1-induced EMT. ICG-001 also reduced the expression of ck-5 and -19 independent of TGFβ1. Exposure to ICG-001 significantly inhibited epithelial cell proliferation and migration, coincident with a down regulation of ITGβ1 and fibronectin expression. These data support our hypothesis that modulating the β-catenin/CBP signaling axis plays a key role in epithelial plasticity and function. Copyright © 2015 Elsevier Ltd. All rights reserved.

Epithelial-to-Mesenchymal Transition and MicroRNAs in Lung Cancer

PubMed Central

Pécuchet, Nicolas; Imbeaud, Sandrine; Pallier, Karine; Didelot, Audrey; Roussel, Hélène; Gibault, Laure; Fabre, Elizabeth; Le Pimpec-Barthes, Françoise; Laurent-Puig, Pierre; Blons, Hélène

2017-01-01

Despite major advances, non-small cell lung cancer (NSCLC) remains the major cause of cancer-related death in developed countries. Metastasis and drug resistance are the main factors contributing to relapse and death. Epithelial-to-mesenchymal transition (EMT) is a complex molecular and cellular process involved in tissue remodelling that was extensively studied as an actor of tumour progression, metastasis and drug resistance in many cancer types and in lung cancers. Here we described with an emphasis on NSCLC how the changes in signalling pathways, transcription factors expression or microRNAs that occur in cancer promote EMT. Understanding the biology of EMT will help to define reversing process and treatment strategies. We will see that this complex mechanism is related to inflammation, cell mobility and stem cell features and that it is a dynamic process. The existence of intermediate phenotypes and tumour heterogeneity may be debated in the literature concerning EMT markers, EMT signatures and clinical consequences in NSCLC. However, given the role of EMT in metastasis and in drug resistance the development of EMT inhibitors is an interesting approach to counteract tumour progression and drug resistance. This review describes EMT involvement in cancer with an emphasis on NSCLC and microRNA regulation. PMID:28771186

Emodin ameliorates high glucose induced-podocyte epithelial-mesenchymal transition in-vitro and in-vivo.

PubMed

Chen, Tingfang; Zheng, Li Yang; Xiao, Wenzhen; Gui, Dingkun; Wang, Xiaoxia; Wang, Niansong

2015-01-01

Epithelial-to-mesenchymal transition (EMT) is a potential pathway leading to podocyte depletion and proteinuria in diabetic kidney disease (DKD). Here, we investigated the protective effects of Emodin (EMO) on high glucose (HG) induced-podocyte EMT in-vitro and in-vivo. Conditionally immortalized mouse podocytes were exposed to HG with 30 μg /ml of EMO and 1 μmol/ml of integrin-linked kinase (ILK) inhibitor QLT0267 for 24 h. Streptozotocin (STZ)-induced diabetic rats were treated with EMO at 20 mg· kg(-1)· d(-1) and QLT0267 at 10 mg· kg(-1)· w(-1) p.o., for 12 weeks. Albuminuria and blood glucose level were measured. Immunohistochemistry, immunofluorescence, western blotting and real-time PCR were used to detect expression of ILK, the epithelial marker of nephrin and the mesenchymal marker of desmin in-vitro and in-vivo. HG increased podocyte ILK and desmin expression while decreased nephrin expression. However, EMO significantly inhibited ILK and desmin expression and partially restored nephrin expression in HG-stimulated podocytes. These in-vitro observations were further confirmed in-vivo. Treatment with EMO for 12 weeks attenuated albuminuria, renal histopathology and podocyte foot process effacement in diabetic rats. EMO also repressed renal ILK and desmin expression, preserved nephrin expression, as well as ameliorated albuminuria in STZ-induced diabetic rats. EMO ameliorated glucose-induced EMT and subsequent podocyte dysfunction partly through ILK and desmin inhibition as well as nephrin upregulatiotion, which might provide a potential novel therapeutic option for DKD. © 2015 S. Karger AG, Basel.

Kindlin-2 regulates renal tubular cell plasticity by activation of Ras and its downstream signaling.

PubMed

Wei, Xiaofan; Wang, Xiang; Xia, Yang; Tang, Yan; Li, Feng; Fang, Weigang; Zhang, Hongquan

2014-01-01

Kindlin-2 is an adaptor protein that contributes to renal tubulointerstitial fibrosis (TIF). Epithelial-to-mesenchymal transition (EMT) in tubular epithelial cells was regarded as one of the key events in TIF. To determine whether kindlin-2 is involved in the EMT process, we investigated its regulation of EMT in human kidney tubular epithelial cells (TECs) and explored the underlying mechanism. In this study, we found that overexpression of kindlin-2 suppressed epithelial marker E-cadherin and increased the expression of fibronectin and the myofibroblast marker α-smooth muscle actin (SMA). Kindlin-2 significantly activated ERK1/2 and Akt, and inhibition of ERK1/2 or Akt reversed kindlin-2-induced EMT in human kidney TECs. Mechanistically, kindlin-2 interacted with Ras and son of sevenless (Sos)-1. Furthermore, overexpression of kindlin-2 increased Ras activation through recruiting Sos-1. Treatment with a Ras inhibitor markedly repressed kindlin-2-induced ERK1/2 and Akt activation, leading to restraint of EMT. We further demonstrated that knockdown of kindlin-2 inhibited EGF-induced Ras-Sos-1 interaction, resulting in reduction of Ras activation and suppression of EMT stimulated by EGF. Importantly, we found that depletion of kindlin-2 significantly inhibited activation of ERK1/2 and Akt signaling in mice with unilateral ureteral obstruction. We conclude that kindlin-2, through activating Ras and the downstream ERK1/2 and Akt signaling pathways, plays an important role in regulating renal tubular EMT and could be a potential therapeutic target for the treatment of fibrotic kidney diseases.

TMPRSS4 induces cancer stem cell-like properties in lung cancer cells and correlates with ALDH expression in NSCLC patients.

PubMed

de Aberasturi, Arrate L; Redrado, Miriam; Villalba, Maria; Larzabal, Leyre; Pajares, Maria J; Garcia, Javier; Evans, Stephanie R; Garcia-Ros, David; Bodegas, Maria Elena; Lopez, Lissett; Montuenga, Luis; Calvo, Alfonso

2016-01-28

Metastasis involves a series of changes in cancer cells that promote their escape from the primary tumor and colonization to a new organ. This process is related to the transition from an epithelial to a mesenchymal phenotype (EMT). Recently, some authors have shown that migratory cells with an EMT phenotype share properties of cancer stem cells (CSCs), which allow them to form a new tumor mass. The type II transmembrane serine protease TMPRSS4 is highly expressed in some solid tumors, promotes metastasis and confers EMT features to cancer cells. We hypothesized that TMPRSS4 could also provide CSC properties. Overexpression of TMPRSS4 reduces E-cadherin and induces N-cadherin and vimentin in A549 lung cancer cells, supporting an EMT phenotype. These changes are accompanied by enhanced migration, invasion and tumorigenicity in vivo. TMPRSS4 expression was highly increased in a panel of lung cancer cells cultured as tumorspheres (a typical assay to enrich for CSCs). H358 and H441 cells with knocked-down TMPRSS4 levels were significantly less able to form primary and secondary tumorspheres than control cells. Moreover, they showed a lower proportion of ALDH+ cells (examined by FACS analysis) and lower expression of some CSC markers than controls. A549 cells overexpressing TMPRSS4 conferred the opposite phenotype and were also more sensitive to the CSC-targeted drug salinomycin than control cells, but were more resistant to regular chemotherapeutic drugs (cisplatin, gemcitabine and 5-fluorouracil). Analysis of 70 NSCLC samples from patients revealed a very significant correlation between TMPRSS4 expression and CSC markers ALDH (p = 0.0018) and OCT4 (p = 0.0004), suggesting that TMPRSS4 is associated with a CSC phenotype in patients' tumors. These results show that TMPRSS4, in addition to inducing EMT, can also promote CSC features in lung cancer; therefore, CSC-targeting drugs could be an appropriate treatment for TMPRSS4+ tumors. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Expression of the extracellular matrix protein periostin in liver tumours and bile duct carcinomas.

PubMed

Riener, Marc-Oliver; Fritzsche, Florian R; Soll, Christopher; Pestalozzi, Bernhard C; Probst-Hensch, Nicole; Clavien, Pierre-Alain; Jochum, Wolfram; Soltermann, Alex; Moch, Holger; Kristiansen, Glen

2010-04-01

To study the relevance of periostin, known to be involved in epithelial-mesenchymal transition (EMT), in hepatocellular and bile duct cancer. Immunohistochemical periostin expression was semiquantitatively analysed in normal liver tissue (n = 20), hepatocellular carcinoma (HCC; n = 91), liver-cell adenoma (n = 9), focal nodular hyperplasia (n = 13) and bile duct carcinomas (BDC; n = 116) using tissue microarrays. Normal bile ducts, gallbladder epithelium and hepatocytes showed weak cytoplasmic periostin expression. In HCC, there was strong epithelial periostin expression in 19/91 (20.9%) and strong stromal periostin expression in 10/91 cases (11%). Epithelial expression in tumour cells was significantly associated with a higher tumour grade (P < 0.05) and hepatitis B virus infection (P = 0.007). Importantly, there was no strong periostin expression in benign liver tumours. Strong stromal periostin expression was detected in 78/116 (67.2%) BDC and strong epithelial expression in 39/116 (33.6%) BDC. pT stage, differentiation grade and proliferation rate in primary BDC were independent of periostin expression. Epithelial periostin expression was associated with reduced overall survival on univariate and multivariate analysis. The EMT protein periostin is expressed in the stroma and epithelium of a subset of BDC and HCC. Epithelial periostin expression is a marker for malignant transformation of hepatocytes and a novel prognostic marker in BDC.

Dioxin Receptor Expression Inhibits Basal and Transforming Growth Factor β-induced Epithelial-to-mesenchymal Transition*

PubMed Central

Rico-Leo, Eva M.; Alvarez-Barrientos, Alberto; Fernandez-Salguero, Pedro M.

2013-01-01

Recent studies have emphasized the role of the dioxin receptor (AhR) in maintaining cell morphology, adhesion, and migration. These novel AhR functions depend on the cell phenotype, and although AhR expression maintains mesenchymal fibroblasts migration, it inhibits keratinocytes motility. These observations prompted us to investigate whether AhR modulates the epithelial-to-mesenchymal transition (EMT). For this, we have used primary AhR+/+ and AhR−/− keratinocytes and NMuMG cells engineered to knock down AhR levels (sh-AhR) or to express a constitutively active receptor (CA-AhR). Both AhR−/− keratinocytes and sh-AhR NMuMG cells had increased migration, reduced levels of epithelial markers E-cadherin and β-catenin, and increased expression of mesenchymal markers Snail, Slug/Snai2, vimentin, fibronectin, and α-smooth muscle actin. Consistently, AhR+/+ and CA-AhR NMuMG cells had reduced migration and enhanced expression of epithelial markers. AhR activation by the agonist FICZ (6-formylindolo[3,2-b]carbazole) inhibited NMuMG migration, whereas the antagonist α-naphthoflavone induced migration as did AhR knockdown. Exogenous TGFβ exacerbated the promigratory mesenchymal phenotype in both AhR-expressing and AhR-depleted cells, although the effects on the latter were more pronounced. Rescuing AhR expression in sh-AhR cells reduced Snail and Slug/Snai2 levels and cell migration and restored E-cadherin levels. Interference of AhR in human HaCaT cells further supported its role in EMT. Interestingly, co-immunoprecipitation and immunofluorescence assays showed that AhR associates in common protein complexes with E-cadherin and β-catenin, suggesting the implication of AhR in cell-cell adhesion. Thus, basal or TGFβ-induced AhR down-modulation could be relevant in the acquisition of a motile EMT phenotype in both normal and transformed epithelial cells. PMID:23382382

Silencing of BAG3 inhibits the epithelial-mesenchymal transition in human cervical cancer.

PubMed

Song, Fei; Wang, Geng; Ma, Zhifang; Ma, Yuebing; Wang, Yingying

2017-11-10

Bcl2-associated athanogene 3 (BAG3) has been reported to be involved in aggressive progression of many tumors. In the present study, we examined the expression of BAG3 in human cervical cancer (CC) tissues and investigated the role of BAG3 in SiHa and HeLa cell growth, migration, and invasion. Here, we found that most of CC tissues highly expressed the protein and mRNA of BAG3, while their expression was obviously lower in paired normal tissues (all p<0.001). BAG3 expression was associated with FIGO stage and metastasis (all p<0.05). In-vitro analysis demonstrated that BAG3 siRNAs inhibited SiHa and HeLa cell growth, invasion and migration. Mechanically, BAG3 siRNAs inhibited the expression of EMT-regulating markers, involving MMP2, Slug and N-cadherin, and increased the expression of E-cadherin. In a xenograft nude model, BAG3 siRNAs inhibited tumor growth and the expression of EMT biomarkers. In conclusion, BAG3 is involved in the EMT process, including cell growth, invasion and migration in the development of CC. Thus, BAG3 target might be recommended as a novel therapeutic approach.

Silencing of BAG3 inhibits the epithelial-mesenchymal transition in human cervical cancer

PubMed Central

Song, Fei; Wang, Geng; Ma, Zhifang; Ma, Yuebing; Wang, Yingying

2017-01-01

Bcl2-associated athanogene 3 (BAG3) has been reported to be involved in aggressive progression of many tumors. In the present study, we examined the expression of BAG3 in human cervical cancer (CC) tissues and investigated the role of BAG3 in SiHa and HeLa cell growth, migration, and invasion. Here, we found that most of CC tissues highly expressed the protein and mRNA of BAG3, while their expression was obviously lower in paired normal tissues (all p<0.001). BAG3 expression was associated with FIGO stage and metastasis (all p<0.05). In-vitro analysis demonstrated that BAG3 siRNAs inhibited SiHa and HeLa cell growth, invasion and migration. Mechanically, BAG3 siRNAs inhibited the expression of EMT-regulating markers, involving MMP2, Slug and N-cadherin, and increased the expression of E-cadherin. In a xenograft nude model, BAG3 siRNAs inhibited tumor growth and the expression of EMT biomarkers. In conclusion, BAG3 is involved in the EMT process, including cell growth, invasion and migration in the development of CC. Thus, BAG3 target might be recommended as a novel therapeutic approach. PMID:29221135

The relevance of EMT in breast cancer metastasis: Correlation or causality?

PubMed

Bill, Ruben; Christofori, Gerhard

2015-06-22

Although major progress has been achieved in treating breast cancer patients, metastatic breast cancer still remains a deadly disease. A full understanding of the process of systemic cancer cell dissemination is therefore critical to develop next generation therapies. A plethora of experimental data points toward a central role of an epithelial to mesenchymal transition (EMT) in the multistep cascade of metastasis formation. However, in patients the data are based on correlative studies which often, but not always, tie the expression of EMT markers to cancer invasion, metastasis and poor clinical outcome. Moreover, the notion that cancer cells are able to switch between different modes of migration asks for a thorough review of the actual relevance of EMT in cancer metastasis. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Curcumin inhibits tumor epithelial-mesenchymal transition by downregulating the Wnt signaling pathway and upregulating NKD2 expression in colon cancer cells

PubMed Central

ZHANG, ZEWEI; CHEN, HAITAO; XU, CHAO; SONG, LU; HUANG, LULU; LAI, YUEBIAO; WANG, YUQI; CHEN, HANLU; GU, DANLIN; REN, LILI; YAO, QINGHUA

2016-01-01

Tumor invasion and metastasis are closely associated with epithelial-mesenchymal transition (EMT). EMT refers to epithelial cells under physiological and pathological conditions that are specific to mesenchymal transition. Curcumin inhibits EMT progression via Wnt signaling. The Wnt signaling pathway is a conservative EMT-related signaling pathway that is involved in the development of various tumors. In the present study, MTS assays were employed to analyze the proliferation of curcumin-treated cells. Naked cuticle homolog 2 (NKD2), chemokine receptor 4 (CXCR4) and antibodies associated with EMT were examined in SW620 colorectal cancer cell lines using western blot analysis and real-time qPCR. NKD2 small-interfering RNA (siRNA) and CXCR4 expression plasmid was synthesized and transfected into the colorectal cancer cell lines, and NKD2 and CXCR4 expression levels were detected. The results showed that curcumin significantly inhibited the proliferation of colorectal cancer cells and upregulated the expression of NKD2 in SW620 colorectal cancer cells and in the xenograft, resulting in the downregulation of key markers in the Wnt signaling. In addition, the progression of ETM was inhibited due to the overexpression of E-cadherin as well as the downregulation of vimentin. Curcumin also inhibited tumor metastasis by downregulating the expression of CXCR4 significantly. The results suggested involvement of the NKD2-Wnt-CXCR4 signaling pathway in colorectal cancer cells. In addition, curcumin is inhibit this signaling and the development of colorectal cancer. PMID:26985708

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Tao; Li, Xiao-Na; Li, Xing-Guang

Highlights: • DNAJC6 is up-regulated in hepatocellular carcinoma tissues. • DNAJC6 promotes hepatocellular carcinoma cell proliferation and invasion. • DNAJC6 induces epithelial–mesenchymal transition by activating transforming growth factor β signaling. - Abstract: Epithelial–mesenchymal transition (EMT) is a developmental program, which is associated with hepatocellular carcinoma (HCC) development and progression. DNAJC6 (DNA/HSP40 homolog subfamily C member 6) encodes auxilin, which is responsible for juvenile Parkinsonism with phenotypic variability. However, the role of DNAJC6 in HCC development and progression is limited. Here, we report that DNAJC6 is up-regulated in HCC tissues and up-regulation of DNAJC6 expression predicts poor outcome in patients withmore » HCC. Furthermore, overexpression of DNAJC6 enhances the ability for acquisition of mesenchymal traits, enhanced cell proliferation and invasion. DNAJC6 positively regulated expression of EMT-related transcription factor, also activating transforming growth factor β (TGF-β) pathway to contribute to EMT. Our findings demonstrated an important function of DNAJC6 in the progression of HCC by induction of EMT, and they implicate DNAJC6 as a marker of poor outcome in HCC.« less

Hibiscus sabdariffa polyphenols alleviate insulin resistance and renal epithelial to mesenchymal transition: a novel action mechanism mediated by type 4 dipeptidyl peptidase.

PubMed

Peng, Chiung-Huei; Yang, Yi-Sun; Chan, Kuei-Chuan; Wang, Chau-Jong; Chen, Mu-Lin; Huang, Chien-Ning

2014-10-08

The epithelial to mesenchymal transition (EMT) is important in renal fibrosis. Ser307 phosphorylation of insulin receptor substrate-1 (IRS-1 (S307)) is a hallmark of insulin resistance. We report that polyphenol extracts of Hibiscus sabdariffa (HPE) ameliorate diabetic nephropathy and EMT. Recently it has been observed that type 4 dipeptidyl peptidase (DPP-4) inhibitor linagliptin is effective for treating type 2 diabetes and albuminuria. We investigated if DPP-4 and insulin resistance are involved in renal EMT and explored the role of HPE. In high glucose-stimulated tubular cells, HPE, like linagliptin, inhibited DPP-4 activation, thereby regulating vimentin (EMT marker) and IRS-1 (S307). IRS-1 knockdown revealed its essential role in mediating downstream EMT. In type 2 diabetic rats, pIRS-1 (S307) abundantly surrounds the tubular region, with increased vimentin in kidney. Both the expressions were reduced by HPE. In conclusion, HPE exerts effects similar to those of linagliptin, which improves insulin resistance and EMT, and could be an adjuvant to prevent diabetic nephropathy.

African Americans with pancreatic ductal adenocarcinoma exhibit gender differences in Kaiso expression

PubMed Central

Mukherjee, Angana; Jones, Jacqueline; Karanam, Balasubramanyam; Davis, Melissa; Jaynes, Jesse; Reams, R. Renee; Dean-Colomb, Windy; Yates, Clayton

2016-01-01

Kaiso, a bi-modal transcription factor, regulates gene expression, and is elevated in breast, prostate, and colon cancers. Depletion of Kaiso in other cancer types leads to a reduction in markers for the epithelial–mesenchymal transition (EMT) (Jones et al., 2014), however its clinical implications in pancreatic ductal adenocarcinoma (PDCA) have not been widely explored. PDCA is rarely detected at an early stage but is characterized by rapid progression and invasiveness. We now report the significance of the subcellular localization of Kaiso in PDCAs from African Americans. Kaiso expression is higher in the cytoplasm of invasive and metastatic pancreatic cancers. In males, cytoplasmic expression of Kaiso correlates with cancer grade and lymph node positivity. In male and female patients, cytoplasmic Kaiso expression correlates with invasiveness. Also, nuclear expression of Kaiso increases with increased invasiveness and lymph node positivity. Further, analysis of the largest PDCA dataset available on ONCOMINE shows that as Kaiso increases, there is an overall increase in Zeb1, which is the inverse for E-cadherin. Hence, these findings suggest a role for Kaiso in the progression of PDCAs, involving the EMT markers, E-cadherin and Zeb1. PMID:27424525

YB-1 expression promotes epithelial-to-mesenchymal transition in prostate cancer that is inhibited by a small molecule fisetin

PubMed Central

Khan, Mohammad Imran; Adhami, Vaqar Mustafa; Lall, Rahul Kumar; Sechi, Mario; Joshi, Dinesh C.; Haidar, Omar M.; Syed, Deeba Nadeem; Siddiqui, Imtiaz Ahmad; Chiu, Shing-Yan; Mukhtar, Hasan

2014-01-01

Epithelial-to-mesenchymal transition (EMT) plays an important role in prostate cancer (PCa) metastasis. The transcription/translation regulatory Y-box binding protein-1 (YB-1) is known to be associated with cancer metastasis. We observed that YB-1 expression increased with tumor grade and showed an inverse relationship with E-cadherin in a human PCa tissue array. Forced YB-1 expression induced a mesenchymal morphology that was associated with down regulation of epithelial markers. Silencing of YB-1 reversed mesenchymal features and decreased cell proliferation, migration and invasion in PCa cells. YB-1 is activated directly via Akt mediated phosphorylation at Ser102 within the cold shock domain (CSD). We next identified fisetin as an inhibitor of YB-1 activation. Computational docking and molecular dynamics suggested that fisetin binds on the residues from β1 - β4 strands of CSD, hindering Akt's interaction with YB-1. Calculated free binding energy ranged from −11.9845 to −9.6273 kcal/mol. Plasmon Surface Resonance studies showed that fisetin binds to YB-1 with an affinity of approximately 35 μM, with both slow association and dissociation. Fisetin also inhibited EGF induced YB-1 phosphorylation and markers of EMT both in vitro and in vivo. Collectively our data suggest that YB-1 induces EMT in PCa and identify fisetin as an inhibitor of its activation. PMID:24770864

A Novel Differentiation Therapy Approach to Reduce the Metastatic Potential of Basal, Highly Metastatic, Triple-Negative Breast Cancers

DTIC Science & Technology

2012-05-01

subset enriched in epithelial-to- mesenchymal transition and stem cell characteristics. Cancer Res 69: 4116–4124. Hoenerhoff MJ, Chu I, Barkan D, Liu ZY...expression of epithelial markers and loss of mesenchymal markers in MB- 231 cells (Task1a) Our analyses of m icroarray data com paring 231-Empty cells ...is considered a hallmark of EMT (Yang and Weinberg 2008). MB-231 cells lack E-cadherin expression and exhibit a more mesenchymal phenotype

Genome-wide identification of epithelial-mesenchymal transition-associated microRNAs reveals novel targets for glioblastoma therapy

PubMed Central

Zhang, Yong; Zeng, Ailiang; Liu, Shuheng; Li, Rui; Wang, Xiefeng; Yan, Wei; Li, Hailin; You, Yongping

2018-01-01

MicroRNAs (miRNA) regulate a number of cellular processes. Recent studies have indicated that these molecules function in the epithelial-mesenchymal transition (EMT). However, the crucial systematic role of EMT and miRNAs together in glioblastoma (GBM) remains poorly understood. The present study demonstrated that EMT was closely associated with malignant progression and clinical outcome using three independent glioma databases (GSE16011, Rembrandt and The Cancer Genome Atlas). Furthermore, integrated analysis of miRNAs and mRNA profiling in 491 GBM samples revealed an EMT biological process associated with an miRNA profile (19 positively and 18 negatively correlated miRNAs). Among these miRNAs, miR-95 and miR-223 indicated a high level of functional validation, reflecting their positive correlation with EMT. Additionally, the upregulation of miR-95, which was negatively correlated with EMT, inhibited cellular invasion in glioma U251 and LN229 cells and decreased the expression of the mesenchymal marker N-catenin, whereas an miRNA positively correlated with EMT, miR-223, exhibited the opposite effect. Therefore, the results of the present study could further enhance the current understanding of the functions of miRNAs in GBM, indicating that the EMT-specific miRNA signature may represent a novel target for GBM therapy. PMID:29740486

GSK-3β inhibitor attenuates urinary albumin excretion in type 2 diabetic db/db mice, and delays epithelial-to-mesenchymal transition in mouse kidneys and podocytes.

PubMed

Wan, Jia; Li, Peng; Liu, Dong-Wei; Chen, Ying; Mo, Hai-Zhen; Liu, Ben-Guo; Chen, Wen-Jie; Lu, Xiao-Qing; Guo, Jia; Zhang, Qian; Qiao, Ying-Jin; Liu, Zhang-Suo; Wan, Guang-Rui

2016-08-01

The mechanism underlying epithelial‑to‑mesenchymal transition (EMT) caused by high glucose (HG) stimulation in diabetic nephropathy (DN) remains to be fully elucidated. The present study investigated the effects of HG on EMT and the activity of glycogen synthase kinase 3β (GSK‑3β) in podocytes and the kidneys of db/db mice, and assessed the effects of (2'Z, 3'E)‑6‑bromoindirubin‑3'‑oxime (BIO), an inhibitor of GSK‑3β, on EMT and glomerular injury. The resulting data showed that the activity of GSK‑3β was upregulated by HG and downregulated by BIO in the podocytes and the renal cortex. The expression levels of epithelial markers, including nephrin, podocin and synaptopodin, were decreased by HG and increased by BIO, whereas the reverse were true for mesenchymal markers, including α‑smooth muscle actin (α‑SMA) and fibronectin. The expression levels of β‑catenin and Snail, in contrast to current understanding of the Wnt signaling pathway, were increased by HG and decreased by BIO. In addition, expression of the vitamin D receptor (VDR) was decreased by HG and increased by BIO. In conclusion, the present study revealed that the mechanism by which BIO inhibited HG‑mediated EMT in podocytes and the renal cortex was primarily due to the VDR. Treatment with BIO protected renal function by maintaining the integrity of the filtration membrane and decreasing UAE, but not by regulating blood glucose. Therefore, GSK‑3β may be used as a sensitive biomarker of DN, and its inhibition by BIO may be effective in the treatment of DN.

Systems analysis reveals a transcriptional reversal of the mesenchymal phenotype induced by SNAIL-inhibitor GN-25

PubMed Central

2013-01-01

Background HMLEs (HMLE-SNAIL and Kras-HMLE, Kras-HMLE-SNAIL pairs) serve as excellent model system to interrogate the effect of SNAIL targeted agents that reverse epithelial-to-mesenchymal transition (EMT). We had earlier developed a SNAIL-p53 interaction inhibitor (GN-25) that was shown to suppress SNAIL function. In this report, using systems biology and pathway network analysis, we show that GN-25 could cause reversal of EMT leading to mesenchymal-to-epithelial transition (MET) in a well-recognized HMLE-SNAIL and Kras-HMLE-SNAIL models. Results GN-25 induced MET was found to be consistent with growth inhibition, suppression of spheroid forming capacity and induction of apoptosis. Pathway network analysis of mRNA expression using microarrays from GN-25 treated Kras-HMLE-SNAIL cells showed an orchestrated global re-organization of EMT network genes. The expression signatures were validated at the protein level (down-regulation of mesenchymal markers such as TWIST1 and TWIST2 that was concurrent with up-regulation of epithelial marker E-Cadherin), and RNAi studies validated SNAIL dependent mechanism of action of the drug. Most importantly, GN-25 modulated many major transcription factors (TFs) such as inhibition of oncogenic TFs Myc, TBX2, NR3C1 and led to enhancement in the expression of tumor suppressor TFs such as SMAD7, DD1T3, CEBPA, HOXA5, TFEB, IRF1, IRF7 and XBP1, resulting in MET as well as cell death. Conclusions Our systems and network investigations provide convincing pre-clinical evidence in support of the clinical application of GN-25 for the reversal of EMT and thereby reducing cancer cell aggressiveness. PMID:24004452

Syndecan-1 suppresses epithelial-mesenchymal transition and migration in human oral cancer cells.

PubMed

Wang, Xiaofeng; He, Jinting; Zhao, Xiaoming; Qi, Tianyang; Zhang, Tianfu; Kong, Chenfei

2018-04-01

Epithelial-mesenchymal transition (EMT) is one of the major processes that contribute to the occurrence of cancer metastasis. EMT has been associated with the development of oral cancer. Syndecan‑1 (SDC1) is a key cell‑surface adhesion molecule and its expression level inversely correlates with tumor differentiation and prognosis. In the present study, we aimed to determine the role of SDC1 in oral cancer progression and investigate the molecular mechanisms through which SDC1 regulates the EMT and invasiveness of oral cancer cells. We demonstrated that basal SDC1 expression levels were lower in four oral cancer cell lines (KB, Tca8113, ACC2 and CAL‑27), than in normal human periodontal ligament fibroblasts. Ectopic overexpression of SDC1 resulted in morphological transformation, decreased expression of EMT‑associated markers, as well as decreased migration, invasiveness and proliferation of oral cancer cells. In contrast, downregulation of the expression of SDC1 caused the opposite results. Furthermore, the knockdown of endogenous SDC1 activated the extracellular signal‑regulated kinase (ERK) cascade, upregulated the expression of Snail and inhibited the expression of E‑cadherin. In conclusion, our findings revealed that SDC1 suppressed EMT via the modulation of the ERK signaling pathway that, in turn, negatively affected the invasiveness of human oral cancer cells. Our results provided useful evidence about the potential use of SDC1 as a molecular target for therapeutic interventions in human oral cancer.

Bmi-1 promotes invasion and metastasis, and its elevated expression is correlated with an advanced stage of breast cancer

PubMed Central

2011-01-01

Background B-lymphoma Moloney murine leukemia virus insertion region-1 (Bmi-1) acts as an oncogene in various tumors, and its overexpression correlates with a poor outcome in several human cancers. Ectopic expression of Bmi-1 can induce epithelial-mesenchymal transition (EMT) and enhance the motility and invasiveness of human nasopharyngeal epithelial cells (NPECs), whereas silencing endogenous Bmi-1 expression can reverse EMT and reduce the metastatic potential of nasopharyngeal cancer cells (NPCs). Mouse xenograft studies indicate that coexpression of Bmi-1 and H-Ras in breast cancer cells can induce an aggressive and metastatic phenotype with an unusual occurrence of brain metastasis; although, Bmi-1 overexpression did not result in oncogenic transformation of MCF-10A cells. However, the underlying molecular mechanism of Bmi-1-mediated progression and the metastasis of breast cancer are not fully elucidated at this time. Results Bmi-1 expression is more pronouncedly increased in primary cancer tissues compared to matched adjacent non-cancerous tissues. High Bmi-1 expression is correlated with advanced clinicopathologic classifications (T, N, and M) and clinical stages. Furthermore, a high level of Bmi-1 indicates an unfavorable overall survival and serves as a high risk marker for breast cancer. In addition, inverse transcriptional expression levels of Bmi-1 and E-cadherin are detected between the primary cancer tissues and the matched adjacent non-cancerous tissues. Higher Bmi-1 levels are found in the cancer tissue, whereas the paired adjacent non-cancer tissue shows higher E-cadherin levels. Overexpression of Bmi-1 increases the motility and invasive properties of immortalized human mammary epithelial cells, which is concurrent with the increased expression of mesenchymal markers, the decreased expression of epithelial markers, the stabilization of Snail and the dysregulation of the Akt/GSK3β pathway. Consistent with these observations, the repression of Bmi-1 in highly metastatic breast cancer cells remarkably reduces cellular motility, invasion and transformation, as well as tumorigenesis and lung metastases in nude mice. In addition, the repression of Bmi-1 reverses the expression of EMT markers and inhibits the Akt/GSK3β/Snail pathway. Conclusions This study demonstrates that Bmi-1 promotes the invasion and metastasis of human breast cancer and predicts poor survival. PMID:21276221

Role of Epithelial–Mesenchymal Transition in Repair of the Lacrimal Gland after Experimentally Induced Injury

PubMed Central

You, Samantha; Avidan, Orna; Tariq, Ayesha; Ahluwalia, Ivy; Stark, Paul C.; Kublin, Claire L.

2012-01-01

Purpose. Ongoing studies demonstrate that the murine lacrimal gland is capable of repair after experimentally induced injury. It was recently reported that repair of the lacrimal gland involved the mobilization of mesenchymal stem cells (MSCs). These cells expressed the type VI intermediate filament protein nestin whose expression was upregulated during the repair phase. The aim of the present study was to investigate the roles of vimentin, a type III intermediate filament protein and a marker of epithelial–mesenchymal transition (EMT) in repair of the lacrimal gland. Methods. Injury was induced by direct injection of interleukin (IL)-1 into the exorbital lacrimal gland. MSCs were prepared from injured glands using tissue explants. Expression of vimentin and the transcription factor Snai1, a master regulator of EMT, was determined by RT-PCR, Western blotting analysis, and immunofluorescence. Results. These data show that vimentin expression, at both the mRNA and the protein levels, was upregulated during the repair phase (2–3 days postinjury) and returned to the control level when repair ended. Temporal expression of Snai1 mirrored that of vimentin and was localized in cell nuclei. Cultured MSCs isolated from injured lacrimal glands expressed Snai1 and vimentin alongside nestin and alpha smooth muscle actin (another biomarker of EMT). There was a strong positive correlation between Snai1 expression and vimentin expression. Conclusions. It was found that EMT is induced during repair of the lacrimal gland to generate MSCs to initiate repair, and that mesenchymal–epithelial transition is then activated to form acinar and ductal epithelial cells. PMID:22025566

Valproic acid inhibits epithelial‑mesenchymal transition in renal cell carcinoma by decreasing SMAD4 expression.

PubMed

Mao, Shaowei; Lu, Guoliang; Lan, Xiaopeng; Yuan, Chuanwei; Jiang, Wei; Chen, Yougen; Jin, Xunbo; Xia, Qinghua

2017-11-01

Renal cell carcinoma (RCC) is the most common malignancy in urogenital neoplasms worldwide. According to previous studies, valproic acid (VPA), an anticonvulsant drug, can suppress tumor metastasis and decrease the expression level of Mothers against decapentaplegic homolog 4 (SMAD4) and therefore may inhibit epithelial‑mesenchymal transition (EMT), which is responsible for cancer metastasis. However, the association between VPA, EMT and SMAD4 in RCC metastasis remains obscure. In the present study, it was demonstrated that in the RCC cell lines 786‑O and Caki‑1 treated with VPA, the neural (N)‑cadherin, vimentin and SMAD4 protein and mRNA levels were decreased, accompanied with an increase in expression of epithelial (E)‑cadherin. Silencing SMAD4 expression decreased the expression of EMT markers, including N‑cadherin and simultaneously upregulated E‑cadherin in RCC cell lines. SMAD4 overexpression counteracted the VPA‑mediated EMT‑inhibitory effect (P<0.05). The present study demonstrates that VPA inhibited EMT in RCC cells via altering SMAD4 expression. In addition, immunohistochemical staining demonstrated that transforming growth factor‑β (TGF‑β) and low expression of SMAD4 was associated with a lower Fuhrman grade and low expression of transcription intermediary factor 1‑γ was associated with a higher tumor Fuhrman grade (P<0.05), Therefore, based on the regulatory effect of SMAD4 on EMT‑associated transcription factors, SMAD4 which can form a SMAD3/SMAD4 complex induced by TGF‑β, could be a potential anticancer drug target inhibiting tumor invasion and metastasis in RCC.

K-ras mutation promotes ionizing radiation-induced invasion and migration of lung cancer in part via the Cathepsin L/CUX1 pathway.

PubMed

Wang, Long; Zhao, Yifan; Xiong, Yajie; Wang, Wenjuan; Fei, Yao; Tan, Caihong; Liang, Zhongqin

2018-01-15

K-ras mutation is involved in cancer progression including invasion and migration, but the underlying mechanism is not yet clear. Cathepsin L is a lysosomal cysteine protease and has recently been associated with invasion and migration in human cancers when it is overexpressed. Our recent studies have shown that ionizing radiation (IR) enhanced expression of cathepsin L and increased invasion and migration of tumor cells, but the molecular mechanism is still unclear. In the present study, the effects of K-ras mutation and IR induced invasion and migration of lung cancer as well as the underlying mechanisms were investigated both in vitro and in vivo. Firstly, the levels of cathepsin L and epithelial mesenchymal transition (EMT) marker proteins remarkably changed in A549 (K-ras mutant) after irradiation compared with H1299 (K-ras wild), thereby promoting invasion and migration. Additionally, cathepsin L and its downstream transcription factor CUX1/p110 were increased after irradiation in A549 transfected with CUX1/p200, and the proteolytic processing of CUX1 by cathepsin L was remarkably increased after co-transfection of CUX1/p200 and cathepsin L-lentivirus in H1299. In addition, delivery of a mutant K-ras (V12) into HEK 293 cells stimulated EMT after irradiation due to the accumulation of cathepsin L. Moreover, mutated K-ras was associated with IR-induced cathepsin L and EMT in BALB/c nude mice. Finally, the level of cathepsin L expression was higher in samples carrying a K-ras mutation than in wild-type K-ras samples and the mesenchymal markers were upregulated in the samples of mutant K-ras, whereas the epithelial marker E-cadherin was downregulated in non-small cell lung cancers tissues. In conclusion, the findings demonstrated that mutated K-ras promotes cathepsin L expression and plays a pivotal role in EMT of human lung cancer. The regulatory effect of IR-induced cathepsin L on lung cancer invasion and migration was partially attributed to the Cathepsin L /CUX1-mediated EMT signaling pathway. This study will provide cathepsin L as a potential target for tumor therapy. Copyright © 2017 Elsevier Inc. All rights reserved.

Upregulation of CD147 promotes cell invasion, epithelial-to-mesenchymal transition and activates MAPK/ERK signaling pathway in colorectal cancer.

PubMed

Xu, Tao; Zhou, Mingliang; Peng, Lipan; Kong, Shuai; Miao, Ruizheng; Shi, Yulong; Sheng, Hongguang; Li, Leping

2014-01-01

Colorectal cancer (CRC) is one of the most common cancers in the world. CD147, a transmembrane protein, has been reported to be correlated with various cancers. In this study, we aimed to investigate the mechanism of CD147 in regulating drug resistance, cell invasion and epithelial-to-mesenchymal transition (EMT) in CRC cells. qRT-PCR and western blotting were used to evaluated the expression of CD147 in 40 CRC cases and 4 cell lines. Increased expression of CD147 at both mRNA and protein levels was found in CRC samples, and the level of CD147 was correlated with lymph node metastasis. CD147 overexpression increased the 5-Fluorouracil (5-FU) resistance, enhanced the invasion and EMT of CRC cells by regulating EMT markers and MMPs. Adverse results were obtained in CD147 knockdown CRC cell line. Further investigation revealed that CD147 activated MAPK/ERK pathway, ERK inhibitor U0126 suppressed the CD147-induced cell invasion, migration and MMP-2, MMP-9 expression. Taken together, our study indicates that CD147 promotes the 5-FU resistance, and MAPK/ERK signaling pathway is involved in CD147-promoted invasion and EMT of CRC cells.

Upregulation of CD147 promotes cell invasion, epithelial-to-mesenchymal transition and activates MAPK/ERK signaling pathway in colorectal cancer

PubMed Central

Xu, Tao; Zhou, Mingliang; Peng, Lipan; Kong, Shuai; Miao, Ruizheng; Shi, Yulong; Sheng, Hongguang; Li, Leping

2014-01-01

Colorectal cancer (CRC) is one of the most common cancers in the world. CD147, a transmembrane protein, has been reported to be correlated with various cancers. In this study, we aimed to investigate the mechanism of CD147 in regulating drug resistance, cell invasion and epithelial-to-mesenchymal transition (EMT) in CRC cells. qRT-PCR and western blotting were used to evaluated the expression of CD147 in 40 CRC cases and 4 cell lines. Increased expression of CD147 at both mRNA and protein levels was found in CRC samples, and the level of CD147 was correlated with lymph node metastasis. CD147 overexpression increased the 5-Fluorouracil (5-FU) resistance, enhanced the invasion and EMT of CRC cells by regulating EMT markers and MMPs. Adverse results were obtained in CD147 knockdown CRC cell line. Further investigation revealed that CD147 activated MAPK/ERK pathway, ERK inhibitor U0126 suppressed the CD147-induced cell invasion, migration and MMP-2, MMP-9 expression. Taken together, our study indicates that CD147 promotes the 5-FU resistance, and MAPK/ERK signaling pathway is involved in CD147-promoted invasion and EMT of CRC cells. PMID:25550778

FOXQ1 promotes gastric cancer metastasis through upregulation of Snail.

PubMed

Zhang, Jing; Liu, Yimin; Zhang, Jia; Cui, Xiaohai; Li, Gang; Wang, Jiansheng; Ren, Hong; Zhang, Yunfeng

2016-06-01

Gastric cancer (GC) is one of the most common cancers, and the second most common cause of cancer deaths worldwide. Forkhead box Q1 (FOXQ1) is a member of the forkhead transcription factor family and its upregulation is closely correlated with tumor progression and prognosis of multiple cancer types, including GC. FOXQ1 has been shown to regulate EMT and function in human cancers. However, the role of FOXQ1 in regulating EMT in GC and the exactly mechanism has not been clarified. The purpose of this study was to investigate the effects of FOXQ1 on EMT in human GC. FOXQ1 protein was detected by immunohistochemistry in human GC specimens and their clinical significance evaluated. We examined the cell biology and molecular biology changes after overexpression and knockdown of FOXQ1 in gastric cancer cells in vitro. To further understand the underlying mechanisms of EMT promoted by FOXQ1, we examined the changes of target genes of FOXQ1 after overexpression and knockdown of FOXQ1 in gastric cancer cells. In the present study, we demonstrate that FOXQ1 is overexpressed in GC tissues and its expression level is closely correlated with histologic differentiation, pTNM stage, and lymphatic metastasis of GC. Kaplan-Meier survival analysis showed that a high expression level of FOXQ1 resulted in a significantly poor prognosis of GC patients. FOXQ1 modulated GC cell invasion in vitro, and induced E-cadherin repression. FOXQ1 also upregulated the expression of vimentin in vitro. The Snail signaling pathway was likely involved in the induction of EMT by FOXQ1 in GC. Our results demonstrate that FOXQ1 is a prognostic marker for patients with GC, FOXQ1 over-expression is involved in acquisition of the mesenchymal phenotype of gastric cancer cells, and that subsequent Snail expression is essential for induction of EMT. The results suggest that FOXQ1 is a potential therapeutic target for the development of therapies for GC.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jiang, Fujin; Department of Urinary Surgery, Huai'an Hospital to Xuzhou Medical University, Huai'an, Jiangsu; Ma, Song

Lactate dehydrogenase-A(LDH-A) is an important rate-limiting enzyme in the Warburg effect. Survival analysis indicated poor clinical outcomes in MIBC with high LDH-A expression. The results of in vitro experiment indicated that LDH-A promotes MIBC cells proliferation, invasion and migration. The positive relationship between LDH-A expression and CSC/EMT markers was confirmed both in invasive bladder cell line and in 136 MIBC specimens. Thus, we conclude that LDH-A may be a promising target for MIBC. - Highlights: • Survival analysis indicated poor clinical outcomes in MIBC with high LDH-A expression. • IHC analysis of 136 MIBC specimens revealed increased LDH-A is correlated withmore » positive Oct4 and negative E-cadherin. • In vitro experiments demonstrated LDH-A promotes MIBC progression by positive regulation of EMT/CSC.« less

Mastl overexpression is associated with epithelial to mesenchymal transition and predicts a poor clinical outcome in gastric cancer.

PubMed

Sun, Xian-Jun; Li, Yan-Liang; Wang, Long-Gang; Liu, Li-Qing; Ma, Heng; Hou, Wen-Hong; Yu, Jin-Ming

2017-12-01

Microtubule-associated serine/threonine kinase like (Mastl) is deregulated in a number of types of human malignancy and may be a kinase target for cancer treatment. The aim of the present study was to determine the Mastl expression in gastric cancer and to clarify its clinical and prognostic significance. Immunohistochemistry was performed on a cohort of 126 postoperative gastric cancer samples to detect the expression of Mastl and two epithelial to mesenchymal transition (EMT) markers, epithelial-cadherin and Vimentin. The χ 2 test, Kaplan-Meier estimator analysis and Cox's regression model were used to analyze the data. Upregulated Mastl protein expression was observed in the gastric cancer tissues compared with that in the adjacent non-cancerous gastric tissues. Increased Mastl expression was identified in 54/126 (42.9%) gastric cancer samples, and was significantly associated with lymph node metastasis, tumor relapse, EMT status and poor overall survival. Additional analysis demonstrated that the Mastl expression level stratified the patient outcome in stage III, but not stage II tumor subgroups. Cox's regression analysis revealed that increased Mastl expression was an independent prognostic factor for patients with gastric cancer. Mastl expression may be a valuable prognostic marker and a potential target for patients with gastric cancer.

Curcumin reverses irinotecan resistance in colon cancer cell by regulation of epithelial-mesenchymal transition.

PubMed

Zhang, Chunhong; Xu, Yangjie; Wang, Haowen; Li, Gang; Yan, Han; Fei, Zhenghua; Xu, Yunsheng; Li, Wenfeng

2018-04-01

The objective of this study was to investigate the effect and the mechanism by which curcumin reverses irinotecan-induced chemotherapy resistance in colon cancer. Construction of irinotecan-resistant colon cancer model LoVo/CPT-11R cells was performed by increasing drug concentration. The Cell Counting Kit-8 assay was used to detect inhibition of proliferation; cell morphology was observed by an optical microscope. Quantitative RT-PCR and western blotting were performed to detect molecular marker expressions during epithelial-mesenchymal transition (EMT); drug-resistant cells were treated with curcumin at different concentrations and Cell Counting Kit-8 was reperformed to detect cell proliferation after treatments. Drug-resistant cells were then divided into four groups: control group, irinotecan group, curcumin group, and irinotecan+curcumin group; quantitative RT-PCR and western blotting were performed to detect molecular marker expressions during epithelial-mesenchymal transition. Flow cytometry was used to detect cell apoptosis after grouping, and apoptosis-related protein was detected by western blotting. LoVo/CPT-11R cells could survive in culture medium containing irinotecan at 60 μg/ml and the drug-resistance index was 5.69; the drug-resistant cells had a larger volume than normal cells and were poorly connected to each other. E-cadherin expression was downregulated, whereas vimentin and N-cadherin expressions were upregulated. After curcumin treatment, drug-resistant cell proliferation was significantly inhibited; in the curcumin+irinotecan treatment group, E-cadherin expression was upregulated, whereas vimentin and N-cadherin expressions were downregulated. Curcumin could significantly increase cell apoptosis. EMT is involved in the development of irinotecan resistance and curcumin can reverse this drug resistance through reversion of the EMT process.

The mechanisms for lung cancer risk of PM2.5 : Induction of epithelial-mesenchymal transition and cancer stem cell properties in human non-small cell lung cancer cells.

PubMed

Wei, Hongying; Liang, Fan; Cheng, Wei; Zhou, Ren; Wu, Xiaomeng; Feng, Yan; Wang, Yan

2017-11-01

Fine particulate matter (PM 2.5 ) is a major component of air pollutions that are closely associated with increased risk of lung cancer. However, the role of PM 2.5 in the etiology of lung cancer is largely unknown. In this study, we performed acute (24 hours) and chronic (five passages) exposure models to investigate the carcinogenetic mechanisms of PM 2.5 by targeting the induction of epithelial-mesenchymal transition (EMT) and cancer stem cells (CSC) properties in human non-small cell lung cancer cell line A549. We found that both acute and chronic PM 2.5 exposure enhanced cell migration and invasion, decreased mRNA expression of epithelial markers and increased mRNA expression of mesenchymal markers. Chronic PM 2.5 exposure further induced notable EMT morphology and CSC properties, indicating the developing process of cell malignant behaviors from acute to chronic PM 2.5 exposure. CSC properties induced by chronic PM 2.5 exposure characterized with increased cell-surface markers (CD44, ABCG2), self-renewal genes (SOX2 and OCT4), side population cells and neoplastic capacity. Furthermore, the levels of three stemness-associated microRNAs, Let-7a, miR-16 and miR-34a, were found to be significantly downregulated by chronic PM 2.5 exposure, with microarray data analysis from TCGA database showing their lower expression in human lung adenocarcinoma tissues than that in the adjacent normal lung tissues. These data revealed that the induction of EMT and CSC properties were involved in the lung cancer risk of PM 2.5 , and implicated CSC properties and related microRNAs as possible biomarkers for carcinogenicity prediction of PM 2.5 . © 2017 Wiley Periodicals, Inc.

Inhibition of cyclin D1 enhances sensitivity to radiotherapy and reverses epithelial to mesenchymal transition for esophageal cancer cells.

PubMed

Su, Huafang; Jin, Xiance; Shen, Lanxiao; Fang, Ya; Fei, Zhenghua; Zhang, Xuebang; Xie, Congying; Chen, Xiaolei

2016-04-01

Acquired radioresistance during radiotherapy has significantly affected the treatment efficacy in esophageal cancer. Many of radioresistant cancer cells demonstrated epithelial-mesenchymal transition (EMT).We found in previous study that a radioresistant cell line (KYSE-150R) possessed EMT characteristic with cyclin D1 overexpression. Cyclin D1 has been demonstrated to affect the radiation sensitivity in cancer cells. To elucidate the molecular functions of cyclin D1 on EMT phenotypes and esophageal cancer radiosensitivity, we treated the radioresistant esophageal cancer cells (KYSE-150R) and parental cells (KYSE-150) with cyclin D1 small interfering RNA (siRNA). The cell proliferation rate of KYSE-150R and the radiation survival fraction were significantly decreased in cyclin D1 siRNA treatment group. Knocking down cyclin D1 resulted in G0/G1 arrest in KYSE-150R cells. The average number of irradiation-induced γ-H2AX foci increased in the cells treated with cyclin D1 siRNA, indicating impaired DNA double-strand break (DSB) repair in KYSE-150R cells. Cyclin D1 also reversed EMT phenotypes with significantly increased expression of E-cadherin in KYSE-150R cells. However, cyclin D1 siRNA have no radiosensitizing effects on KYSE-150 cells, with no obvious change in EMT marker expression .Our work showed that EMT phenotypes can be reduced and the radiosensitivity of esophageal cancer cells can be enhanced by inhibiting cyclin D1.

Ketamine-induced bladder fibrosis involves epithelial-to-mesenchymal transition mediated by transforming growth factor-β1.

PubMed

Wang, Junpeng; Chen, Yang; Gu, Di; Zhang, Guihao; Chen, Jiawei; Zhao, Jie; Wu, Peng

2017-10-01

Bladder wall fibrosis is a major complication of ketamine-induced cystitis (KC), but the underlying pathogenesis is poorly understood. The aim of the present study was to elucidate the mechanism of ketamine-induced fibrosis in association with epithelial-to-mesenchymal transition (EMT) mediated by transforming growth factor-β1 (TGF-β1). Sprague-Dawley rats were randomly distributed into four groups, which received saline, ketamine, ketamine combined with a TGF-β receptor inhibitor (SB-505124) for 16 wk, or 12 wk of ketamine and 4 wk of abstinence. In addition, the profibrotic effect of ketamine was confirmed in SV-40 immortalized human uroepithelial (SV-HUC-1) cells. The ketamine-treated rats displayed voiding dysfunction and decreased bladder compliance. Bladder fibrosis was accompanied by the appearance of a certain number of cells expressing both epithelial and mesenchymal markers, indicating that epithelial cells might undergo EMT upon ketamine administration. Meanwhile, the expression level of TGF-β1 was significantly upregulated in the urothelium of bladders in ketamine-treated rats. Treatment of SV-HUC-1 cells with ketamine increased the expression of TGF-β1 and EMT-inducing transcription factors, resulting in the downregulation of E-cadherin and upregulation of fibronectin and α-smooth muscle actin. Administration of SB-505124 inhibited EMT and fibrosis both in vitro and vivo. In addition, withdrawal from ketamine did not lead to recovery of bladder urinary function or decreased fibrosis. Taken together, our study shows for the first time that EMT might contribute to bladder fibrosis in KC. TGF-β1 may have an important role in bladder fibrogenesis via an EMT mechanism. Copyright © 2017 the American Physiological Society.

Effects of Curcumin on Tobacco Smoke-induced Hepatic MAPK Pathway Activation and Epithelial-Mesenchymal Transition In Vivo.

PubMed

Liang, Zhaofeng; Wu, Rui; Xie, Wei; Xie, Chunfeng; Wu, Jieshu; Geng, Shanshan; Li, Xiaoting; Zhu, Mingming; Zhu, Weiwei; Zhu, Jianyun; Huang, Cong; Ma, Xiao; Xu, Wenrong; Zhong, Caiyun; Han, Hongyu

2017-08-01

Tobacco smoke is a major risk factor for hepatic cancer. Epithelial-mesenchymal transition (EMT) induced by tobacco smoke is crucially involved in the initiation and development of cancer. Mitogen-activated protein kinase (MAPK) pathways play important roles in tobacco smoke-associated carcinogenesis including EMT process. The chemopreventive effect of curcumin supplementation against cancers has been reported. In this study, we investigated the effects of tobacco smoke on MAPK pathway activation and EMT alterations, and then the preventive effect of curcumin was examined in the liver of BALB/c mice. Our results indicated that exposure of mice to tobacco smoke for 12 weeks led to activation of ERK1/2, JNK, p38 and ERK5 pathways as well as activator protein-1 (AP-1) proteins in liver tissue. Exposure of mice to tobacco smoke reduced the hepatic mRNA and protein expression of the epithelial markers, while the hepatic mRNA and protein levels of the mesenchymal markers were increased. Treatment of curcumin effectively attenuated tobacco smoke-induced activation of ERK1/2 and JNK MAPK pathways, AP-1 proteins and EMT alterations in the mice liver. Our data suggested the protective effect of curcumin in tobacco smoke-triggered MAPK pathway activation and EMT in the liver of BALB/c mice, thus providing new insights into the chemoprevention of tobacco smoke-associated hepatic cancer. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

The roles of connective tissue growth factor and integrin-linked kinase in high glucose-induced phenotypic alterations of podocytes.

PubMed

Dai, Hou-Yong; Zheng, Min; Lv, Lin-Li; Tang, Ri-Ning; Ma, Kun-Ling; Liu, Dan; Wu, Min; Liu, Bi-Cheng

2012-01-01

Emerging evidence has suggested that podocytes undergo epithelial-mesenchymal transition (EMT) in diabetic nephropathy (DN). Connective tissue growth factor (CTGF) and integrin-linked kinase (ILK) are involved in the progression of DN. However, the underlying mechanisms of EMT are not well understood. The study aimed to investigate the roles of CTGF and ILK in high glucose-induced phenotypic alterations of podocytes and determine whether ILK signaling is downstream of CTGF. The epithelial marker of nephrin and the mesenchymal marker of desmin were investigated by real-time RT-PCR and Western blotting. The results demonstrated that podocytes displayed a spreading, arborized morphology in normal glucose, whereas they had a cobblestone morphology in high glucose conditions, accompanied by decreased nephrin expression and increased desmin expression, suggesting podocytes underwent EMT. In response to high glucose, CTGF and ILK expression in podocytes were increased in a dose- and time-dependent manner, whereas the increase did not occur in the osmotic control. Furthermore, the inhibition of CTGF with anti-CTGF antibody prevented the phenotypic transition, as demonstrated by the preservation of epithelial morphology, the suppression of high glucose-induced desmin overexpression and the restoration of nephrin. Of note, the upregulation of ILK induced by high glucose was partially blocked by the inhibition of CTGF. In summary, these findings suggested that CTGF and ILK were involved in high glucose-induced phenotypic alterations of podocytes. ILK acted as a downstream kinase of CTGF and high glucose-induced ILK expression might occur through CTGF-dependent and -independent pathways. Copyright © 2011 Wiley Periodicals, Inc.

Src is activated by the nuclear receptor peroxisome proliferator-activated receptor β/δ in ultraviolet radiation-induced skin cancer.

PubMed

Montagner, Alexandra; Delgado, Maria B; Tallichet-Blanc, Corinne; Chan, Jeremy S K; Sng, Ming K; Mottaz, Hélén; Degueurce, Gwendoline; Lippi, Yannick; Moret, Catherine; Baruchet, Michael; Antsiferova, Maria; Werner, Sabine; Hohl, Daniel; Saati, Talal Al; Farmer, Pierre J; Tan, Nguan S; Michalik, Liliane; Wahli, Walter

2014-01-01

Although non-melanoma skin cancer (NMSC) is the most common human cancer and its incidence continues to rise worldwide, the mechanisms underlying its development remain incompletely understood. Here, we unveil a cascade of events involving peroxisome proliferator-activated receptor (PPAR) β/δ and the oncogene Src, which promotes the development of ultraviolet (UV)-induced skin cancer in mice. UV-induced PPARβ/δ activity, which directly stimulated Src expression, increased Src kinase activity and enhanced the EGFR/Erk1/2 signalling pathway, resulting in increased epithelial-to-mesenchymal transition (EMT) marker expression. Consistent with these observations, PPARβ/δ-null mice developed fewer and smaller skin tumours, and a PPARβ/δ antagonist prevented UV-dependent Src stimulation. Furthermore, the expression of PPARβ/δ positively correlated with the expression of SRC and EMT markers in human skin squamous cell carcinoma (SCC), and critically, linear models applied to several human epithelial cancers revealed an interaction between PPARβ/δ and SRC and TGFβ1 transcriptional levels. Taken together, these observations motivate the future evaluation of PPARβ/δ modulators to attenuate the development of several epithelial cancers.

Src is activated by the nuclear receptor peroxisome proliferator-activated receptor β/δ in ultraviolet radiation-induced skin cancer

PubMed Central

Montagner, Alexandra; Delgado, Maria B; Tallichet-Blanc, Corinne; Chan, Jeremy S K; Sng, Ming K; Mottaz, Hélène; Degueurce, Gwendoline; Lippi, Yannick; Moret, Catherine; Baruchet, Michael; Antsiferova, Maria; Werner, Sabine; Hohl, Daniel; Al Saati, Talal; Farmer, Pierre J; Tan, Nguan S; Michalik, Liliane; Wahli, Walter

2014-01-01

Although non-melanoma skin cancer (NMSC) is the most common human cancer and its incidence continues to rise worldwide, the mechanisms underlying its development remain incompletely understood. Here, we unveil a cascade of events involving peroxisome proliferator-activated receptor (PPAR) β/δ and the oncogene Src, which promotes the development of ultraviolet (UV)-induced skin cancer in mice. UV-induced PPARβ/δ activity, which directly stimulated Src expression, increased Src kinase activity and enhanced the EGFR/Erk1/2 signalling pathway, resulting in increased epithelial-to-mesenchymal transition (EMT) marker expression. Consistent with these observations, PPARβ/δ-null mice developed fewer and smaller skin tumours, and a PPARβ/δ antagonist prevented UV-dependent Src stimulation. Furthermore, the expression of PPARβ/δ positively correlated with the expression of SRC and EMT markers in human skin squamous cell carcinoma (SCC), and critically, linear models applied to several human epithelial cancers revealed an interaction between PPARβ/δ and SRC and TGFβ1 transcriptional levels. Taken together, these observations motivate the future evaluation of PPARβ/δ modulators to attenuate the development of several epithelial cancers. PMID:24203162

Apigenin inhibits NF-κB and snail signaling, EMT and metastasis in human hepatocellular carcinoma.

PubMed

Qin, Yuan; Zhao, Dong; Zhou, Hong-Gang; Wang, Xing-Hui; Zhong, Wei-Long; Chen, Shuang; Gu, Wen-Guang; Wang, Wei; Zhang, Chun-Hong; Liu, Yan-Rong; Liu, Hui-Juan; Zhang, Qiang; Guo, Yuan-Qiang; Sun, Tao; Yang, Cheng

2016-07-05

Apigenin is a naturally occurring compound with anti-inflammatory, antioxidant, and anticancer properties. In this study, we investigated the effects of apigenin on migration and metastasis in experimental human hepatocellular carcinoma (HCC) cell lines in vitro and in vivo. Apigenin dose-dependently inhibited proliferation, migration, and invasion by PLC and Bel-7402 human HCC cells. It also suppressed tumor growth in PLC cell xenografts without altering body weight, thereby prolonging survival. Apigenin reduced Snai1 and NF-κB expression, reversed increases in epithelial-mesenchymal transition (EMT) marker levels, increased cellular adhesion, regulated actin polymerization and cell migration, and inhibited invasion and migration by HCC cells. Apigenin may therefore inhibit EMT by inhibiting the NF-κB/Snail pathway in human HCC.

Apigenin inhibits NF-κB and Snail signaling, EMT and metastasis in human hepatocellular carcinoma

PubMed Central

Zhong, Wei-long; Chen, Shuang; Gu, Wen-guang; Wang, Wei; Zhang, Chun-hong; Liu, Yan-rong; Liu, Hui-juan; Zhang, Qiang; Guo, Yuan-qiang; Sun, Tao; Yang, Cheng

2016-01-01

Apigenin is a naturally occurring compound with anti-inflammatory, antioxidant, and anticancer properties. In this study, we investigated the effects of apigenin on migration and metastasis in experimental human hepatocellular carcinoma (HCC) cell lines in vitro and in vivo. Apigenin dose-dependently inhibited proliferation, migration, and invasion by PLC and Bel-7402 human HCC cells. It also suppressed tumor growth in PLC cell xenografts without altering body weight, thereby prolonging survival. Apigenin reduced Snai1 and NF-κB expression, reversed increases in epithelial-mesenchymal transition (EMT) marker levels, increased cellular adhesion, regulated actin polymerization and cell migration, and inhibited invasion and migration by HCC cells. Apigenin may therefore inhibit EMT by inhibiting the NF-κB/Snail pathway in human HCC. PMID:27203387

AXL Inhibition Suppresses the DNA Damage Response and Sensitizes Cells to PARP Inhibition in Multiple Cancers

PubMed Central

Diao, Lixia; Tong, Pan; Fan, Youhong; Carey, Jason PW; Bui, Tuyen N.; Warner, Steve; Heymach, John V; Hunt, Kelly K; Wang, Jing

2016-01-01

Epithelial to mesenchymal transition (EMT) is associated with a wide range of changes in cancer cells, including stemness, chemo- and radio-resistance and metastasis. The mechanistic role of upstream mediators of EMT has not yet been well characterized. Recently, we showed that non-small cell lung cancers (NSCLCs) that have undergone EMT overexpress AXL, a receptor tyrosine kinase. AXL is also overexpressed in a subset of triple-negative breast cancers (TNBCs) and head and neck squamous cell carcinomas (HNSCCs) and its overexpression has been associated with more aggressive tumor behavior and linked to resistance to chemotherapy, radiation, and targeted therapy. Since the DNA repair pathway is also altered in patient tumor specimens overexpressing AXL, it is hypothesized that modulation of AXL in cells that have undergone EMT will sensitize them to agents targeting the DNA repair pathway. Downregulation or inhibition of AXL directly reversed the EMT phenotype, led to decreased expression of DNA repair genes and diminished efficiency of homologous recombination (HR) and RAD51 foci formation. As a result, AXL inhibition caused a state of HR-deficiency in the cells, making them sensitive to inhibition of the DNA repair protein, PARP1. AXL inhibition synergized with PARP inhibition, leading to apoptotic cell death. AXL expression also associated positively with markers of DNA repair across TNBC, HNSCC and NSCLC patient cohorts. PMID:27671334

FSP1-specific SMAD2 knockout in renal tubular, endothelial, and interstitial cells reduces fibrosis and epithelial-to-mesenchymal transition in murine STZ-induced diabetic nephropathy.

PubMed

Loeffler, Ivonne; Liebisch, Marita; Allert, Stefanie; Kunisch, Elke; Kinne, Raimund W; Wolf, Gunter

2018-04-01

Extracellular matrix deposition during tubulointerstitial fibrosis (TIF), a central pathological process in patients with diabetic nephropathy (DN), is driven by locally activated, disease-relevant myofibroblasts. Myofibroblasts can arise from various cellular sources, e.g., tubular epithelial cells via a process named epithelial-to-mesenchymal transition (EMT). Transforming growth factor beta 1 (TGF-β1) and its downstream Smad signaling play a critical role in both TIF and EMT. Whereas Smad3 is one central mediator, the role of the other prominently expressed variant, Smad2, is not completely understood. In this study, we sought to analyze the role of renal Smad2 in the development of TIF and EMT during streptozotocin-induced DN by using a fibroblast-specific protein 1 (FSP1)-promotor-driven SMAD2 knockout mouse model with decreased tubular, endothelial, and interstitial Smad2 expression. In contrast to wild-type diabetic mice, diabetic SMAD2 knockout mice showed the following features: (1) significantly reduced DN and TIF (shown by KIM1 expression; periodic acid Schiff staining; collagen I and III, fibronectin, and connective tissue growth factor deposition); (2) significantly reduced tubular EMT-like changes (e.g., altered Snail1, E-cadherin, matrix metalloproteinase 2, and vimentin deposition); and (3) significantly decreased expression of myofibroblast markers (α-smooth muscle actin, FSP1). As one mechanism for the protection against diabetes-induced TIF and EMT, decreased Smad3 protein levels and, as a possible consequence, reduced TGF-β1 levels were observed in diabetic SMAD2 knockout mice. Our findings thus support the important role of Smad2 for pro-fibrotic TGF-β/Smad3 signaling in experimental DN.

IL-27 inhibits epithelial-mesenchymal transition and angiogenic factor production in a STAT1-dominant pathway in human non-small cell lung cancer

PubMed Central

2013-01-01

Background Interleukin-27 signaling is mediated by the JAK-STAT pathway via activation of STAT1 and STAT3, which have tumor suppressive and oncogenic activities, respectively. Epithelial–mesenchymal transition (EMT) and angiogenesis are key processes in carcinogenesis. Although IL-27 has been shown to have potent anti-tumor activity in various cancer models, the role of IL-27 in EMT and angiogenesis is poorly understood. In this study, we investigated the role of IL-27 in regulating EMT and angiogenesis through modulation of the STAT pathways in human non-small cell lung carcinoma (NSCLC) cells. Methods STAT activation following IL-27 exposure was measured in human NSCLC cell lines. Expression of epithelial (E-cadherin, γ-catenin) and mesenchymal (N-cadherin, vimentin) markers were assessed by Western blot analysis. Production of pro-angiogenic factors (VEGF, IL-8/CXCL8, CXCL5) were examined by ELISA. Cell motility was examined by an in vitro scratch and transwell migration assays. Selective inhibitors of STAT1 (STAT1 siRNAs) and STAT3 (Stattic) were used to determine whether both STAT1 and STAT3 are required for IL-27 mediated inhibition of EMT and secretion of angiogenic factors. Results Our results demonstrate that IL-27 stimulation in NSCLC resulted in 1) STAT1 and STAT3 activation in a JAK-dependent manner, 2) development of epithelial phenotypes, including a decrease in the expression of a transcriptional repressor for E-cadherin (SNAIL), and mesenchymal marker (vimentin) with a reciprocal increase in the expression of epithelial markers, 3) inhibition of cell migration, and 4) reduced production of pro-angiogenic factors. STAT1 inhibition in IL-27–treated cells reversed the IL-27 effect with resultant increased expression of Snail, vimentin and the pro-angiogenic factors. The inhibition of STAT3 activation had no effect on the development of the epithelial phenotype. Conclusion IL-27 induces mesenchymal to epithelial transition and inhibits the production of pro-angiogenic factors in a STAT1–dominant pathway. These findings highlight the importance of STAT1 in repressing lung carcinogenesis and describe a new anti-tumor mechanism of IL-27. PMID:24274066

Uric acid activates NRLP3 inflammasome in an in-vivo model of epithelial to mesenchymal transition in the kidney.

PubMed

Romero, César Andrés; Remor, Aline; Latini, Alexandra; De Paul, Ana Lucía; Torres, Alicia Inés; Mukdsi, Jorge Humberto

2017-06-01

Uric acid (UA) has been associated with renal fibrosis and progression of chronic kidney disease. However, the underlying mechanisms of this process have still not been identified. Here, we studied the role of the innate imunity receptor NLRP3/ASC in UA induced epithelial-mesenchymal transition (EMT) in kidney. Wistar rats were fed with oxonic acid 2% and UA 2% (OXA + U), OXA + U plus allopurinol (ALL) or regular chow (C) for 7 weeks. We analyzed the presence of EMT markers, the expression of NLRP3, ASC, Caspase-1 and Smad 2/3 molecules and the mitochondrial morphological and functional characteristics. High UA induced renal fibrosis, mild chronic inflammation, as well as morphological and biochemical evidence of EMT. High UA also increased the expression of NLRP3/ASC with activation of both inflammasome related caspase-1 and inflammasome unrelated Smad 2/3 pathways. Ultrastructural co-localization of NLRP3 and Smad 2/3 indicated physical interaction between the two molecules. No morphological or functional changes were found between mitochondria exposed to high UA. In conclusion, kidney epithelial NLRP3/ASC expression was increased in high UA state in rats and both inflammasome related caspase-1 and non-inflammasome related P-Smad 2/3 pathways were associated with the observed EMT, inflammation and fibrosis induced by UA in the kidney.

Estrogen-related receptor α participates transforming growth factor-β (TGF-β) induced epithelial-mesenchymal transition of osteosarcoma cells

PubMed Central

Chen, Yantao; Zhang, Kunshui; Li, Yang; He, Qing

2017-01-01

ABSTRACT Osteosarcoma patients often exhibit pulmonary metastasis, which results in high patient mortality. Understanding the mechanisms of advanced metastasis in osteosarcoma cell is important for the targeted treatment and drug development. Our present study revealed that transforming growth factor-β (TGF-β) treatment can significantly promote the in vitro migration and invasion of human osteosarcoma MG-63 and HOS cells. The loss of epithelial characteristics E-cadherin (E-Cad) and up regulation of mesenchymal markers Vimentin (Vim) suggested TGF-β induced epithelial-mesenchymal transition (EMT) of osteosarcoma cells. TGF-β treatment obviously increased the expression of Snail, a key EMT-related transcription factor, in both MG-63 and HOS cells. Silencing of Snail markedly attenuated TGF-β induced down regulation of E-cad and up regulation of Vim. TGF-β treatment also significantly increased the expression and nuclear translocation of estrogen-related receptors α (ERRα), while had no obvious effect on the expression of ERα, ERβ, or ERRγ. Knock down of ERRα or its inhibitor XCT-790 significantly attenuated TFG-β induced EMT and transcription of Snail in osteosarcoma cells. Collectively, our present study revealed that TGF-β treatment can trigger the EMT of osteosarcoma cells via ERRα/Snail pathways. Our data suggested that ERRα/Snail pathways might be potential therapeutic targets of metastasis of osteosarcoma cells. PMID:27532429

Long Non-Coding RNA (LncRNA) HOXA11-AS Promotes Breast Cancer Invasion and Metastasis by Regulating Epithelial-Mesenchymal Transition

PubMed Central

Li, Wenlei; Jia, Guotao; Qu, Yanwen; Du, Qian; Liu, Baoguo; Liu, Bin

2017-01-01

Background To detect the expression of lncRNA HOXA11-AS and its biological effect in breast cancer. Material/Methods In this study, fluorescent quantitative real-time PCR (qRT-PCR), MTT assay and clone formation assay, flow cytometry, Transwell assay and wound healing assay, immunofluorescence, and Western blot analysis were conducted to detect the expression of lncRNA HOXA11-AS, cell proliferation activity, cell apoptosis rate and cell cycle distribution, the changes of cell invasion and metastasis capacity, and the expressions of molecular markers of epithelial-mesenchymal transition (EMT), respectively. Additionally, a nude mouse metastatic tumor model was established to study the influence of lncRNA HOXA11-AS on invasion and metastasis capacity of breast cancer cells. Results The qRT-PCR experiment results showed that HOXA11-AS expression in breast cancer tissue of 50 patients was relatively higher than that in tissue adjacent to cancer. MTT assay suggested that tumor cell proliferation capacity was suppressed followed by the knockdown of lncRNA HOXA11-AS expression in MDA-MB-231 and MCF-7 cells; flow cytometry results demonstrated that interfering in lncRNA HOXA11-AS could induce tumor cell apoptosis and promote cell cycle progression to be arrested in G1/G0 stage; experiments in vivo/vitro manifested that interfering in lncRNA HOXA11-AS could inhibit tumor cell invasion and migration capacity by affecting the expressions of EMT-related molecular markers (E-cadherin, N-cadherin, Vimentin). Conclusions High expression of lncRNA HOXA11-AS promotes breast cancer invasion and metastasis by affecting EMT, and interfering in lncRAN HOXA11-AS expression provides a theoretical basis and important molecular target for inhibiting the distant metastasis of breast cancer in clinical practice. PMID:28701685

Sustainability of CD24 expression, cell proliferation and migration, cisplatin-resistance, and caspase-3 expression during mesenchymal-epithelial transition induced by the removal of TGF-β1 in A549 lung cancer cells.

PubMed

Kim, Seong-Kwan; Park, Jin-A; Zhang, Dan; Cho, Sang-Hyun; Yi, Hee; Cho, Soo-Min; Chang, Byung-Joon; Kim, Jin-Suk; Shim, Jae-Han; Abd El-Aty, A M; Shin, Ho-Chul

2017-08-01

Epithelial-mesenchymal transition (EMT) is a notable mechanism underlying cancer cell metastasis. Transforming growth factor β1 (TGF-β1) has been used to induce EMT; however, there is a lack of information regarding the role of TGF-β1 in mesenchymal-epithelial transition (MET). In the present study, EMT was induced in A549 lung cancer cells using TGF-β1 (TGF-β1-treated group) and MET was induced sequentially from the TGF-β1-treated group by removing the TGF-β1 (MET/return group). Untreated A549 lung cancer cells were used as a control. Characteristic features, including cancer stem cell markers [cluster of differentiation (CD)24, CD44 and CD133], cell proliferation and migration and diverse intracellular mechanisms, were observed in all groups. Using western blot analysis, the TGF-β1-treated group demonstrated increased vimentin and reduced E-cadherin expression, whereas the MET/return group demonstrated the opposite trend. Among cancer stem cell markers, the population of CD24 low cells was reduced in the TGF-β1-treated group. Furthermore, the G2/M phase cell cycle population, cisplatin-sensitivity, and cell proliferation and migration ability were increased in the TGF-β1-treated group. These features were unaltered in the MET/return group when compared to the TGF-β1-treated group. Immunoblotting revealed an increase in the levels of SMAD3, phosphorylated SMAD3, phosphorylated extracellular signal-regulated kinase and caspase-3, and a decrease in active caspase-3 levels in the TGF-β1-treated group. Increased caspase-3 and reduced active caspase-3 levels were observed in the MET/return group, similar to those in the TGF-β1-treated group; however, levels of other signalling proteins were unchanged compared with the control group. EMT induced by TGF-β1 was not preserved; however, stemness-associated properties (CD24 expression, caspase-3 expression, cell proliferation and cisplatin-resistance) were sustained following removal of TGF-β1.

Emodin Inhibits the Epithelial to Mesenchymal Transition of Epithelial Ovarian Cancer Cells via ILK/GSK-3β/Slug Signaling Pathway

PubMed Central

Lu, Jingjing; Xu, Ying; Wei, Xuan; Zhao, Zhe; Xue, Jing

2016-01-01

Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy. Despite the anticancer capabilities of emodin observed in many cancers, including EOC, the underlying molecular mechanism remains to be elucidated. A crucial link has been discovered between the acquisition of metastatic traits and the epithelial-mesenchymal transition (EMT). The present study aimed to determine whether emodin could inhibit the EMT of EOC cells and explore the underlying mechanism. The CCK-8 assay and transwell assay showed that emodin effectively repressed the abilities of proliferation, invasion, and migration in A2780 and SK-OV-3 cells. The Western blot showed that emodin upregulated epithelial markers (E-cadherin and Claudin) while it downregulated mesenchymal markers (N-cadherin and Vimentin) and transcription factor (Slug) in a dose-dependent fashion. After transfection of siRNA-Slug, both Slug and N-cadherin were downregulated in EOC cells while E-cadherin was upregulated, which was intensified by emodin. Besides, emodin decreased the expression of ILK, p-GSK-3β, β-catenin, and Slug. Transfection of siRNA-ILK also achieved the same effects, which was further strengthened by following emodin treatment. Nevertheless, SB216763, an inhibitor of GSK-3β, could reverse the effects of emodin except for ILK expression. These findings suggest that emodin inhibited the EMT of EOC cells via ILK/GSK-3β/Slug signaling pathway. PMID:28097141

Emodin Inhibits the Epithelial to Mesenchymal Transition of Epithelial Ovarian Cancer Cells via ILK/GSK-3β/Slug Signaling Pathway.

PubMed

Lu, Jingjing; Xu, Ying; Wei, Xuan; Zhao, Zhe; Xue, Jing; Liu, Peishu

2016-01-01

Epithelial ovarian cancer (EOC) is the most lethal gynecologic malignancy. Despite the anticancer capabilities of emodin observed in many cancers, including EOC, the underlying molecular mechanism remains to be elucidated. A crucial link has been discovered between the acquisition of metastatic traits and the epithelial-mesenchymal transition (EMT). The present study aimed to determine whether emodin could inhibit the EMT of EOC cells and explore the underlying mechanism. The CCK-8 assay and transwell assay showed that emodin effectively repressed the abilities of proliferation, invasion, and migration in A2780 and SK-OV-3 cells. The Western blot showed that emodin upregulated epithelial markers (E-cadherin and Claudin) while it downregulated mesenchymal markers (N-cadherin and Vimentin) and transcription factor (Slug) in a dose-dependent fashion. After transfection of siRNA-Slug, both Slug and N-cadherin were downregulated in EOC cells while E-cadherin was upregulated, which was intensified by emodin. Besides, emodin decreased the expression of ILK, p-GSK-3 β , β -catenin, and Slug. Transfection of siRNA-ILK also achieved the same effects, which was further strengthened by following emodin treatment. Nevertheless, SB216763, an inhibitor of GSK-3 β , could reverse the effects of emodin except for ILK expression. These findings suggest that emodin inhibited the EMT of EOC cells via ILK/GSK-3 β /Slug signaling pathway.

Curcumin inhibits tumor epithelial‑mesenchymal transition by downregulating the Wnt signaling pathway and upregulating NKD2 expression in colon cancer cells.

PubMed

Zhang, Zewei; Chen, Haitao; Xu, Chao; Song, Lu; Huang, Lulu; Lai, Yuebiao; Wang, Yuqi; Chen, Hanlu; Gu, Danlin; Ren, Lili; Yao, Qinghua

2016-05-01

Tumor invasion and metastasis are closely associated with epithelial‑mesenchymal transition (EMT). EMT refers to epithelial cells under physiological and pathological conditions that are specific to mesenchymal transition. Curcumin inhibits EMT progression via Wnt signaling. The Wnt signaling pathway is a conservative EMT‑related signaling pathway that is involved in the development of various tumors. In the present study, MTS assays were employed to analyze the proliferation of curcumin‑treated cells. Naked cuticle homolog 2 (NKD2), chemokine receptor 4 (CXCR4) and antibodies associated with EMT were examined in SW620 colorectal cancer cell lines using western blot analysis and real‑time qPCR. NKD2 small‑interfering RNA (siRNA) and CXCR4 expression plasmid was synthesized and transfected into the colorectal cancer cell lines, and NKD2 and CXCR4 expression levels were detected. The results showed that curcumin significantly inhibited the proliferation of colorectal cancer cells and upregulated the expression of NKD2 in SW620 colorectal cancer cells and in the xenograft, resulting in the downregulation of key markers in the Wnt signaling. In addition, the progression of ETM was inhibited due to the overexpression of E‑cadherin as well as the downregulation of vimentin. Curcumin also inhibited tumor metastasis by downregulating the expression of CXCR4 significantly. The results suggested involvement of the NKD2‑Wnt‑CXCR4 signaling pathway in colorectal cancer cells. In addition, curcumin is inhibit this signaling and the development of colorectal cancer.

Metformin inhibits the radiation-induced invasive phenotype of esophageal squamous cell carcinoma.

PubMed

Nakayama, Akira; 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

2016-11-01

Esophageal cancer is one of the most aggressive tumor types because of its invasiveness and metastatic potential. Several reports have described an association between increased invasiveness after ionizing radiation (IR) treatment and epithelial-to-mesenchymal transition (EMT). The biguanide metformin is reported to prevent transforming growth factor-β (TGF-β)-induced EMT and proliferation of cancer. This study examined whether IR induces EMT and promotes the invasive potential of TE-9 esophageal squamous cell carcinoma cells and the effect of metformin on IR-induced EMT. After IR exposure, TE-9 cells showed a spindle-shaped morphology and lost cell-cell adhesion. Immunoblotting showed that IR induced expression of mesenchymal markers (vimentin and N-cadherin), transcription factors (Slug, Snail, and Twist), and matrix metalloproteinases. A scratch wound assay and Matrigel invasion assay showed that IR enhanced the invasive potential and migratory capacity of TE-9 cells. Expression of hypoxia-related factor-1α and TGF-β was increased after IR. IR also induced phosphorylation of Smad2 and Smad3. Metformin inhibited radiation-induced EMT-like morphological changes, and enhanced invasion and migration of TE-9 cells. Metformin inhibited IR-induced phosphorylation of Smad2 and Smad3. Although phosphorylation of AMP-activated protein kinase was enhanced by IR and metformin, phosphorylation of mammalian target of rapamycin was enhanced by IR and suppressed by metformin. These results indicated that metformin suppressed IR-induced EMT via suppression of the TGF-β-Smad phosphorylation pathway, and a part of the non-Smad pathway. Metformin might be useful to prevent IR-induced invasion and metastasis of esophageal squamous cell carcinoma.

TCDD promoted EMT of hFPECs via AhR, which involved the activation of EGFR/ERK signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gao, Zhan; The Fifth Affiliated Hospital, Zhengzhou University, 450052; Bu, Yongjun

2016-05-01

One critical step of second palatal fusion is the newly formed medial epithelia seam (MES) disintegration, which involves apoptosis, epithelial to mesenchymal transition (EMT), and cell migration. Although the environmental toxicant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) produces cleft palate at high rates, little is known about the effects of TCDD exposure on the fate of palatal epithelial cells. By using primary epithelial cells isolated from human fetal palatal shelves (hFPECs), we show that TCDD increased cell proliferation and EMT, as demonstrated by increased the epithelial markers (E-cadherin and cytokeratin14) and enhanced the mesenchymal markers (vimentin and fibronectin), but had no effect on cellmore » migration and apoptosis. TCDD exposure led to a dose-dependent increase in Slug protein expression. Coimmunoprecipitation revealed that TCDD promoted AhR to form a protein complex with Slug. ChIP assay confirmed that TCDD exposure recruited AhR to the xenobiotic responsive element of Slug promoter. Knockdown of AhR by siRNA remarkably weakened TCDD-induced binding of AhR to the XRE promoter of slug, thereby suppressed TCDD-induced vimentin. Further experiment showed that TCDD stimulated EGFR phosphorylation did not influence the TGFβ3/Smad signaling; whereas TCDD increased phosphorylation of ERK1/2 and p38 with no effect on activation of JNK. By using varieties of inhibitors, we confirmed that TCDD promoted proliferation and EMT of hFPECs via activation of EGFR/ERK pathway. These data make a novel contribution to the molecular mechanism of cleft palate by TCDD. - Highlights: • TCDD exposure promoted cell proliferation and EMT of hFPECs; • AhR signaling was activated and required for TCDD-induced EMT; • TCDD-mediated EMT of hFPECs involved the activation of EGFR/ERK signaling; • TCDD exposure had no effect on TGFβ3/Smad pathway.« less

Role of macrophage migration inhibitory factor (MIF) in the effects of oxidative stress on human retinal pigment epithelial cells.

PubMed

Ko, Ji-Ae; Sotani, Yasuyuki; Ibrahim, Diah Gemala; Kiuchi, Yoshiaki

2017-10-01

Proliferative vitreoretinopathy (PVR) is the major cause of treatment failure in individuals who undergo surgery for retinal detachment. The epithelial-mesenchymal transition (EMT) in retinal pigment epithelium (RPE) cells contributes to the pathogenesis of PVR. Oxidative stress is thought to play a role in the progression of retinal diseases including PVR. We have now examined the effects of oxidative stress on the EMT and related processes in the human RPE cell line. We found that H 2 O 2 induced the contraction of RPE cells in a three-dimensional collagen gel. Analysis of a cytokine array revealed that H 2 O 2 specifically increased the release of macrophage migration inhibitory factor (MIF) from RPE cells. Reverse transcription-polymerase chain reaction and immunoblot analyses showed that H 2 O 2 increased the expression of MIF in RPE cells. Immunoblot and immunofluorescence analyses revealed that H 2 O 2 upregulated the expression of α-SMA and vimentin and downregulated that of ZO-1 and N-cadherin. Consistent with these observations, the transepithelial electrical resistance of cell was reduced by exposure to H 2 O 2 . The effects of oxidative stress on EMT-related and junctional protein expression as well as on transepithelial electrical resistance were inhibited by antibodies to MIF, but they were not mimicked by treatment with recombinant MIF. Finally, analysis with a profiling array for mitogen-activated protein kinase signalling revealed that H 2 O 2 specifically induced the phosphorylation of p38 mitogen-activated protein kinase. Our results thus suggest that MIF may play a role in induction of the EMT and related processes by oxidative stress in RPE cells and that it might thereby contribute to the pathogenesis of PVR. Proliferative vitreoretinopathy is a major complication of rhegmatogenous retinal detachment, and both oxidative stress and induction of the EMT in RPE cells are thought to contribute to the pathogenesis of this condition. We have now examined the effects of oxidative stress on the EMT and related processes in the human RPE cell line ARPE19. Our results thus implicate MIF in induction of the EMT and related processes by oxidative stress in RPE cells and the regulated expression of EMT markers. They further suggest that MIF may play an important role in the pathogenesis of PVR. Copyright © 2017 John Wiley & Sons, Ltd.

Functional MUC4 suppress epithelial-mesenchymal transition in lung adenocarcinoma metastasis.

PubMed

Gao, Liuwei; Liu, Jun; Zhang, Bin; Zhang, Hua; Wang, Daowei; Zhang, Tiemei; Liu, Yang; Wang, Changli

2014-02-01

The mucin MUC4 is a high molecular weight membrane-bound transmembrane glycoprotein that is frequently detected in invasive and metastatic cancer. The overexpression of MUC4 is associated with increased risks for several types of cancer. However, the functional role of MUC4 is poorly understood in lung adenocarcinoma. Using antisense-MUC4-RNA transfected adenocarcinoma cells, we discovered that the loss of MUC4 expression results in epithelial-mesenchymal transition (EMT). We found morphological alterations and the repression of the epithelial marker E-cadherin in transfected cells. Additionally, the loss of MUC4 caused the upregulation of the mesenchymal marker vimentin compared to control cells. Using a MUC4-knockdown versus control LTEP xenograft mice model (129/sv mice), we also found that EMT happened in lung tissues of MUC4-knockdown-LTEP xenograft mice. Moreover, antisense-MUC4-RNA transfected cells had a significantly increased cellular migration ability in vitro. The loss of MUC4 also occurred in lung adenocarcinoma patients with lymph node metastases. We further investigated MUC4 and found that it plays a critical role in regulating EMT by modulating β-catenin. Taken together, our study reveals a novel role for MUC4 in suppressing EMT and suggests that the assessment of MUC4 may function as a prognostic biomarker and could be a potential therapeutic target for lung adenocarcinoma metastasis.

3,3′-Diindolylmethane Suppressed Cyprodinil-Induced Epithelial-Mesenchymal Transition and Metastatic-Related Behaviors of Human Endometrial Ishikawa Cells via an Estrogen Receptor-Dependent Pathway

PubMed Central

Kim, Bo-Gyoung; Kim, Jin-Wook; Kim, Soo-Min; Go, Ryeo-Eun; Hwang, Kyung-A

2018-01-01

Cyprodinil (CYP) is a pyrimidine amine fungicide that has been extensively used in agricultural areas. 3,3′-Diindolylmethane (DIM) is a derivative of the dietary phytoestrogen, indole-3-carbinol (I3C), which is derived from cruciferous vegetables and considered to be a cancer-preventive phytonutrient agent. In this study, the effects of CYP and DIM were examined on the cell viability, invasion, and metastasis of human endometrial cancer cells, Ishikawa, via epithelial mesenchymal transition (EMT). CYP increased the level of cell viability of Ishikawa cells compared to DMSO as a control, as did E2. Ishikawa cells lost cell-to-cell contact and obtained a spindle-shaped or fibroblast-like morphology in response to the application of E2 or CYP by the cell morphology assay. In the cell migration and invasion assay, CYP enhanced the ability of migration and invasion of Ishikawa cells, as did E2. E2 and CYP increased the expressions of N-cadherin and Snail proteins, while decreasing the expression of E-cadherin protein as EMT-related markers. In addition, E2 and CYP increased the protein expressions of cathepsin D and MMP-9, metastasis-related markers. Conversely, CYP-induced EMT, cell migration, and invasion were reversed by fulvestrant (ICI 182,780) as an estrogen receptor (ER) antagonist, indicating that CYP exerts estrogenic activity by mediating these processes via an ER-dependent pathway. Similar to ICI 182,780, DIM significantly suppressed E2 and CYP-induced proliferation, EMT, migration, and invasion of Ishikawa cancer cells. Overall, the present study revealed that DIM has an antiestrogenic chemopreventive effect to withdraw the cancer-enhancing effect of E2 and CYP, while CYP has the capacity to enhance the metastatic potential of estrogen-responsive endometrial cancer. PMID:29316692

MUC4 mucin-induced epithelial to mesenchymal transition: a novel mechanism for metastasis of human ovarian cancer cells.

PubMed

Ponnusamy, M P; Lakshmanan, I; Jain, M; Das, S; Chakraborty, S; Dey, P; Batra, S K

2010-10-21

The acquisition of invasiveness in ovarian cancer (OC) is accompanied by the process of epithelial-to-mesenchymal transition (EMT). The MUC4 mucin is overexpressed in ovarian tumors and has a role in the invasiveness of OC cells. The present study was aimed at evaluating the potential involvement of MUC4 in the metastasis of OC cells by inducing EMT. Ectopic overexpression of MUC4 in OC cells (SKOV3-MUC4) resulted in morphological alterations along with a decreased expression of epithelial markers (E-cadherin and cytokeratin (CK)-18) and an increased expression of mesenchymal markers (N-cadherin and vimentin) compared with the control cells (SKOV3-vector). Also, pro-EMT transcription factors TWIST1, TWIST2 and SNAIL showed an upregulation in SKOV3-MUC4 cells. We further investigated the pathways upstream of N-cadherin, such as focal adhesion kinase (FAK), MKK7, JNK1/2 and c-Jun, which were also activated in the SKOV3-MUC4 cells compared with SKOV3-vector cells. Inhibition of phospho-FAK (pFAK) and pJNK1/2 decreased N-cadherin expression in the MUC4-overexpressing cells, which further led to a significant decrease in cellular motility. Knockdown of N-cadherin decreased the activation of extracellular signal-regulated kinase-1/2 (ERK1/2), AKT and matrix metalloproteinase 9 (MMP9), and inhibited the motility in the SKOV3-MUC4 cells. Upon in vivo tumorigenesis and metastasis analysis, the SKOV3-MUC4 cells produced significantly larger tumors and demonstrated a higher incidence of metastasis to distance organs (peritoneal wall, colon, intestine, stomach, lymph nodes, liver and diaphragm). Taken together, our study reveals a novel role for MUC4 in inducing EMT through the upregulation of N-cadherin and promoting metastasis of OC cells.

Epithelial–mesenchymal transition markers expressed in circulating tumor cells in hepatocellular carcinoma patients with different stages of disease

PubMed Central

Li, Y-M; Xu, S-C; Li, J; Han, K-Q; Pi, H-F; Zheng, L; Zuo, G-H; Huang, X-B; Li, H-Y; Zhao, H-Z; Yu, Z-P; Zhou, Z; Liang, P

2013-01-01

The presence of circulating tumor cells (CTCs) in peripheral blood is associated with metastasis and prognosis in hepatocellular carcinoma (HCC) patients. The epithelial–mesenchymal transition (EMT) has a pivotal role in tumor invasion and dissemination. To identify more sensitive biomarkers for evaluating metastasis and prognosis, we investigated the expression of EMT markers, including vimentin, twist, ZEB1, ZEB2, snail, slug and E-cadherin in CTCs, primary HCC tumors and adjacent non-tumoral liver tissues. After isolating viable CTCs from the peripheral blood of HCC patients using asialoglycoprotein receptors (ASGPRs), the CTCs were identified with immunofluorescence staining. CTCs were detected in the peripheral blood obtained from 46 of 60 (76.7%) HCC patients. Triple-immunofluorescence staining showed that twist and vimentin expression could be detected in CTCs obtained from 39 (84.8%) and 37 (80.4%) of the 46 patients, respectively. The expression of both twist and vimentin in CTCs was significantly correlated with portal vein tumor thrombus. Coexpression of twist and vimentin in CTCs could be detected in 32 (69.6%) of the 46 patients and was highly correlated with portal vein tumor thrombus, TNM classification and tumor size. Quantitative fluorescence western blot analysis revealed that the expression levels of E-cadherin, vimentin and twist in HCC tumors were significantly associated with the positivity of isolated CTCs (P=0.013, P=0.012, P=0.009, respectively). However, there was no significant difference in ZEB1, ZEB2, snail and slug expression levels in CTCs, primary HCC tumors and adjacent non-tumoral liver tissues across samples with regard to the clinicopathological parameters. Our results demonstrate that the EMT has a role in promoting the blood-borne dissemination of primary HCC cells, and the twist and vimentin expression levels in CTCs could serve as promising biomarkers for evaluating metastasis and prognosis in HCC patients. PMID:24091674

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Haiwen; Li, Hongliang, E-mail: honglianglity@sina.com; Chen, Qi

Docetaxel efficiency in the therapy of prostate cancer (PCa) patients is limited due to the development of chemoresistance. Recent studies have implied a role of INPP4B in tumor chemoresistance, while the effects of INPP4B on docetaxel resistance in PCa have not been elucidated. In the present study, the docetaxel-resistant human PCa cell lines PC3-DR and DU-145-DR were established from the parental cell lines PC3 and DU-145, and the expression and role of INPP4B in docetaxel-resistant PCa cells were investigated. The results demonstrated that INPP4B expression was significantly downregulated in docetaxel-resistant cells. Overexpression of INPP4B increased the sensitivity to docetaxel andmore » promoted cell apoptosis in PC3-DR and DU-145-DR cells. In addition, INPP4B overexpression downregulated the expression of the mesenchymal markers fibronectin, N-cadherin, and vimentin, and upregulated the expression level of the epithelial maker E-cadherin. Furthermore, INPP4B overexpression markedly inhibited the PI3K/Akt pathway. We also found that IGF-1, the inhibitor of PI3K/Akt, markedly blocked the change in EMT markers induced by overexpression of INPP4B, and reversed the resistance of PC3-DR and DU-145-DR cells to docetaxel, which is sensitized by Flag-INPP4B. In summary, the presented data indicate that INPP4B is crucial for docetaxel-resistant PCa cell survival, potentially by regulating EMT through the PI3K/Akt signaling pathway. - Highlights: • INPP4B is downregulated in docetaxel-resistant PCa cells. • INPP4B inhibits cell proliferation. • INPP4B induces cell apoptosis. • INPP4B inhibits PCa cell EMT.« less

Gastrin-releasing peptide receptor (GRPr) promotes EMT, growth, and invasion in canine prostate cancer.

PubMed

Elshafae, Said M; Hassan, Bardes B; Supsavhad, Wachiraphan; Dirksen, Wessel P; Camiener, Rachael Y; Ding, Haiming; Tweedle, Michael F; Rosol, Thomas J

2016-06-01

The gastrin-releasing peptide receptor (GRPr) is upregulated in early and late-stage human prostate cancer (PCa) and other solid tumors of the mammary gland, lung, head and neck, colon, uterus, ovary, and kidney. However, little is known about its role in prostate cancer. This study examined the effects of a heterologous GRPr agonist, bombesin (BBN), on growth, motility, morphology, gene expression, and tumor phenotype of an osteoblastic canine prostate cancer cell line (Ace-1) in vitro and in vivo. The Ace-1 cells were stably transfected with the human GRPr and tumor cells were grown in vitro and as subcutaneous and intratibial tumors in nude mice. The effect of BBN was measured on cell proliferation, cell migration, tumor growth (using bioluminescence), tumor cell morphology, bone tumor phenotype, and epithelial-mesenchymal transition (EMT) and metastasis gene expression (quantitative RT-PCR). GRPr mRNA expression was measured in primary canine prostate cancers and normal prostate glands. Bombesin (BBN) increased tumor cell proliferation and migration in vitro and tumor growth and invasion in vivo. BBN upregulated epithelial-to-mesenchymal transition (EMT) markers (TWIST, SNAIL, and SLUG mRNA) and downregulated epithelial markers (E-cadherin and β-catenin mRNA), and modified tumor cell morphology to a spindle cell phenotype. Blockade of GRPr upregulated E-cadherin and downregulated VIMENTIN and SNAIL mRNA. BBN altered the in vivo tumor phenotype in bone from an osteoblastic to osteolytic phenotype. Primary canine prostate cancers had increased GRPr mRNA expression compared to normal prostates. These data demonstrated that the GRPr is important in prostate cancer growth and progression and targeting GRPr may be a promising strategy for treatment of prostate cancer. Prostate 76:796-809, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Bax/Tubulin/Epithelial-Mesenchymal Pathways Determine the Efficacy of Silybin Analog HM015k in Colorectal Cancer Cell Growth and Metastasis.

PubMed

Amawi, Haneen; Hussein, Noor A; Ashby, Charles R; Alnafisah, Rawan; Sanglard, Leticia M; Manivannan, Elangovan; Karthikeyan, Chandrabose; Trivedi, Piyush; Eisenmann, Kathryn M; Robey, Robert W; Tiwari, Amit K

2018-01-01

The inhibition of apoptosis, disruption of cellular microtubule dynamics, and over-activation of the epithelial mesenchymal transition (EMT), are involved in the progression, metastasis, and resistance of colorectal cancer (CRC) to chemotherapy. Therefore, the design of a molecule that can target these pathways could be an effective strategy to reverse CRC progression and metastasis. In this study, twelve novel silybin derivatives, HM015a-HM015k (15a-15k) and compound 17, were screened for cytotoxicity in CRC cell lines. Compounds HM015j and HM015k (15k and 15j) significantly decreased cell proliferation, inhibited colony formation, and produced cell cycle arrest in CRC cells. Furthermore, 15k significantly induced the formation of reactive oxygen species and apoptosis. It induced the cleavage of the intrinsic apoptotic protein (Bax p21) to its more efficacious fragment, p18. Compound 15k also inhibited tubulin expression and disrupted its structure. Compound 15k significantly decreased metastatic LOVO cell migration and invasion. Furthermore, 15k reversed mesenchymal morphology in HCT116 and LOVO cells. Additionally, 15k significantly inhibited the expression of the mesenchymal marker N-cadherin and upregulated the expression of the epithelial marker, E-cadherin. Compound 15k inhibited the expression of key proteins known to induce EMT (i.e., DVL3, β-catenin, c-Myc) and upregulated the anti-metastatic protein, cyclin B1. Overall, in vitro , 15k significantly inhibited CRC progression and metastasis by inhibiting apoptosis, tubulin activity and the EMT pathways. Overall, these data suggest that compound 15k should be tested in vivo in a CRC animal model for further development.

Leucine-rich repeat-containing G protein-coupled receptor 4 (Lgr4) is necessary for prostate cancer metastasis via epithelial-mesenchymal transition.

PubMed

Luo, Weijia; Tan, Peng; Rodriguez, Melissa; He, Lian; Tan, Kunrong; Zeng, Li; Siwko, Stefan; Liu, Mingyao

2017-09-15

Prostate cancer is a highly penetrant disease among men in industrialized societies, but the factors regulating the transition from indolent to aggressive and metastatic cancer remain poorly understood. We found that men with prostate cancers expressing high levels of the G protein-coupled receptor LGR4 had a significantly shorter recurrence-free survival compared with patients with cancers having low LGR4 expression. LGR4 expression was elevated in human prostate cancer cell lines with metastatic potential. We therefore generated a novel transgenic adenocarcinoma of the mouse prostate (TRAMP) mouse model to investigate the role of Lgr4 in prostate cancer development and metastasis in vivo TRAMP Lgr4 -/- mice exhibited an initial delay in prostate intraepithelial neoplasia formation, but the frequency of tumor formation was equivalent between TRAMP and TRAMP Lgr4 -/- mice by 12 weeks. The loss of Lgr4 significantly improved TRAMP mouse survival and dramatically reduced the occurrence of lung metastases. LGR4 knockdown impaired the migration, invasion, and colony formation of DU145 cells and reversed epithelial-mesenchymal transition (EMT), as demonstrated by up-regulation of E-cadherin and decreased expression of the EMT transcription factors ZEB, Twist, and Snail. Overexpression of LGR4 in LNCaP cells had the opposite effects. Orthotopic injection of DU145 cells stably expressing shRNA targeting LGR4 resulted in decreased xenograft tumor size, reduced tumor EMT marker expression, and impaired metastasis, in accord with our findings in TRAMP Lgr4 -/- mice. In conclusion, we propose that Lgr4 is a key protein necessary for prostate cancer EMT and metastasis. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

High glucose induces podocyte epithelial-to-mesenchymal transition by demethylation-mediated enhancement of MMP9 expression

PubMed Central

Ling, Li; Chen, Libo; Zhang, Changning; Gui, Shuyan; Zhao, Haiyan; Li, Zhengzhang

2018-01-01

Abnormal expression of matrix metalloproteinase 9 (MMP9) is correlated with podocyte epithelial-to-mesenchymal transition (EMT) in diabetic nephropathy (DN). However, the mechanisms underlying this process are not well defined. Site-specific demethylation may sustain high expression levels of target genes. In the present study, in order to investigate the association between DNA demethylation of MMP9 promoter and podocyte EMT in DN, human podocytes were cultured in high-glucose (HG) medium and a rat model of DN was established by intraperitoneal injection of streptozotocin (STZ) to determine whether site-specific demethylation of the MMP9 promoter was involved in regulating podocyte EMT in DN. The MTT assay was used to assess the effects of HG culture on the growth of podocytes, and the demethylation status of the MMP9 promoter was assessed by bisulfite sequencing polymerase chain reaction. mRNA and protein expression levels of MMP9, α-smooth muscle actin (α-SMA), podocalyxin and fibronectin-1 in podocytes were assessed by reverse transcription-quantitative PCR (RT-qPCR) and western blot analyses. The results demonstrated that HG treatment up regulated the expression of MMP9, α-SMA and fibronectin-1, but down regulated the expression of podocalyxin in podocytes. The MMP9 promoter region was revealed to contain a variety of demethylated CpG sites, and HG treatment reduced the rate of MMP9 promotermethylation, which, in turn, enhanced its promoter activity. In summary, these data suggested that demethylation of the MMP9 promoter may serve an important role in podocyte EMT in DN. The demethylation status of the MMP9 promoter maybe used as an important prognostic marker of DN in clinic. PMID:29436620

The effects of pleiotrophin in proliferative vitreoretinopathy.

PubMed

Ding, Xue; Bai, Yujing; Zhu, Xuemei; Li, Tianqi; Jin, Enzhong; Huang, Lvzhen; Yu, Wenzhen; Zhao, Mingwei

2017-05-01

The purpose of our study was to investigate the effects of pleiotrophin (PTN) in proliferative vitreoretinopathy (PVR) both in vitro and in vivo. Immunofluorescence was used to observe the PTN expression in periretinal membrane samples from patients with PVR and controls. ARPE-19 cells were exposed to TGF-β1. The epithelial-to-mesenchymal transition (EMT) of the ARPE-19 cells was confirmed by observed morphological changes and the increased expression of α-SMA and fibronectin at both the mRNA and protein levels. We used specific small interfering (si)RNA to knock down the expression of PTN. The subsequent effects of PTN inhibition were assessed with regard to the EMT, migration, proliferation, cytoskeletal arrangement, TGF-β signaling, PTN signaling, integral tight junction protein expression (e.g., claudin-1 and occludin), and p38 MAPK and p-p38 MAPK levels. Additionally, a PVR rat model was established by the intravitreal injection of ARPE-19 cells transfected with PTN-siRNA and was evaluated accordingly. PTN was highly expressed in PVR membranes compared to controls. PTN knockdown attenuated the TGF-β1-induced migration, proliferation, cytoskeletal rearrangement, and expression of EMT markers such as α-SMA and fibronectin in the ARPE-19 cells, and these effects may have been mediated through p38 MAPK signaling pathway activation. PTN silencing inhibited the up-regulation of claudin-1 and occludin stimulated by TGF-β1, and PTN knockdown inhibited the proliferative aspects of severe PVR in vivo. PTN is involved in the process of EMT induced by TGF-β1 in human ARPE-19 cells in vitro, and PTN knockdown attenuated the progression of experimental PVR in vivo. These findings provide new insights into the pathogenesis of PVR.

Protons sensitize epithelial cells to mesenchymal transition.

PubMed

Wang, Minli; Hada, Megumi; Saha, Janapriya; Sridharan, Deepa M; Pluth, Janice M; Cucinotta, Francis A

2012-01-01

Proton radiotherapy has gained more favor among oncologists as a treatment option for localized and deep-seated tumors. In addition, protons are a major constituent of the space radiation astronauts receive during space flights. The potential for these exposures to lead to, or enhance cancer risk has not been well studied. Our objective is to study the biological effects of low energy protons on epithelial cells and its propensity to enhance transforming growth factor beta 1 (TGFβ1)-mediated epithelial-mesenchymal transition (EMT), a process occurring during tumor progression and critical for invasion and metastasis. Non-transformed mink lung epithelial cells (Mv1Lu) and hTERT- immortalized human esophageal epithelial cells (EPC) were used in this study. EMT was identified by alterations in cell morphology, EMT-related gene expression changes determined using real-time PCR, and EMT changes in specific cellular markers detected by immunostaining and western blotting. Although TGFβ1 treatment alone is able to induce EMT in both Mv1Lu and EPC cells, low energy protons (5 MeV) at doses as low as 0.1 Gy can enhance TGFβ1 induced EMT. Protons alone can also induce a mild induction of EMT. SD208, a potent TGFβ Receptor 1 (TGFβR1) kinase inhibitor, can efficiently block TGFβ1/Smad signaling and attenuate EMT induction. We suggest a model for EMT after proton irradiation in normal and cancerous tissue based on our results that showed that low and high doses of protons can sensitize normal human epithelial cells to mesenchymal transition, more prominently in the presence of TGFβ1, but also in the absence of TGFβ1.

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.

PubMed

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 (P<0.05). We found significantly more positively expressed CTGF protein in ESCC tissues was than in normal adjacent esophageal tissues (P<0.01). Dual luciferase reporter gene assay showed that miR-145 can specifically bind with the 3'UTR of CTGF and significantly inhibit the luciferase activity by 55% (P<0.01). Up-regulation of miR-145 or down-regulation of CTGF can suppress the proliferation, migration, invasion, and EMT process of ESCC cells. CONCLUSIONS MiR-145 was significantly down-regulated in ESCC tissues 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.

Epithelial–mesenchymal transition during oncogenic transformation induced by hexavalent chromium involves reactive oxygen species-dependent mechanism in lung epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ding, Song-Ze, E-mail: dingsongze@hotmail.com; Graduate Center for Toxicology, College of Medicine, University of Kentucky, Lexington, KY 40536; Yang, Yu-Xiu

2013-05-15

Hexavalent chromium [Cr(VI)] is an important human carcinogen associated with pulmonary diseases and lung cancer. Exposure to Cr(VI) induces DNA damage, cell morphological change and malignant transformation in human lung epithelial cells. Despite extensive studies, the molecular mechanisms remain elusive, it is also not known if Cr(VI)-induced transformation might accompany with invasive properties to facilitate metastasis. We aimed to study Cr(VI)-induced epithelial–mesenchymal transition (EMT) and invasion during oncogenic transformation in lung epithelial cells. The results showed that Cr(VI) at low doses represses E-cadherin mRNA and protein expression, enhances mesenchymal marker vimentin expression and transforms the epithelial cell into fibroblastoid morphology.more » Cr(VI) also increases cell invasion and promotes colony formation. Further studies indicated that Cr(VI) uses multiple mechanisms to repress E-cadherin expression, including activation of E-cadherin repressors such as Slug, ZEB1, KLF8 and enhancement the binding of HDAC1 in E-cadherin gene promoter, but DNA methylation is not responsible for the loss of E-cadherin. Catalase reduces Cr(VI)-induced E-cadherin and vimentin protein expression, attenuates cell invasion in matrigel and colony formation on soft agar. These results demonstrate that exposure to a common human carcinogen, Cr(VI), induces EMT and invasion during oncogenic transformation in lung epithelial cells and implicate in cancer metastasis and prevention. - Graphical abstract: Epithelial–mesenchymal transition during oncogenic transformation induced by hexavalent chromium involves reactive oxygen species-dependent mechanisms in lung epithelial cells. - Highlights: • We study if Cr(VI) might induce EMT and invasion in epithelial cells. • Cr(VI) induces EMT by altering E-cadherin and vimentin expression. • It also increases cell invasion and promotes oncogenic transformation. • Catalase reduces Cr(VI)-induced EMT, invasion and transformation.« less

SOX5 predicts poor prognosis in lung adenocarcinoma and promotes tumor metastasis through epithelial-mesenchymal transition

PubMed Central

Chen, Xin; Fu, Yufei; Xu, Hongfei; Teng, Peng; Xie, Qiong; Zhang, Yiran; Yan, Caochong; Xu, Yiqiao; Li, Chunqi; Zhou, Jianying; Ni, Yiming; Li, Weidong

2018-01-01

Lung cancer is the leading cause of cancer-related death worldwide. Epithelial-mesenchymal transition (EMT) promotes lung cancer progression and metastasis, especially in lung adenocarcinoma. Sex determining region Y-box protein 5 (SOX5) is known to stimulate the progression of various cancers. Here, we used immunohistochemical analysis to reveal that SOX5 levels were increased in 90 lung adenocarcinoma patients. The high SOX5 expression in lung adenocarcinoma and non-tumor counterparts correlated with the patients’ poor prognosis. Inhibiting SOX5 expression attenuated metastasis and progression in lung cancer cells, while over-expressing SOX5 accelerated lung adenocarcinoma progression and metastasis via EMT. An in vivo zebrafish xenograft cancer model also showed SOX5 knockdown was followed by reduced lung cancer cell proliferation and metastasis. Our results indicate SOX5 promotes lung adenocarcinoma tumorigenicity and can be a novel diagnosis and prognosis marker of the disease. PMID:29541384

Hand1 overexpression inhibits medulloblastoma metastasis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Asuthkar, Swapna; Guda, Maheedhara R.; Martin, Sarah E.

2016-08-19

Medulloblastoma (MB) is the most frequent malignant pediatric brain tumor. Current treatment includes surgery, radiation and chemotherapy. However, ongoing treatment in patients is further classified according to the presence or absence of metastasis. Since metastatic medulloblastoma are refractory to current treatments, there is need to identify novel biomarkers that could be used to reduce metastatic potential, and more importantly be targeted therapeutically. Previously, we showed that ionizing radiation-induced uPAR overexpression is associated with increased accumulation of β-catenin in the nucleus. We further demonstrated that uPAR protein act as cytoplasmic sequestration factor for a novel basic helix-loop-helix transcription factor, Hand1. Amongmore » the histological subtypes classical and desmoplastic subtypes account for the majority while large cell/anaplastic variant is most commonly associated with metastatic disease. In this present study using immunohistochemical approach and patient data mining for the first time, we demonstrated that Hand1 expression is observed to be downregulated in all the subtypes of medulloblastoma. Previously we showed that Hand1 overexpression regulated medulloblastoma angiogenesis and here we investigated the role of Hand1 in the context of Epithelial-Mesenchymal Transition (EMT). Moreover, UW228 and D283 cells overexpressing Hand1 demonstrated decreased-expression of mesenchymal markers (N-cadherin, β-catenin and SOX2); metastatic marker (SMA); and increased expression of epithelial marker (E-cadherin). Strikingly, human pluripotent stem cell antibody array showed that Hand1 overexpression resulted in substantial decrease in pluripotency markers (Nanog, Oct3/4, Otx2, Flk1) suggesting that Hand1 expression may be essential to attenuate the EMT and our findings underscore a novel role for Hand1 in medulloblastoma metastasis. - Highlights: • Hand1 expression is downregulated in Medulloblastoma. • Hand1 over expression reduce the expression of signaling from WNT, SHH and Group 3 medulloblastoma subgroups. • Hand1 overexpression reduced metastatic abilities by reducing the expression of β-catenin and N-cadherin.« less

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kikuta, Kazuhiro; Masamune, Atsushi, E-mail: amasamune@med.tohoku.ac.jp; Watanabe, Takashi

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 ismore » 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 by treatment with anti-TGF-{beta}-neutralizing antibody, excluding a central role of TGF-{beta} in this process. In conclusion, PSCs promoted EMT in pancreatic cancer cells suggesting a novel mechanism by which PSCs contribute to the aggressive behavior of pancreatic cancer cells.« less

Glioma progression through the prism of heat shock protein mediated extracellular matrix remodeling and epithelial to mesenchymal transition.

PubMed

Rajesh, Y; Biswas, Angana; Mandal, Mahitosh

2017-10-15

Glial tumor is one of the intrinsic brain tumors with high migratory and infiltrative potential. This essentially contributes to the overall poor prognosis by circumvention of conventional treatment regimen in glioma. The underlying mechanism in gliomagenesis is bestowed by two processes- Extracellular matrix (ECM) Remodeling and Epithelial to mesenchymal transition (EMT). Heat Shock Family of proteins (HSPs), commonly known as "molecular chaperons" are documented to be upregulated in glioma. A positive correlation also exists between elevated expression of HSPs and invasive capacity of glial tumor. HSPs overexpression leads to mutational changes in glioma, which ultimately drive cells towards EMT, ECM modification, malignancy and invasion. Differential expression of HSPs - a factor providing cytoprotection to glioma cells, also contributes towards its radioresistance /chemoresistance. Various evidences also display upregulation of EMT and ECM markers by various heat shock inducing proteins e.g. HSF-1. The aim of this review is to study in detail the role of HSPs in EMT and ECM leading to radioresistance/chemoresistance of glioma cells. The existing treatment regimen for glioma could be enhanced by targeting HSPs through immunotherapy, miRNA and exosome mediated strategies. This could be envisaged by better understanding of molecular mechanisms underlying glial tumorigenesis in relation to EMT and ECM remodeling under HSPs influence. Our review might showcase fresh potential for the development of next generation therapeutics for effective glioma management. Copyright © 2017 Elsevier Inc. All rights reserved.

PRRX2 as a novel TGF-β-induced factor enhances invasion and migration in mammary epithelial cell and correlates with poor prognosis in breast cancer.

PubMed

Juang, Yu-Lin; Jeng, Yung-Ming; Chen, Chi-Long; Lien, Huang-Chun

2016-12-01

TGF-β and cancer progression share a multifaceted relationship. Despite the knowledge of TGF-β biology in the development of cancer, several factors that mediate the cancer-promoting role of TGF-β continue to be identified. This study aimed to identify and characterise novel factors potentially related to TGF-β-mediated tumour aggression in breast cells. We treated the human mammary epithelial cell line MCF10A with TGF-β and identified TGF-β-dependent upregulation of PRRX2, the gene encoding paired-related homeobox 2 transcription factor. Overexpression of PRRX2 enhanced migration, invasion and anchorage-independent growth of MCF10A cells and induced partial epithelial mesenchymal transition (EMT), as determined by partial fibroblastoid morphology of cells, upregulation of EMT markers and partially disrupted acinar structure in a three-dimensional culture. We further identified PLAT, the gene encoding tissue-type plasminogen activator (tPA), as the highest differentially expressed gene in PRRX2-overexpressing MCF10A cells, and demonstrated direct binding and transactivation of the PLAT promoter by PRRX2. Furthermore, PLAT knockdown inhibited PRRX2-mediated enhanced migration and invasion, suggesting that tPA may mediate PRRX2-induced migration and invasion. Finally, the significant correlation of PRRX2 expression with poor survival in 118 primary breast tumour samples (P = 0.027) and the increased PRRX2 expression in metaplastic breast carcinoma samples, which is pathogenetically related to EMT, validated the biological importance of PRRX2-enhanced migration and invasion and PRRX2-induced EMT. Thus, our data suggest that upregulation of PRRX2 may be a mechanism contributing to TGF-β-induced invasion and EMT in breast cancer. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Pancreatic Cancer Cells Enhance the Ability of Collagen Internalization during Epithelial–Mesenchymal Transition

PubMed Central

Ikenaga, Naoki; Ohuchida, Kenoki; Mizumoto, Kazuhiro; Akagawa, Shin; Fujiwara, Kenji; Eguchi, Daiki; Kozono, Shingo; Ohtsuka, Takao; Takahata, Shunichi; Tanaka, Masao

2012-01-01

Background Extracellular matrix (ECM) remodeling is predominantly mediated by fibroblasts using intracellular and extracellular pathways. Although it is well known that extracellular degradation of the ECM by proteases derived from cancer cells facilitates cellular invasion, the intracellular degradation of ECM components by cancer cells has not been clarified. The aim of this study was to characterize collagen internalization, which is the initial step of the intracellular degradation pathway in pancreatic cancer cells, in light of epithelial–mesenchymal transition (EMT). Methodology/Principal Findings We analyzed the function of collagen internalization in two pancreatic cancer cell lines, SUIT-2 and KP-2, and pancreatic stellate cells (PSCs) using Oregon Green 488-gelatin. PSCs had a strong ability for collagen uptake, and the pancreatic cancer cells also internalized collagen although less efficiently. The collagen internalization abilities of SUIT-2 and KP-2 cells were promoted by EMT induced by human recombinant transforming growth factor β1 (P<0.05). Expression of Endo180, a collagen uptake receptor, was high in mesenchymal pancreatic cancer cell lines, as determined by EMT marker expression (P<0.01). Quantitative RT-PCR and western blot analyses showed that Endo180 expression was also increased by EMT induction in SUIT-2 and KP-2 cells. Endo180 knockdown by RNA interference attenuated the collagen uptake (P<0.01) and invasive abilities (P<0.05) of SUIT-2 and KP-2 cells. Conclusions/Significance Pancreatic cancer cells are capable of collagen internalization, which is enhanced by EMT. This ECM clearance system may be a novel mechanism for cellular invasion and a potential therapeutic target in pancreatic cancer. PMID:22792318

Lysyl oxidase-like 2 (LOXL2), a new regulator of cell polarity required for metastatic dissemination of basal-like breast carcinomas

PubMed Central

Moreno-Bueno, Gema; Salvador, Fernando; Martín, Alberto; Floristán, Alfredo; Cuevas, Eva P; Santos, Vanesa; Montes, Amalia; Morales, Saleta; Castilla, Maria Angeles; Rojo-Sebastián, Alejandro; Martínez, Alejandra; Hardisson, David; Csiszar, Katalin; Portillo, Francisco; Peinado, Héctor; Palacios, José; Cano, Amparo

2011-01-01

Basal-like breast carcinoma is characterized by the expression of basal/myoepithelial markers, undifferentiated phenotype, highly aggressive behaviour and frequent triple negative status (ESR−, PR−, Her2neu−). We have previously shown that epithelial–mesenchymal transition (EMT) occurs in basal-like breast tumours and identified Lysyl-oxidase-like 2 (LOXL2) as an EMT player and poor prognosis marker in squamous cell carcinomas. We now show that LOXL2 mRNA is overexpressed in basal-like human breast carcinomas. Breast carcinoma cell lines with basal-like phenotype show a specific cytoplasmic/perinuclear LOXL2 expression, and this subcellular distribution is significantly associated with distant metastatic incidence in basal-like breast carcinomas. LOXL2 silencing in basal-like carcinoma cells induces a mesenchymal-epithelial transition (MET) associated with a decrease of tumourigenicity and suppression of metastatic potential. Mechanistic studies indicate that LOXL2 maintains the mesenchymal phenotype of basal-like carcinoma cells by a novel mechanism involving transcriptional downregulation of Lgl2 and claudin1 and disorganization of cell polarity and tight junction complexes. Therefore, intracellular LOXL2 is a new candidate marker of basal-like carcinomas and a target to block metastatic dissemination of this aggressive breast tumour subtype. PMID:21732535

MicroRNA-33b, upregulated by EF24, a curcumin analog, suppresses the epithelial-to-mesenchymal transition (EMT) and migratory potential of melanoma cells by targeting HMGA2.

PubMed

Zhang, Pu; Bai, Huiyuan; Liu, Gentao; Wang, Heyong; Chen, Feng; Zhang, Baoshun; Zeng, Panying; Wu, Chengxiang; Peng, Cong; Huang, Changjin; Song, Yang; Song, Erqun

2015-05-05

Diphenyl difluoroketone (EF24), a curcumin analog, exhibits potent anti-tumor activities by arresting cell cycle and inducing apoptosis. However, the efficacy and modes of action of EF24 on melanoma metastasis remain elusive. In this study, we found that at non-cytotoxic concentrations, EF24 suppressed cell motility and epithelial-to-mesenchymal Transition (EMT) of melanoma cell lines, Lu1205 and A375. EF24 also suppressed HMGA2 expression at mRNA and protein levels. miR-33b directly bound to HMGA2 3' untranslated region (3'-UTR) to suppress its expression as measured by dual-luciferase assay. EF24 increased expression of E-cadherin and decreased STAT3 phosphorylation and expression of the mesenchymal markers, vimentin and N-cadherin. miR-33b inhibition or HMGA2 overexpression reverted EF24-mediated suppression of EMT phenotypes. In addition, EF24 modulated the HMGA2-dependent actin stress fiber formation, focal adhesion assembly and FAK, Src and RhoA activation by targeting miR-33b. Thus, the results suggest that EF24 suppresses melanoma metastasis via upregulating miR-33b and concomitantly reducing HMGA2 expression. The observed activities of EF24 support its further evaluation as an anti-metastatic agent in melanoma therapy. Copyright © 2015. Published by Elsevier Ireland Ltd.

Hand1 overexpression inhibits medulloblastoma metastasis.

PubMed

Asuthkar, Swapna; Guda, Maheedhara R; Martin, Sarah E; Antony, Reuben; Fernandez, Karen; Lin, Julian; Tsung, Andrew J; Velpula, Kiran K

2016-08-19

Medulloblastoma (MB) is the most frequent malignant pediatric brain tumor. Current treatment includes surgery, radiation and chemotherapy. However, ongoing treatment in patients is further classified according to the presence or absence of metastasis. Since metastatic medulloblastoma are refractory to current treatments, there is need to identify novel biomarkers that could be used to reduce metastatic potential, and more importantly be targeted therapeutically. Previously, we showed that ionizing radiation-induced uPAR overexpression is associated with increased accumulation of β-catenin in the nucleus. We further demonstrated that uPAR protein act as cytoplasmic sequestration factor for a novel basic helix-loop-helix transcription factor, Hand1. Among the histological subtypes classical and desmoplastic subtypes account for the majority while large cell/anaplastic variant is most commonly associated with metastatic disease. In this present study using immunohistochemical approach and patient data mining for the first time, we demonstrated that Hand1 expression is observed to be downregulated in all the subtypes of medulloblastoma. Previously we showed that Hand1 overexpression regulated medulloblastoma angiogenesis and here we investigated the role of Hand1 in the context of Epithelial-Mesenchymal Transition (EMT). Moreover, UW228 and D283 cells overexpressing Hand1 demonstrated decreased-expression of mesenchymal markers (N-cadherin, β-catenin and SOX2); metastatic marker (SMA); and increased expression of epithelial marker (E-cadherin). Strikingly, human pluripotent stem cell antibody array showed that Hand1 overexpression resulted in substantial decrease in pluripotency markers (Nanog, Oct3/4, Otx2, Flk1) suggesting that Hand1 expression may be essential to attenuate the EMT and our findings underscore a novel role for Hand1 in medulloblastoma metastasis. Copyright © 2016 Elsevier Inc. All rights reserved.

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

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miura, Yuka; Hagiwara, Natsumi; Radisky, Derek C.

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.more » 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.« less

Substrate stiffness affects epithelial-mesenchymal transition of cervical cancer cells through miR-106b and its target protein DAB2.

PubMed

Piao, Jinlan; You, Ke; Guo, Yanli; Zhang, Youyi; Li, Zijian; Geng, Li

2017-06-01

The effects of different substrate stiffness were investigated on epithelial-mesenchymal transition (EMT) of cervical cancer cell lines and the role of miR-106b and its target protein DAB2 therein. Cervical cancer cell lines HeLa and SiHa were cultured on artificial substrates with different stiffness prepared using different ratios of acrylamide and bis-acrylamide. Changes of microRNA profiles were detected using microRNA chip analysis, and the expression levels of EMT-related markers E-cadherin and vimentin were detected using western blotting and real-time PCR. In addition, the effects of miR-106b overexpression as well as miR-106b and DAB2 knockdown on expression of E-cadherin and vimentin were also examined using western blotting and real-time PCR. The results showed that i) cervical cancer cell lines SiHa and HeLa cultured on substrate with stiffness of 20 kPa had the strongest EMT ability, showed the highest levels of vimentin and lowest levels of E-cadherin, compared with cells cultured on substrate with stiffness of 1 kPa; ii) miR-106b knockdown reversed the effects of substrate stiffness on EMT of cervical cancer cells, while miR-106 overexpression and DAB2 knockdown induced EMT of cervical cancer cells cultured on substrate with stiffness of 20 kPa. Overall, the results indicated that substrate stiffness could regulate EMT of cervical cancer cell lines HeLa and SiHa at least partially through miR-106b and its downstream target DAB2.

An EMT–Driven Alternative Splicing Program Occurs in Human Breast Cancer and Modulates Cellular Phenotype

PubMed Central

Flytzanis, Nicholas C.; Balsamo, Michele; Condeelis, John S.; Oktay, Maja H.; Burge, Christopher B.; Gertler, Frank B.

2011-01-01

Epithelial-mesenchymal transition (EMT), a mechanism important for embryonic development, plays a critical role during malignant transformation. While much is known about transcriptional regulation of EMT, alternative splicing of several genes has also been correlated with EMT progression, but the extent of splicing changes and their contributions to the morphological conversion accompanying EMT have not been investigated comprehensively. Using an established cell culture model and RNA–Seq analyses, we determined an alternative splicing signature for EMT. Genes encoding key drivers of EMT–dependent changes in cell phenotype, such as actin cytoskeleton remodeling, regulation of cell–cell junction formation, and regulation of cell migration, were enriched among EMT–associated alternatively splicing events. Our analysis suggested that most EMT–associated alternative splicing events are regulated by one or more members of the RBFOX, MBNL, CELF, hnRNP, or ESRP classes of splicing factors. The EMT alternative splicing signature was confirmed in human breast cancer cell lines, which could be classified into basal and luminal subtypes based exclusively on their EMT–associated splicing pattern. Expression of EMT–associated alternative mRNA transcripts was also observed in primary breast cancer samples, indicating that EMT–dependent splicing changes occur commonly in human tumors. The functional significance of EMT–associated alternative splicing was tested by expression of the epithelial-specific splicing factor ESRP1 or by depletion of RBFOX2 in mesenchymal cells, both of which elicited significant changes in cell morphology and motility towards an epithelial phenotype, suggesting that splicing regulation alone can drive critical aspects of EMT–associated phenotypic changes. The molecular description obtained here may aid in the development of new diagnostic and prognostic markers for analysis of breast cancer progression. PMID:21876675

CD44S-hyaluronan interactions protect cells resulting from EMT against anoikis

PubMed Central

Cieply, Benjamin; Koontz, Colton; Frisch, Steven M.

2016-01-01

The detachment of normal epithelial cells from matrix triggers an apoptotic response known as anoikis, during homeostatic turnover. Metastatic tumor cells evade anoikis, by mechanisms that are only partly characterized. In particular, the epithelial–mesenchymal transition (EMT) in a subset of invasive tumor cells confers anoikis-resistance. In some cases, EMT up-regulates the cancer stem cell marker CD44S and the enzyme hyaluronic acid synthase-2 (HAS2). CD44S is the major receptor for hyaluronan in the extracellular matrix. Herein, we demonstrate that CD44S, unlike the CD44E isoform expressed in normal epithelial cells, contributes to the protection against anoikis. This protection requires the interaction of CD44S with hyaluronan (HA). CD44S–HA interaction is proposed to play an important role in tumor metastasis through enhanced cell survival under detached conditions. PMID:25937513

Advanced Oxidation Protein Products Induce Epithelial-Mesenchymal Transition of Intestinal Epithelial Cells via a PKC δ-Mediated, Redox-Dependent Signaling Pathway.

PubMed

Xu, Xiaoping; Sun, Shibo; Xie, Fang; Ma, Juanjuan; Tang, Jing; He, Shuying; Bai, Lan

2017-07-01

Epithelial-mesenchymal transition (EMT) has been considered a fundamental mechanism in complications of Crohn's disease (CD), especially intestinal fibrosis. However, the mechanism underlying EMT regulation in intestinal fibrosis remains unclear. This study aimed to investigate the role of advanced oxidation protein products (AOPPs) in the occurrence of intestinal EMT. AOPPs accumulated in CD tissues and were associated with EMT marker expression in fibrotic lesions from CD patients. Challenge with AOPPs induced intestinal epithelial cell (IEC) phenotype transdifferentiation, fibroblast-like phenotype acquisition, and production of extracellular matrix, both in vitro and in vivo. The effect of AOPPs was mainly mediated by a protein kinase C (PKC) δ-mediated redox-dependent pathway, including phosphorylation of PKC δ, recruitment of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, production of reactive oxygen species, and NF-κB p65 activation. Inhibition of AOPP-redox signaling activation effectively blocked AOPP-induced EMT in vitro. Studies performed in normal rats showed that chronic administration of AOPPs triggered the occurrence of EMT in rat intestinal epithelia, accompanied by disruption of intestinal integrity, and by promotion of collagen deposition. These effects could be reversed by inhibition of NADPH oxidase. Innovation and Conclusion: This is the first study to demonstrate that AOPPs triggered the occurrence of EMT in IECs in vitro and in vivo through PKC δ-mediated redox-dependent signaling. Our study identifies the role of AOPPs and, in turn, EMT in intestinal fibrosis and provides novel potential targets for the treatment of intestinal fibrotic diseases. Antioxid. Redox Signal. 27, 37-56.

In vitro and ex vivo vanadium antitumor activity in (TGF-β)-induced EMT. Synergistic activity with carboplatin and correlation with tumor metastasis in cancer patients.

PubMed

Petanidis, Savvas; Kioseoglou, Efrosini; Domvri, Kalliopi; Zarogoulidis, Paul; Carthy, Jon M; Anestakis, Doxakis; Moustakas, Aristidis; Salifoglou, Athanasios

2016-05-01

Epithelial to mesenchymal transition (EMT) plays a key role in tumor progression and metastasis as a crucial event for cancer cells to trigger the metastatic niche. Transforming growth factor-β (TGF-β) has been shown to play an important role as an EMT inducer in various stages of carcinogenesis. Previous reports had shown that antitumor vanadium inhibits the metastatic potential of tumor cells by reducing MMP-2 expression and inducing ROS-dependent apoptosis. However, the role of vanadium in (TGF-β)-induced EMT remains unclear. In the present study, we report for the first time on the inhibitory effects of vanadium on (TGF-β)-mediated EMT followed by down-regulation of ex vivo cancer stem cell markers. The results demonstrate blockage of (TGF-β)-mediated EMT by vanadium and reduction in the mitochondrial potential of tumor cells linked to EMT and cancer metabolism. Furthermore, combination of vanadium and carboplatin (a) resulted in synergistic antitumor activity in ex vivo cell cultures, and (b) prompted G0/G1 cell cycle arrest and sensitization of tumor cells to carboplatin-induced apoptosis. Overall, the findings highlight the multifaceted antitumor action of vanadium and its synergistic antitumor efficacy with current chemotherapy drugs, knowledge that could be valuable for targeting cancer cell metabolism and cancer stem cell-mediated metastasis in aggressive chemoresistant tumors. Copyright © 2016 Elsevier Ltd. All rights reserved.

Protons Sensitize Epithelial Cells to Mesenchymal Transition

PubMed Central

Wang, Minli; Hada, Megumi; Saha, Janapriya; Sridharan, Deepa M.; Pluth, Janice M.; Cucinotta, Francis A.

2012-01-01

Proton radiotherapy has gained more favor among oncologists as a treatment option for localized and deep-seated tumors. In addition, protons are a major constituent of the space radiation astronauts receive during space flights. The potential for these exposures to lead to, or enhance cancer risk has not been well studied. Our objective is to study the biological effects of low energy protons on epithelial cells and its propensity to enhance transforming growth factor beta 1 (TGFβ1)-mediated epithelial-mesenchymal transition (EMT), a process occurring during tumor progression and critical for invasion and metastasis. Non-transformed mink lung epithelial cells (Mv1Lu) and hTERT- immortalized human esophageal epithelial cells (EPC) were used in this study. EMT was identified by alterations in cell morphology, EMT-related gene expression changes determined using real-time PCR, and EMT changes in specific cellular markers detected by immunostaining and western blotting. Although TGFβ1 treatment alone is able to induce EMT in both Mv1Lu and EPC cells, low energy protons (5 MeV) at doses as low as 0.1 Gy can enhance TGFβ1 induced EMT. Protons alone can also induce a mild induction of EMT. SD208, a potent TGFβ Receptor 1 (TGFβR1) kinase inhibitor, can efficiently block TGFβ1/Smad signaling and attenuate EMT induction. We suggest a model for EMT after proton irradiation in normal and cancerous tissue based on our results that showed that low and high doses of protons can sensitize normal human epithelial cells to mesenchymal transition, more prominently in the presence of TGFβ1, but also in the absence of TGFβ1. PMID:22844446

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

PubMed Central

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

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.

PubMed

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. Copyright © 2017 American Society for Microbiology.

Endothelin-1 promotes epithelial-mesenchymal transition in human chondrosarcoma cells by repressing miR-300.

PubMed

Wu, Min-Huan; Huang, Pei-Han; Hsieh, Mingli; Tsai, Chun-Hao; Chen, Hsien-Te; Tang, Chih-Hsin

2016-10-25

Chondrosarcoma is a malignant tumor of mesenchymal origin predominantly composed of cartilage-producing cells. This type of bone cancer is extremely resistant to radiotherapy and chemotherapy. Surgical resection is the primary treatment, but is often difficult and not always practical for metastatic disease, so more effective treatments are needed. In particular, it would be helpful to identify molecular markers as targets for therapeutic intervention. Endothelin-1 (ET-1), a potent vasoconstrictor, has been shown to enhance chondrosarcoma angiogenesis and metastasis. We report that ET-1 promotes epithelial-mesenchymal transition (EMT) in human chondrosarcoma cells. EMT is a key pathological event in cancer progression, during which epithelial cells lose their junctions and apical-basal polarity and adopt an invasive phenotype. Our study verifies that ET-1 induces the EMT phenotype in chondrosarcoma cells via the AMP-activated protein kinase (AMPK) pathway. In addition, we show that ET-1 increases EMT by repressing miR-300, which plays an important role in EMT-enhanced tumor metastasis. We also show that miR-300 directly targets Twist, which in turn results in a negative regulation of EMT. We found a highly positive correlation between ET-1 and Twist expression levels as well as tumor stage in chondrosarcoma patient specimens. Therefore, ET-1 may represent a potential novel molecular therapeutic target in chondrosarcoma metastasis.

Endothelin-1 promotes epithelial–mesenchymal transition in human chondrosarcoma cells by repressing miR-300

PubMed Central

Wu, Min-Huan; Huang, Pei-Han; Hsieh, Mingli; Tsai, Chun-Hao; Chen, Hsien-Te; Tang, Chih-Hsin

2016-01-01

Chondrosarcoma is a malignant tumor of mesenchymal origin predominantly composed of cartilage-producing cells. This type of bone cancer is extremely resistant to radiotherapy and chemotherapy. Surgical resection is the primary treatment, but is often difficult and not always practical for metastatic disease, so more effective treatments are needed. In particular, it would be helpful to identify molecular markers as targets for therapeutic intervention. Endothelin-1 (ET-1), a potent vasoconstrictor, has been shown to enhance chondrosarcoma angiogenesis and metastasis. We report that ET-1 promotes epithelial–mesenchymal transition (EMT) in human chondrosarcoma cells. EMT is a key pathological event in cancer progression, during which epithelial cells lose their junctions and apical-basal polarity and adopt an invasive phenotype. Our study verifies that ET-1 induces the EMT phenotype in chondrosarcoma cells via the AMP-activated protein kinase (AMPK) pathway. In addition, we show that ET-1 increases EMT by repressing miR-300, which plays an important role in EMT-enhanced tumor metastasis. We also show that miR-300 directly targets Twist, which in turn results in a negative regulation of EMT. We found a highly positive correlation between ET-1 and Twist expression levels as well as tumor stage in chondrosarcoma patient specimens. Therefore, ET-1 may represent a potential novel molecular therapeutic target in chondrosarcoma metastasis. PMID:27602960

Bleomycin induced epithelial–mesenchymal transition (EMT) in pleural mesothelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Li-Jun; Ye, Hong; Key Laboratory of Pulmonary Diseases, Ministry of Health of China, Wuhan, Hubei

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease characterized by the development of subpleural foci of myofibroblasts that contribute to the exuberant fibrosis. Recent studies revealed that pleural mesothelial cells (PMCs) undergo epithelial–mesenchymal transition (EMT) and play a pivotal role in IPF. In animal model, bleomycin induces pulmonary fibrosis exhibiting subpleural fibrosis similar to what is seen in human IPF. It is not known yet whether bleomycin induces EMT in PMCs. In the present study, PMCs were cultured and treated with bleomycin. The protein levels of collagen-I, mesenchymal phenotypic markers (vimentin and α-smooth muscle actin), and epithelial phenotypicmore » markers (cytokeratin-8 and E-cadherin) were measured by Western blot. PMC migration was evaluated using wound-healing assay of culture PMCs in vitro, and in vivo by monitoring the localization of PMC marker, calretinin, in the lung sections of bleomycin-induced lung fibrosis. The results showed that bleomycin induced increases in collagen-I synthesis in PMC. Bleomycin induced significant increases in mesenchymal phenotypic markers and decreases in epithelial phenotypic markers in PMC, and promoted PMC migration in vitro and in vivo. Moreover, TGF-β1-Smad2/3 signaling pathway involved in the EMT of PMC was demonstrated. Taken together, our results indicate that bleomycin induces characteristic changes of EMT in PMC and the latter contributes to subpleural fibrosis. - Highlights: • Bleomycin induces collagen-I synthesis in pleural mesothelial cells (PMCs). • Bleomycin induces increases in vimentin and α-SMA protein in PMCs. • Bleomycin induces decreases in cytokeratin-8 and E-cadherin protein in PMCs • TGF-β1-Smad2/3 signaling pathway is involved in the PMC EMT induced by bleomycin.« less

Berberine induces neuronal differentiation through inhibition of cancer stemness and epithelial-mesenchymal transition in neuroblastoma cells.

PubMed

Naveen, C R; Gaikwad, Sagar; Agrawal-Rajput, Reena

2016-06-15

Berberine, a plant alkaloid, has been used since many years for treatment of gastrointestinal disorders. It also shows promising medicinal use against metabolic disorders, neurodegenerative disorders and cancer; however its efficacy in neuroblastoma (NB) is poorly explored. EMT is important in cancer stemness and metastasis resulting in failure to differentiate; thus targeting EMT and related pathways can have clinical benefits. Potential of berberine was investigated for (i) neuronal differentiation and cancer stemness inhibition, (ii) underlying molecular mechanisms regulating cancer-stemness and (iii) EMT reversal. Using neuro2a (N2a) neuroblastoma cells (NB); we investigated effect of berberine on neuronal differentiation, cancer-stemness, EMT and underlying signalling by immunofluorescence, RT-PCR, Western blot. High glucose-induced TGF-β mediated EMT model was used to test EMT reversal potential by Western blot and RT-PCR. STRING analysis was done to determine and validate functional protein-interaction networks. We demonstrate berberine induces neuronal differentiation accompanying increased neuronal differentiation markers like MAP2, β-III tubulin and NCAM; generated neurons were viable. Berberine attenuated cancer stemness markers CD133, β-catenin, n-myc, sox2, notch2 and nestin. Berberine potentiated G0/G1 cell cycle arrest by inhibiting proliferation, cyclin dependent kinases and cyclins resulting in apoptosis through increased bax/bcl-2 ratio. Restoration of tumor suppressor proteins, p27 and p53, indicate promising anti-cancer property. The induction of NCAM and reduction in its polysialylation indicates anti-migratory potential which is supported by down regulation of MMP-2/9. It increased epithelial marker laminin and smad and increased Hsp70 levels also suggest its protective role. Molecular insights revealed that berberine regulates EMT via downregulation of PI3/Akt and Ras-Raf-ERK signalling and subsequent upregulation of p38-MAPK. TGF-β secretion from N2a cells was potentiated by high glucose and negatively regulated by berberine through modulation of TGF-β receptors II and III. Berberine reverted mesenchymal markers, vimentin and fibronectin, with restoration of epithelial marker E-cadherin, highlighting the role of berberine in reversal of EMT. Collectively, the study demonstrates prospective use of berberine against neuroblastoma as elucidated through inhibition of fundamental characteristics of cancer stem cells: tumorigenicity and failure to differentiation and instigates reversal in the EMT. Copyright © 2016 Elsevier GmbH. All rights reserved.

SALL4 as an Epithelial-Mesenchymal Transition and Drug Resistance Inducer through the Regulation of c-Myc in Endometrial Cancer

PubMed Central

Yang, Xiaoming; Fang, Chi; Xu, Huali; Xi, Xiaowei

2015-01-01

SALL4 plays important roles in the development and progression of many cancers. However, the role and molecular mechanism of SALL4 in endometrial cancer remain elusive. In the present research, we have demonstrated that the expression of SALL4 was upregulated in endometrial cancer and correlated positively with tumor stage, metastases and poor survival of patients. The overexpression of SALL4 promoted the invasiveness in endometrial cancer cells, as indicated by the upregulation of mesenchymal cell marker N-cadherin and downregulation of the epithelial marker E-cadherin, and invasion assays in vitro. Additionally, there was also an increase in drug resistance in these cell models due to the upregulation of ATP-binding cassette multidrug transporter ABCB1 expression. Moreover, we also found that ABCB1 was critical for SALL4-induced drug resistance. In contrast, SALL4 knockdown restored drug sensitivity, reversed EMT, diminished cell metastasis and suppressed the downregulation of E-cadherin and the upregulation of N-cadherin and ABCB1. Furthermore, we showed that SALL4 upregulated c-Myc expression and c-Myc was a direct target for SALL4 by ChIP assay, depletion of c-Myc with siRNA abolished the SALL4-induced downregulation of E-cadherin, upregulation of N-cadherin and ABCB1, suggesting that c-Myc was a downstream target for SALL4 and required for SALL4-induced EMT, invasion and drugs resistance in endometrial cancer cells. These results indicated that SALL4 could induce EMT and resistance to antineoplastic drugs through the regulation of c-Myc. SALL4 and c-Myc may be novel therapeutic targets for endometrial cancer. PMID:26407074

IL-1β induced methylation of the estrogen receptor ERα gene correlates with EMT and chemoresistance in breast cancer cells.

PubMed

Jiménez-Garduño, Aura M; Mendoza-Rodríguez, Mónica G; Urrutia-Cabrera, Daniel; Domínguez-Robles, María C; Pérez-Yépez, Eloy A; Ayala-Sumuano, Jorge Tonatiuh; Meza, Isaura

2017-08-26

Inflammation has been recently acknowledged as a key participant in the physiopathology of oncogenesis and tumor progression. The inflammatory cytokine IL-1β has been reported to induce the expression of markers associated with malignancy in breast cancerous cells through Epithelial-Mesenchymal Transition (EMT). Aggressive breast cancer tumors classified as Triple Negative do not respond to hormonal treatment because they lack three crucial receptors, one of which is the estrogen receptor alpha (ERα). Expression of ERα is then considered a good prognostic marker for tamoxifen treatment of this type of cancer, as the binding of this drug to the receptor blocks the transcriptional activity of the latter. Although it has been suggested that inflammatory cytokines in the tumor microenvironment could regulate ERα expression, the mechanism(s) involved in this process have not yet been established. We show here that, in a cell model of breast cancer cells (6D cells), in which the inflammatory cytokine IL-1β induces EMT by activation of the IL-1β/IL-1RI/β-catenin pathway, the up regulation of TWIST1 leads to methylation of the ESR1 gene promoter. This epigenetic modification produced significant decrease of the ERα receptor levels and increased resistance to tamoxifen. The direct participation of IL-1β in these processes was validated by blockage of the cytokine-induced signaling pathway by wortmannin inactivation of the effectors PI3K/AKT. These results support our previous reports that have suggested direct participation of the inflammatory cytokine IL-1β in the transition to malignancy of breast cancer cells. Copyright © 2017 Elsevier Inc. All rights reserved.

GROα overexpression drives cell migration and invasion in triple negative breast cancer cells.

PubMed

Bhat, Kruttika; Sarkissyan, Marianna; Wu, Yanyuan; Vadgama, Jaydutt V

2017-07-01

Triple negative breast cancer (TNBC) is a subtype of highly aggressive breast cancer with poor prognosis. The main characteristic feature of TNBC is its lack of expression of ER, PR and HER2 receptors that are targets for treatments. Hence, it is imperative to identify novel therapeutic strategies to target TNBC. Our aim was to examine whether GROα is a specific marker for TNBC metastasis. For this we performed qPCR, ELISA, migration/invasion assays, western blotting, and siRNA transfections. Evaluation of baseline GROα expression in different breast cancer (BC) subtypes showed that it is significantly upregulated in breast tumor cells, specifically in TNBC cell line. On further evaluation in additional 17 TNBC cell lines we found that baseline GROα expression was significantly elevated in >50% of the cell lines validating GROα overexpression specifically in TNBC cells. Moreover, GROα-stimulation in MCF7 and SKBR3 cells and GROα‑knockdown in MDA-MB‑231 and HCC1937 cells elicited dramatic changes in migration and invasion abilities in vitro. Corresponding changes in EMT markers were also observed in phenotypically modified BC cells. Furthermore, mechanistic studies identified GROα regulating EMT markers and migration/invasion via MAPK pathway and specific inhibition using PD98059 resulted in the reversal of effects induced by GROα on BC cells. In conclusion, our study provides strong evidence to suggest that GROα is a critical modulator of TNBC migration/invasion and proposes GROα as a potential therapeutic target for treatment of TNBC metastasis.

Galectin-8 induces partial epithelial–mesenchymal transition with invasive tumorigenic capabilities involving a FAK/EGFR/proteasome pathway in Madin–Darby canine kidney cells

PubMed Central

Oyanadel, Claudia; Holmes, Christopher; Pardo, Evelyn; Retamal, Claudio; Shaughnessy, Ronan; Smith, Patricio; Cortés, Priscilla; Bravo-Zehnder, Marcela; Metz, Claudia; Feuerhake, Teo; Romero, Diego; Roa, Juan Carlos; Montecinos, Viviana; Soza, Andrea; González, Alfonso

2018-01-01

Epithelial cells can acquire invasive and tumorigenic capabilities through epithelial–mesenchymal-transition (EMT). The glycan-binding protein galectin-8 (Gal-8) activates selective β1-integrins involved in EMT and is overexpressed by certain carcinomas. Here we show that Gal-8 overexpression or exogenous addition promotes proliferation, migration, and invasion in nontumoral Madin–Darby canine kidney (MDCK) cells, involving focal-adhesion kinase (FAK)-mediated transactivation of the epidermal growth factor receptor (EGFR), likely triggered by α5β1integrin binding. Under subconfluent conditions, Gal-8–overexpressing MDCK cells (MDCK-Gal-8H) display hallmarks of EMT, including decreased E-cadherin and up-regulated expression of vimentin, fibronectin, and Snail, as well as increased β-catenin activity. Changes related to migration/invasion included higher expression of α5β1 integrin, extracellular matrix-degrading MMP13 and urokinase plasminogen activator/urokinase plasminogen activator receptor (uPA/uPAR) protease systems. Gal-8–stimulated FAK/EGFR pathway leads to proteasome overactivity characteristic of cancer cells. Yet MDCK-Gal-8H cells still develop apical/basolateral polarity reverting EMT markers and proteasome activity under confluence. This is due to the opposite segregation of Gal-8 secretion (apical) and β1-integrins distribution (basolateral). Strikingly, MDCK-Gal-8H cells acquired tumorigenic potential, as reflected in anchorage-independent growth in soft agar and tumor generation in immunodeficient NSG mice. Therefore, Gal-8 can promote oncogenic-like transformation of epithelial cells through partial and reversible EMT, accompanied by higher proliferation, migration/invasion, and tumorigenic properties. PMID:29298841

Matrix Metalloproteinase-9–Null Mice Are Resistant to TGF-β–Induced Anterior Subcapsular Cataract Formation

PubMed Central

Korol, Anna; Pino, Giuseppe; Dwivedi, Dhruva; Robertson, Jennifer V.; Deschamps, Paula A.; West-Mays, Judith A.

2015-01-01

Epithelial-mesenchymal transition (EMT) is associated with fibrotic diseases in the lens, such as anterior subcapsular cataract (ASC) formation. Often mediated by transforming growth factor (TGF)-β, EMT in the lens involves the transformation of lens epithelial cells into a multilayering of myofibroblasts, which manifest as plaques beneath the lens capsule. TGF-β–induced EMT and ASC have been associated with the up-regulation of two matrix metalloproteinases (MMPs): MMP-2 and MMP-9. The current study used MMP-2 and MMP-9 knockout (KO) mice to further determine their unique roles in TGF-β–induced ASC formation. Adenoviral injection of active TGF-β1 into the anterior chamber of all wild-type and MMP-2 KO mice led to the formation of distinct ASC plaques that were positive for α-smooth muscle actin, a marker of EMT. In contrast, only a small proportion of the MMP-9 KO eyes injected with adenovirus-expressing TGF-β1 exhibited ASC plaques. Isolated lens epithelial explants from wild-type and MMP-2 KO mice that were treated with TGF-β exhibited features indicative of EMT, whereas those from MMP-9 KO mice did not acquire a mesenchymal phenotype. MMP-9 KO mice were further bred onto a TGF-β1 transgenic mouse line that exhibits severe ASC formation, but shows a resistance to ASC formation in the absence of MMP-9. These findings suggest that MMP-9 expression is more critical than MMP-2 in mediating TGF-β–induced ASC formation. PMID:24814605

Rosmarinic Acid Activates AMPK to Inhibit Metastasis of Colorectal Cancer

PubMed Central

Han, Yo-Han; Kee, Ji-Ye; Hong, Seung-Heon

2018-01-01

Rosmarinic acid (RA) has been used as an anti-inflammatory, anti-diabetic, and anti-cancer agent. Although RA has also been shown to exert an anti-metastatic effect, the mechanism of this effect has not been reported to be associated with AMP-activated protein kinase (AMPK). The aim of this study was to elucidate whether RA could inhibit the metastatic properties of colorectal cancer (CRC) cells via the phosphorylation of AMPK. RA inhibited the proliferation of CRC cells through the induction of cell cycle arrest and apoptosis. In several metastatic phenotypes of CRC cells, RA regulated epithelial–mesenchymal transition (EMT) through the upregulation of an epithelial marker, E-cadherin, and the downregulation of the mesenchymal markers, N-cadherin, snail, twist, vimentin, and slug. Invasion and migration of CRC cells were inhibited and expressions of matrix metalloproteinase (MMP)-2 and MMP-9 were decreased by RA treatment. Adhesion and adhesion molecules such as ICAM-1 and integrin β1 expressions were also reduced by RA treatment. In particular, the effects of RA on EMT and MMPs expressions were due to the activation of AMPK. Moreover, RA inhibited lung metastasis of CRC cells by activating AMPK in mouse model. Collectively, these results proved that RA could be potential therapeutic agent against metastasis of CRC. PMID:29459827

Snail regulates cell survival and inhibits cellular senescence in human metastatic prostate cancer cell lines.

PubMed

Emadi Baygi, Modjtaba; Soheili, Zahra Soheila; Schmitz, Ingo; Sameie, Shahram; Schulz, Wolfgang A

2010-12-01

The epithelial-mesenchymal transition (EMT) is regarded as an important step in cancer metastasis. Snail, a master regulator of EMT, has been recently proposed to act additionally as a cell survival factor and inducer of motility. We have investigated the function of Snail (SNAI1) in prostate cancer cells by downregulating its expression via short (21-mer) interfering RNA (siRNA) and measuring the consequences on EMT markers, cell viability, death, cell cycle, senescence, attachment, and invasivity. Of eight carcinoma cell lines, the prostate carcinoma cell lines LNCaP and PC-3 showed the highest and moderate expression of SNAI1 mRNA, respectively, as measured by quantitative RT-PCR. Long-term knockdown of Snail induced a severe decline in cell numbers in LNCaP and PC-3 and caspase activity was accordingly enhanced in both cell lines. In addition, suppression of Snail expression induced senescence in LNCaP cells. SNAI1-siRNA-treated cells did not tolerate detachment from the extracellular matrix, probably due to downregulation of integrin α6. Expression of E-cadherin, vimentin, and fibronectin was also affected. Invasiveness of PC-3 cells was not significantly diminished by Snail knockdown. Our data suggest that Snail acts primarily as a survival factor and inhibitor of cellular senescence in prostate cancer cell lines. We therefore propose that Snail can act as early driver of prostate cancer progression.

Metformin inhibits TGF-β1-induced epithelial-to-mesenchymal transition-like process and stem-like properties in GBM via AKT/mTOR/ZEB1 pathway.

PubMed

Song, Yang; Chen, Yong; Li, Yunqian; Lyu, Xiaoyan; Cui, Jiayue; Cheng, Ye; Zhao, Liyan; Zhao, Gang

2018-01-23

Glioblastoma (GBM) is the most frequent and aggressive brain tumor in adults. In spite of advances in diagnosis and therapy, the prognosis is still relatively poor. The invasive property of GBM is the major cause of death in patients. Epithelial-to-mesenchymal transition-like process (EMT-like process) is considered to play an important role in the invasive property. Metformin has been reported as a regulator of EMT-like process. In this study, we confirmed that metformin inhibited TGF-β1-induced EMT-like process and EMT-associated migration and invasion in LN18 and U87 GBM cells. Our results also showed that metformin significantly suppressed self-renewal capacity of glioblastoma stem cells (GSCs), and expression of stem cell markers Bmi1, Sox2 and Musashi1, indicating that metformin can inhibit cancer stem-like properties of GBM cells. We further clarified that metformin specifically inhibited TGF-β1 activated AKT, the downstream molecular mTOR and the leading transcription factor ZEB1. Taken together, our data demonstrate that metformin inhibits TGF-β1-induced EMT-like process and cancer stem-like properties in GBM cells via AKT/mTOR/ZEB1 pathway and provide evidence of metformin for further clinical investigation targeted GBM.

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

PubMed

Chang, Tae Ik; Kang, Hye-Young; Kim, Kyung Sik; Lee, Sun Ha; Nam, Bo Young; Paeng, Jisun; Kim, Seonghun; Park, Jung Tak; Yoo, Tae-Hyun; Kang, Shin-Wook; Han, Seung Hyeok

2014-01-01

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 significantly greater in PD rats than in C rats (p<0.05). These changes of the peritoneum in PD rats were significantly attenuated by simvastatin. This study demonstrated that PD-related EMT was mediated via the mevalonate pathway, and statin treatment inhibited the EMT changes in HG-treated HPMCs and PDF-stimulated PD rats. These findings suggest that statins may be a promising therapeutic strategy for preservation of peritoneal membrane integrity in long-term PD patients.

ECM1 regulates tumor metastasis and CSC-like property through stabilization of β-catenin.

PubMed

Lee, K-m; Nam, K; Oh, S; Lim, J; Kim, R K; Shim, D; Choi, J-h; Lee, S-J; Yu, J-H; Lee, J W; Ahn, S H; Shin, I

2015-12-10

Extracellular Matrix Protein 1 (ECM1) is a marker for tumorigenesis and is correlated with invasiveness and poor prognosis in various types of cancer. However, the functional role of ECM1 in cancer metastasis is unclear. Here, we detected high ECM1 level in breast cancer patient sera that was associated with recurrence of tumor. The modulation of ECM1 expression affected not only cell migration and invasion, but also sphere-forming ability and drug resistance in breast cancer cell lines. In addition, ECM1 regulated the gene expression associated with the epithelial to mesenchymal transition (EMT) progression and cancer stem cell (CSC) maintenance. Interestingly, ECM1 increased β-catenin expression at the post-translational level through induction of MUC1, which was physically associated with β-catenin. Indeed, the association between β-catenin and the MUC1 cytoplasmic tail was increased by ECM1. Furthermore, forced expression of β-catenin altered the gene expression that potentiated EMT progression and CSC phenotype maintenance in the cells. These data provide evidence that ECM1 has an important role in cancer metastasis through β-catenin stabilization.

The over expression of long non-coding RNA ANRIL promotes epithelial-mesenchymal transition by activating the ATM-E2F1 signaling pathway in pancreatic cancer: An in vivo and in vitro study.

PubMed

Chen, Shi; Zhang, Jia-Qiang; Chen, Jiang-Zhi; Chen, Hui-Xing; Qiu, Fu-Nan; Yan, Mao-Lin; Chen, Yan-Ling; Peng, Cheng-Hong; Tian, Yi-Feng; Wang, Yao-Dong

2017-09-01

This study aims to investigate the roles of lncRNA ANRIL in epithelial-mesenchymal transition (EMT) by regulating the ATM-E2F1 signaling pathway in pancreatic cancer (PC). PC rat models were established and ANRIL overexpression and interference plasmids were transfected. The expression of ANRIL, EMT markers (E-cadherin, N-cadherin and Vimentin) and ATM-E2F1 signaling pathway-related proteins (ATM, E2F1, INK4A, INK4B and ARF) were detected. Small molecule drugs were applied to activate and inhibit the ATM-E2F1 signaling pathway. Transwell assay and the scratch test were adopted to detect cell invasion and migration abilities. ANRIL expression in the PC cells was higher than in normal pancreatic duct epithelial cells. In the PC rat models and PC cells, ANRIL interference promoted the expressions of INK4B, INK4A, ARF and E-cadherin, while reduced N-cadherin and Vimentin expression. Over-expressed ANRIL decreased the expression of INK4B, INK4A, ARF and E-cadherin, but raised N-cadherin and Vimentin expressions. By inhibiting the ATM-E2F1 signaling pathway in PC cells, E-cadherin expression increased but N-cadherin and Vimentin expressions decreased. After ANRIL was silenced or the ATM-E2F1 signaling pathway inhibited, PC cell migration and invasion abilities were decreased. In conclusion, over-expression of lncRNA ANRIL can promote EMT of PC cells by activating the ATM-E2F1 signaling pathway. Copyright © 2017 Elsevier B.V. All rights reserved.

Inhibition of NF-kappa B pathway leads to deregulation of epithelial-mesenchymal transition and neural invasion in pancreatic cancer.

PubMed

Nomura, Alice; Majumder, Kaustav; Giri, Bhuwan; Dauer, Patricia; Dudeja, Vikas; Roy, Sabita; Banerjee, Sulagna; Saluja, Ashok K

2016-12-01

NF-κB has an essential role in the initiation and progression of pancreatic cancer and specifically mediates the induction of epithelial-mesenchymal transition and invasiveness. In this study, we demonstrate the importance of activated NF-κB signaling in EMT induction, lymphovascular metastasis, and neural invasion. Modulation of NF-κB activity was accomplished through the specific NF-κB inhibitor (BAY 11-7085), triptolide, and Minnelide treatment, as well as overexpression of IKBα repressor and IKK activator plasmids. In the classical lymphovascular metastatic cascade, inhibition of NF-κB decreased the expression of several EMT transcription factors (SNAI1, SNAI2, and ZEB1) and mesenchymal markers (VIM and CDH2) and decreased in vitro invasion, which was rescued by IKK activation. This was further demonstrated in vivo via BAY 11-7085 treatment in a orthotopic model of pancreatic cancer. In vivo NF-κB inhibition decreased tumor volume; decreased tumor EMT gene expression, while restoring cell-cell junctions; and decreasing overall metastasis. Furthermore, we demonstrate the importance of active NF-κB signaling in neural invasion. Triptolide treatment inhibits Nerve Growth Factor (NGF) mediated, neural-tumor co-culture in vitro invasion, and dorsal root ganglia (DRG) neural outgrowth through a disruption in tumor-neural cross talk. In vivo, Minnelide treatment decreased neurotrophin expression, nerve density, and sciatic nerve invasion. Taken together, this study demonstrates the importance of NF-κB signaling in the progression of pancreatic cancer through the modulation of EMT induction, lymphovascular invasion, and neural invasion.

The meaning of PIWI proteins in cancer development.

PubMed

Litwin, Monika; Szczepańska-Buda, Anna; Piotrowska, Aleksandra; Dzięgiel, Piotr; Witkiewicz, Wojciech

2017-05-01

Cancer is a histologically and genetically heterogeneous population of tumor cells that exhibits distinct molecular profiles determined by epigenetic alterations. P-element-induced wimpy testis (PIWI) proteins in complex with PIWI-interacting RNA (piRNA) have been previously demonstrated to be involved in epigenetic regulation in germline cells. Recently, reactivation of PIWI expression, primarily PIWI-like protein 1 and 2, through aberrant DNA methylation resulting in genomic silencing has been identified in various types of tumors. It has been suggested that the PIWI-piRNA complex contributes to cancer development and progression by promoting a stem-like state of cancer cells, or cancer stem cells (CSCs). It has been identified that CSCs represent the cells that have undergone epithelial-mesenchymal transition (EMT) and acquired metastatic capacities. However, the molecular association between the EMT process and the stem-cell state remains unclear. Further extensive characterization of CSCs in individual types of tumors is required to identify specific markers for the heterogeneous population of CSCs and therefore selectively target CSCs. Previous studies indicate a reciprocal regulation between PIWI proteins and a complex signaling network linking markers characterized for CSCs and transcription factors involved in EMT. In the present review, studies of PIWI function are summarized, and the potential involvement of PIWI proteins in cancer development and progression is discussed.

The meaning of PIWI proteins in cancer development

PubMed Central

Litwin, Monika; Szczepańska-Buda, Anna; Piotrowska, Aleksandra; Dzięgiel, Piotr; Witkiewicz, Wojciech

2017-01-01

Cancer is a histologically and genetically heterogeneous population of tumor cells that exhibits distinct molecular profiles determined by epigenetic alterations. P-element-induced wimpy testis (PIWI) proteins in complex with PIWI-interacting RNA (piRNA) have been previously demonstrated to be involved in epigenetic regulation in germline cells. Recently, reactivation of PIWI expression, primarily PIWI-like protein 1 and 2, through aberrant DNA methylation resulting in genomic silencing has been identified in various types of tumors. It has been suggested that the PIWI-piRNA complex contributes to cancer development and progression by promoting a stem-like state of cancer cells, or cancer stem cells (CSCs). It has been identified that CSCs represent the cells that have undergone epithelial-mesenchymal transition (EMT) and acquired metastatic capacities. However, the molecular association between the EMT process and the stem-cell state remains unclear. Further extensive characterization of CSCs in individual types of tumors is required to identify specific markers for the heterogeneous population of CSCs and therefore selectively target CSCs. Previous studies indicate a reciprocal regulation between PIWI proteins and a complex signaling network linking markers characterized for CSCs and transcription factors involved in EMT. In the present review, studies of PIWI function are summarized, and the potential involvement of PIWI proteins in cancer development and progression is discussed. PMID:28529570

TGFβ1-induced down-regulation of microRNA-138 contributes to epithelial-mesenchymal transition in primary lung cancer cells.

PubMed

Zhang, Fang; Li, Tiepeng; Han, Lu; Qin, Peng; Wu, Zhao; Xu, Benling; Gao, Quanli; Song, Yongping

2018-02-19

The existence of cancer stem cells within the tumor could lead to cancer therapy resistance. TGFβ1 is considered as one of the most powerful players in the generation of CSCs through induction of epithelial-mesenchymal transition in different types of cancer including lung cancer, however, the detailed mechanisms by which TGFβ1 contribute to EMT induction and CSC maintenance remains unclear. Here, we showed primary lung cancer cells treated by TGFβ1 exhibit mesenchymal features, including morphology and expression of mesenchymal marker in a time-dependent manner. We also observed long-term TGFβ1 exposure leads to an enrichment of a sub-population of CD44 + CD90 + cells which represent CSCs in lung cancer cells. Moreover, the differential expression microRNAs between CSCs and non-CSCs were identified using next-generation sequencing to screen key miRNAs which might contribute to TGFβ1-induced EMT and CSCs generation. Among those differentially expressed miRNAs, the expression of microRNA-138 was time-dependently down-regulated by TGFβ1 treatment. We further demonstrated primary lung cancer cells, in which we knockdown the expression of miR-138, exhibit mesenchymal phenotypes and stem cell properties. Taken together, these findings indicate TGFβ1-induced down-regulation of microRNA-138 contributes to EMT in primary lung cancer cells, and suggest that miR-138 might serve as a potential therapeutic target. Copyright © 2018 Elsevier Inc. All rights reserved.

LRIG1 opposes epithelial to mesenchymal transition and inhibits invasion of basal-like breast cancer cells

PubMed Central

Yokdang, Nucharee; Hatakeyama, Jason; Wald, Jessica H.; Simion, Catalina; Tellez, Joseph D.; Chang, Dennis Z.; Swamynathan, Manojit Mosur; Chen, Mingyi; Murphy, William J.; Carraway, Kermit L.; Sweeney, Colleen

2015-01-01

LRIG1, a member of the LRIG family of transmembrane leucine rich repeat-containing proteins, is a negative regulator of receptor tyrosine kinase signaling and a tumor suppressor. LRIG1 expression is broadly decreased in human cancer and in breast cancer, low expression of LRIG1 has been linked to decreased relapse-free survival. Recently, low expression of LRIG1 was revealed to be an independent risk factor for breast cancer metastasis and death. These findings suggest that LRIG1 may oppose breast cancer cell motility and invasion, cellular processes which are fundamental to metastasis. However, very little is known of LRIG1 function in this regard. In this study, we demonstrate that LRIG1 is down-regulated during epithelial to mesenchymal transition (EMT) of human mammary epithelial cells, suggesting that LRIG1 expression may represent a barrier to EMT. Indeed, depletion of endogenous LRIG1 in human mammary epithelial cells expands the stem cell population, augments mammosphere formation and accelerates EMT. Conversely, expression of LRIG1 in highly invasive Basal B breast cancer cells provokes a mesenchymal to epithelial transition accompanied by a dramatic suppression of tumorsphere formation and a striking loss of invasive growth in three-dimensional culture. LRIG1 expression perturbs multiple signaling pathways and represses markers and effectors of the mesenchymal state. Furthermore, LRIG1 expression in MDA-MB-231 breast cancer cells significantly slows their growth as tumors, providing the first in vivo evidence that LRIG1 functions as a growth suppressor in breast cancer. PMID:26387542

microRNA-145 modulates epithelial-mesenchymal transition and suppresses proliferation, migration and invasion by targeting SIP1 in human cervical cancer cells.

PubMed

Sathyanarayanan, Anusha; Chandrasekaran, Karthik Subramanian; Karunagaran, Devarajan

2017-04-01

Previously, it has been reported that microRNA-145 (miR-145) is lowly expressed in human cervical cancers and that its putative tumour suppressive role may be attributed to epithelial-mesenchymal transition (EMT) regulation. Here, we aimed to assess whether miR-145 may affect EMT-associated markers/genes and suppress cervical cancer growth and motility, and to provide a mechanistic basis for these phenomena. The identification of the SMAD-interacting protein 1 (SIP1) mRNA as putative miR-145 target was investigated using a 3' untranslated region (3'UTR) luciferase assay and Western blotting, respectively. The functional effects of exogenous miR-145 expression, miR-145 suppression or siRNA-mediated SIP1 expression down-regulation in cervical cancer-derived C33A and SiHa cells were analysed using Western blotting, BrdU incorporation (proliferation), transwell migration and invasion assays. In addition, the expression levels of miR-145 and SIP1 were determined in primary human cervical cancer and non-cancer tissue samples using qRT-PCR. We found that miR-145 binds to the wild-type 3'UTR of SIP1, but not to its mutant counterpart, and that, through this binding, miR-145 can effectively down-regulate SIP1 expression. In addition, we found that exogenous miR-145 expression or siRNA-mediated down-regulation of SIP1 expression attenuates the proliferation, migration and invasion of C33A and SiHa cells and alters the expression of the EMT-associated markers CDH1, VIM and SNAI1, whereas inhibition of endogenous miR-145 expression elicited the opposite effects. The expression of miR-145 in cervical cancer tissue samples was found to be low, while that of SIP1 was found to be high compared to non-cancerous cervical tissues. An inverse expression correlation between the two was substantiated through the anlaysis of data deposited in the TCGA database. Our data indicate that low miR-145 expression levels in conjunction with elevated SIP1 expression levels may contribute to cervical cancer development. MiR-145-mediated regulation of SIP1 provides a novel mechanistic basis for its tumour suppressive mode of action in human cervical cancer cells.

TNF-α-inducing protein of Helicobacter pylori induces epithelial-mesenchymal transition (EMT) in gastric cancer cells through activation of IL-6/STAT3 signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Guodong; Tang, Na; Wang, Chao

Tumor necrosis factor (TNF)-α-inducing protein (Tipα) is a newly identified carcinogenic factor secreted by Helicobacter pylori (H. pylori). Although it has been proved that Tipα is a strong inducer of epithelial-mesenchymal transition (EMT), a crucial process of migration, the exact molecular mechanism is unknown. Current evidence indicates that the oncogenic transcription factor signal transducers and activators of transcription 3 (STAT3) is inappropriately activated in multiple malignancies, including gastric cancer. In this study, we showed that Tipα significantly down-regulated the expression of EMT-related markers E-cadherin as well as up-regulated N-cadherin and vimentin in SGC7901 cells, with typical morphological changes of EMT. Tipα alsomore » promoted proliferation and migration of SGC7901 cells. Furthermore, Tipα activated interleukin-6 (IL-6)/STAT3 signaling pathway in SGC7901 cells. The effects of Tipα treatment observed was abolished when we block IL-6/STAT3 signaling pathway. Altogether, our data demonstrated that Tipα may accelerate tumor aggressiveness in gastric cancer by promoting EMT through activation of IL-6/STAT3 pathway. - Highlights: • Tipα induces EMT and activates IL-6/STAT3 pathway in gastric cancer cells. • IL-6/STAT3 pathway inhibition reverses Tipα-induced proliferation and migration in gastric cancer cells. • Tipα induces EMT in gastric cancer cells via IL-6/STAT3 pathway activation.« less

MIR517C inhibits autophagy and the epithelial-to-mesenchymal (-like) transition phenotype in human glioblastoma through KPNA2-dependent disruption of TP53 nuclear translocation

PubMed Central

Lu, Yuntao; Xiao, Limin; Liu, Yawei; Wang, Hai; Li, Hong; Zhou, Qiang; Pan, Jun; Lei, Bingxi; Huang, Annie; Qi, Songtao

2015-01-01

The epithelial-to-mesenchymal (-like) transition (EMT), a crucial embryonic development program, has been linked to the regulation of glioblastoma (GBM) progression and invasion. Here, we investigated the role of MIR517C/miR-517c, which belongs to the C19MC microRNA cluster identified in our preliminary studies, in the pathogenesis of GBM. We found that MIR517C was associated with improved prognosis in patients with GBM. Furthermore, following treatment with the autophagy inducer temozolomide (TMZ) and low glucose (LG), MIR517C degraded KPNA2 (karyopherin alpha 2 [RAG cohort 1, importin alpha 1]) and subsequently disturbed the nuclear translocation of TP53 in the GBM cell line U87 in vitro. Interestingly, this microRNA could inhibit autophagy and reduce cell migration and infiltration in U87 cells harboring wild-type (WT) TP53, but not in U251 cells harboring mutant (MU) TP53. Moreover, the expression of epithelial markers (i.e., CDH13/T-cadherin and CLDN1 [claudin 1]) increased, while the expression of mesenchymal markers (i.e., CDH2/N-cadherin, SNAI1/Snail, and VIM [vimentin]) decreased, indicating that the EMT status was blocked by MIR517C in U87 cells. Compared with MIR517C overexpression, MIR517C knockdown promoted infiltration of U87 cells to the surrounding structures in nude mice in vivo. The above phenotypic changes were also observed in TP53+/+ and TP53-/- HCT116 colon cancer cells. In summary, our study provided support for a link between autophagy and EMT status in WT TP53 GBM cells and provided evidence for the signaling pathway (MIR517C-KPNA2-cytoplasmic TP53) involved in attenuating autophagy and eliminating the increased migration and invasion during the EMT. PMID:26553592

Transformation of Mouse Liver Cells by Methylcholanthrene Leads to Phenotypic Changes Associated with Epithelial-mesenchymal Transition.

PubMed

Oh, Jiyun; Kwak, Jae-Hwan; Kwon, Do-Young; Kim, A-Young; Oh, Dal-Seok; Je, Nam Kyung; Lee, Jaewon; Jung, Young-Suk

2014-12-01

Environmental pollutants such as polycyclic aromatic hydrocarbons (PAHs) have been implicated in cancer development and progression. However, the effects of PAHs on carcinogenesis are still poorly understood. Here, we characterized a mouse cancer cell line BNL 1ME A. 7R.1 (1MEA) derived by transformation of non-tumorigenic liver cell line BNL CL.2 (BNL) using 3-methylcholanthrene (3MC), a carcinogenic PAH. RT-PCR and immunoblot analysis were used to determine the expression level of mRNA and proteins, respectively. To determine functionality, cell motility was assessed in vitro using a transwell migration assay. Both mRNA and protein levels of E-cadherin were significantly decreased in 1MEA cells in comparison with BNL cells. While the expression levels of mesenchymal markers and related transcription factors were enhanced in 1MEA cells, which could lead to increase in cell motility. Indeed, we found that 7-day exposure of BNL cells to 3-MC reduced the level of the adhesion molecule and epithelial marker Ecadherin and increased reciprocally the level of the mesenchymal marker vimentin in a dose-dependent manner. Taken together, these results indicate that the process of epithelial-mesenchymal transition (EMT) may be activated during premalignant transformation induced by 3-MC. A mechanism study to elucidate the relation between 3-MC exposure and EMT is underway in our laboratory.

CLDN1 expression in cervical cancer cells is related to tumor invasion and metastasis.

PubMed

Zhang, Wei-Na; Li, Wei; Wang, Xiao-Li; Hu, Zheng; Zhu, Da; Ding, Wen-Cheng; Liu, Dan; Li, Ke-Zhen; Ma, Ding; Wang, Hui

2016-12-27

Even though infection with human papillomaviruses (HPV) is very important, it is not the sole cause of cervical cancer. Because it is known that genetic variations that result from HPV infection are probably the most important causes of cervical cancer, we used human whole genome array comparative genomic hybridization to detect the copy number variations of genes in cervical squamous cell carcinoma. The results of the array were validated by PCR, FISH and immunohistochemistry. We find that the copy number and protein expression of claudin-1 (CLDN1) increase with the progression of cervical cancer. The strong positive staining of CLDN1 in the cervical lymph node metastasis group received a significantly higher score than the staining in the group with no lymph node metastasis of cervical cancer tissues. The overexpression of CLDN1 in SiHa cells can increase anti-apoptosis ability and promote invasive ability of these cells accompanied by a decrease in expression of the epithelial marker E-cadherin as well as an increase in the expression of the mesenchymal marker vimentin. CLDN1 induces the epithelial-mesenchymal transition (EMT) through its interaction with SNAI1. Furthermore, we demonstrate that CLDN1 overexpression has significant effects on the growth and metastasis of xenografted tumors in athymic mice. These data suggest that CLDN1 promotes invasion and metastasis in cervical cancer cells via the expression of EMT/invasion-related genes. Therefore, CLDN1 could be a potential therapeutic target for the treatment of cervical cancer.

Clinicopathological significance and prognostic value of myoinvasive patterns in endometrial endometrioid carcinoma.

PubMed

Amălinei, Cornelia; Aignătoaei, Anda Maria; Balan, Raluca Anca; Giuşcă, Simona Eliza; Lozneanu, Ludmila; Avădănei, Elena Roxana; Căruntu, Irina Draga

2018-01-01

Endometrioid endometrial carcinoma has an overall good prognosis. However, variable five-year survival rates (92%-42%) have been reported in FIGO stage I, suggesting the involvement of other factors related to tumor biological behavior. These may be related to the role played by epithelial-mesenchymal transition (EMT) and cancer stem cells in endometrial carcinogenesis. In this context, our review highlights the prognostic significance of several types of myoinvasion in low grade, low stage endometrioid endometrial carcinoma, as a reflection of these molecular changes at the invasive front. According to recently introduced myoinvasive patterns, the diffusely infiltrating and microcystic, elongated, and fragmented (MELF) patterns show loss of hormone receptors, along with EMT and high expression of cancer stem cell markers, being associated with a poor prognosis. Additionally, MELF pattern exhibits a high incidence of lymphovascular invasion and lymph node metastases. Conversely, the broad front pattern has a good prognosis and a low expression of EMT and stem cells markers. Similarly, the adenomyosis (AM)-like and adenoma malignum patterns of invasion are associated to a favorable prognosis, but nevertheless, they raise diagnostic challenges. AM-like pattern must be differentiated from carcinoma invasion of AM foci, while adenoma malignum pattern creates difficulties in appreciating the depth of myoinvasion and requires differential diagnosis with other conditions. Another pattern expecting its validation and prognostic significance value is the nodular fasciitis-like stroma and large cystic growth pattern. In practice, the knowledge of these patterns of myoinvasion may be valuable for the correct assessment of stage, may improve prognosis evaluation and may help identify molecules for future targeted therapies.

Hedgehog Signaling Regulates Epithelial-Mesenchymal Transition in Pancreatic Cancer Stem-Like Cells

PubMed Central

Wang, Feng; Ma, Ling; Zhang, Zhengkui; Liu, Xiaoran; Gao, Hongqiao; Zhuang, Yan; Yang, Pei; Kornmann, Marko; Tian, Xiaodong; Yang, Yinmo

2016-01-01

Hedgehog (Hh) signaling is crucially involved in tumorigenesis. This study aimed to assess the role of Hh signaling in the regulation of epithelial-mesenchymal transition (EMT), stemness properties and chemoresistance of human pancreatic Panc-1 cancer stem cells (CSCs). Panc-1 cells were transfected with recombinant lentiviral vectors to silence SMO and serum-free floating-culture system was used to isolate Panc-1 tumorspheres. The expression of CSC and EMT markers was detected by flow cytometry, real-time RT-PCR and Western blot analysis. Malignant behaviors of Panc-1 CSC were evaluated by tumorigenicity assays and nude mouse lung metastasis model. We found that tumorspheres derived from pancreatic cancer cell line Panc-1 possessed self-renewal, differentiation and stemness properties. Hh pathway and EMT were active in Panc-1 tumorspheres. Inhibition of Hh signaling by SMO knockdown inhibited self-renewal, EMT, invasion, chemoresistance, pulmonary metastasis, tumorigenesis of pancreatic CSCs. In conclusion, Hh signaling contributes to the maintenance of stem-like properties and chemoresistance of pancreatic CSC and promotes the tumorigenesis and metastasis of pancreatic cancer. Hh pathway is a potential molecular target for the development of therapeutic strategies for pancreatic CSCs. PMID:26918054

Hedgehog Signaling Regulates Epithelial-Mesenchymal Transition in Pancreatic Cancer Stem-Like Cells.

PubMed

Wang, Feng; Ma, Ling; Zhang, Zhengkui; Liu, Xiaoran; Gao, Hongqiao; Zhuang, Yan; Yang, Pei; Kornmann, Marko; Tian, Xiaodong; Yang, Yinmo

2016-01-01

Hedgehog (Hh) signaling is crucially involved in tumorigenesis. This study aimed to assess the role of Hh signaling in the regulation of epithelial-mesenchymal transition (EMT), stemness properties and chemoresistance of human pancreatic Panc-1 cancer stem cells (CSCs). Panc-1 cells were transfected with recombinant lentiviral vectors to silence SMO and serum-free floating-culture system was used to isolate Panc-1 tumorspheres. The expression of CSC and EMT markers was detected by flow cytometry, real-time RT-PCR and Western blot analysis. Malignant behaviors of Panc-1 CSC were evaluated by tumorigenicity assays and nude mouse lung metastasis model. We found that tumorspheres derived from pancreatic cancer cell line Panc-1 possessed self-renewal, differentiation and stemness properties. Hh pathway and EMT were active in Panc-1 tumorspheres. Inhibition of Hh signaling by SMO knockdown inhibited self-renewal, EMT, invasion, chemoresistance, pulmonary metastasis, tumorigenesis of pancreatic CSCs. In conclusion, Hh signaling contributes to the maintenance of stem-like properties and chemoresistance of pancreatic CSC and promotes the tumorigenesis and metastasis of pancreatic cancer. Hh pathway is a potential molecular target for the development of therapeutic strategies for pancreatic CSCs.

Gestational low-protein intake enhances whole-kidney miR-192 and miR-200 family expression and epithelial-to-mesenchymal transition in rat adult male offspring.

PubMed

Sene, Letícia B; Rizzi, Victor Hugo Gonçalves; Gontijo, José A R; Boer, Patricia A

2018-05-22

Studies have shown that adult offspring of mothers fed a protein-restricted diet during pregnancy present a pronounced reduction of nephron number associated with decreased fractional urinary sodium excretion and arterial hypertension. Additionally, recent advances in our understanding of the molecular pathways that govern the association of gestational nutritional restriction, intrauterine growth retardation and inflammation with impaired nephrogenesis, nephron underdosing and kidney fibrosis point to the epithelial to mesenchymal transition (EMT) as a common factor. In the current study, protein and sodium urinary excretion rates were evaluated in rats, and immunohistochemistry and western blot techniques were used to characterize kidney structure changes in 16 week old male offspring of mothers fed a low-protein diet during pregnancy (LP group) compared with age-matched (NP) controls. We also verified the expression of miRNA, mRNA and protein markers of fibrosis and the EMT in whole kidney prepared from LP offspring. We found, surprisingly, that arterial hypertension and long-term hyperfiltration, manifest by proteinuria, were associated with increased renal miR-192 and miR-200 family expression in 16 week old LP relative to age-matched NP rats. Measurement of kidney fibrosis and EMT-related protein markers, by histochemistry and immunoblot techniques, showed a significant rise of TGF-β1 and type-I collagen content in glomeruli and tubulointerstitial areas, accompanied by enhanced fibronectin and ZEB1 and decreased E-cadherin immunoreactivity in 16 week old LP offspring. The results were partially confirmed by increased gene (mRNA) expression of collagen 1α1, collagen 1α2 and ZEB1 in LP whole kidneys compared with those of age-matched NP offspring. In view of the presumed functional overload in the remaining nephrons, we suggest that hypertension and proteinuria development following maternal protein restriction may be a preponderant factor for EMT and structural kidney changes in LP offspring. However, our study was not wholly able to establish the precise role of miRNAs in LP kidney disorders. Thus, further studies will be required to assess the contribution of the miR family to renal injury in a gestational protein-restricted model of fetal programming. © 2018. Published by The Company of Biologists Ltd.

Cordyceps sinensis attenuates renal fibrosis and suppresses BAG3 induction in obstructed rat kidney.

PubMed

Du, Feng; Li, Si; Wang, Tian; Zhang, Hai-Yan; Zong, Zhi-Hong; Du, Zhen-Xian; Li, De-Tian; Wang, Hua-Qin; Liu, Bo; Miao, Jia-Ning; Bian, Xiao-Hui

2015-01-01

BAG3 regulates a number of cellular processes, including cell proliferation, apoptosis, adhesion and migration, and epithelial-mesenchymal transition (EMT). However, the role of BAG3 in renal tubular EMT and renal interstitial fibrosis remains elusive. This study aimed to examine the dynamic expression of BAG3 during renal fibrosis, and to investigate the efficacy of Cordyceps sinensis (C. sinensis) on renal fibrosis. A rat model of unilateral ureteral obstruction (UUO) was established, and the expression of BAG3 and α-SMA, and the efficacy of C. sinensis on renal fibrosis induced by UUO were examined. The results showed that UUO led to collagen accumulation, which was significantly suppressed by C. sinensis. UUO increased the expression of BAG3 and α-SMA, a mesenchymal marker, while UUO induced BAG3 and α-SMA expression was significantly inhibited by C. sinensis. In addition, immunohistochemical staining demonstrated that BAG3 immunoreactivity was restricted to tubular epithelium. In conclusion, BAG3 is a potential target for the prevention and/or treatment of renal fibrosis, and C. Sinensis is a promising agent for renal fibrosis.

CRISPR/Cas9n-Mediated Deletion of the Snail 1Gene (SNAI1) Reveals Its Role in Regulating Cell Morphology, Cell-Cell Interactions, and Gene Expression in Ovarian Cancer (RMG-1) Cells.

PubMed

Haraguchi, Misako; Sato, Masahiro; Ozawa, Masayuki

2015-01-01

Snail1 is a transcription factor that induces the epithelial to mesenchymal transition (EMT). During EMT, epithelial cells lose their junctions, reorganize their cytoskeletons, and reprogram gene expression. Although Snail1 is a prominent repressor of E-cadherin transcription, its precise roles in each of the phenomena of EMT are not completely understood, particularly in cytoskeletal changes. Previous studies have employed gene knockdown systems to determine the functions of Snail1. However, incomplete protein knockdown is often associated with these systems, which may cause incorrect interpretation of the data. To more precisely evaluate the functions of Snail1, we generated a stable cell line with a targeted ablation of Snail1 (Snail1 KO) by using the CRISPR/Cas9n system. Snail1 KO cells show increased cell-cell adhesion, decreased cell-substrate adhesion and cell migration, changes to their cytoskeletal organization that include few stress fibers and abundant cortical actin, and upregulation of epithelial marker genes such as E-cadherin, occludin, and claudin-1. However, morphological changes were induced by treatment of Snail1 KO cells with TGF-beta. Other transcription factors that induce EMT were also induced by treatment with TGF-beta. The precise deletion of Snail1 by the CRISPR/Cas9n system provides clear evidence that loss of Snail1 causes changes in the actin cytoskeleton, decreases cell-substrate adhesion, and increases cell-cell adhesion. Treatment of RMG1 cells with TGF-beta suggests redundancy among the transcription factors that induce EMT.

Pirfenidone may revert the epithelial-to-mesenchymal transition in human lung adenocarcinoma.

PubMed

Kurimoto, Ryota; Ebata, Takahiro; Iwasawa, Shunichiro; Ishiwata, Tsukasa; Tada, Yuji; Tatsumi, Koichiro; Takiguchi, Yuichi

2017-07-01

The epithelial-to-mesenchymal transition (EMT) in cancer is associated with invasion, metastasis and chemoresistance. Recent studies have revealed the increased expression of programmed death-ligand 1 (PD-L1) in cells undergoing EMT. The underlying mechanism of EMT involves transforming growth factor-β (TGF-β) and fibroblast growth factor-2 (FGF-2). Pirfenidone and the known EMT-suppressor nintedanib suppress pulmonary fibrosis partially through suppression of TGF-β. The present study aimed to determine whether pirfenidone has the potential to induce EMT-reversion, using nintedanib as a reference. The human lung adenocarcinoma cell lines A-549, HCC-827, and PC-9 were treated with TGF-β and FGF-2 to induce EMT. The EMT-induced cells were further treated with pirfenidone or nintedanib. Phenotypic alterations associated with EMT were assessed by examining the following: i) The expression levels of E-cadherin, vimentin, fibronectin and slug, using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and fluorescent immunohistochemistry; ii) cell motility via wound-healing assays; and iii) the expression of PD-L1 using RT-qPCR. The combination of TGF-β and FGF-2 successfully induced EMT in all three cell lines, characterized by a significant reduction in E-cadherin expression in the A-549 and HCC-827 cells, increased expression levels of vimentin, fibronectin, slug and PD-L1, and increased cell motility in all three cell lines. Pirfenidone and nintedanib reverted all of these phenotypes, with the exception of unaltered E-cadherin expression in all three cell lines, and inconsistent expression of vimentin in the HCC-827 and PC-9 cells. Thus, pirfenidone and nintedanib have the ability to induce EMT-reversion in human lung adenocarcinoma.

The enforced expression of c-Myc in pig fibroblasts triggers mesenchymal-epithelial transition (MET) via F-actin reorganization and RhoA/Rock pathway inactivation.

PubMed

Shi, Jun-Wen; Liu, Wei; Zhang, Ting-Ting; Wang, Sheng-Chun; Lin, Xiao-Lin; Li, Jing; Jia, Jun-Shuang; Sheng, Hong-Fen; Yao, Zhi-Fang; Zhao, Wen-Tao; Zhao, Zun-Lan; Xie, Rao-Ying; Yang, Sheng; Gao, Fei; Fan, Quan-Rong; Zhang, Meng-Ya; Yue, Min; Yuan, Jin; Gu, Wei-Wang; Yao, Kai-Tai; Xiao, Dong

2013-04-01

In previous studies from other labs it has been well demonstrated that the ectopic expression of c-Myc in mammary epithelial cells can induce epithelial-mesenchymal transition (EMT), whereas in our pilot experiment, epithelial-like morphological changes were unexpectedly observed in c-Myc-expressing pig fibroblasts [i.e., porcine embryonic fibroblasts (PEFs) and porcine dermal fibroblasts (PDFs)] and pig mesenchymal stem cells, suggesting that the same c-Myc gene is entitled to trigger EMT in epithelial cells and mesenchymal-epithelial transition (MET) in fibroblasts. This prompted us to characterize the existence of a MET in c-Myc-expressing PEFs and PDFs at the molecular level. qRT-PCR, immunofluorescence and western blot analysis illustrated that epithelial-like morphological changes were accompanied by the increased expression of epithelial markers [such as cell adhesion proteins (E-cadherin, α-catenin and Bves), tight junction protein occludin and cytokeratins (Krt8 and Krt18)], the reduced expression of mesenchymal markers [vimentin, fibronectin 1 (FN1), snail1, collagen family of proteins (COL1A1, COL5A2) and matrix metalloproteinase (MMP) family (MMP12 and MMP14)] and the decreased cell motility and increased cell adhesion in c-Myc-expressing PEFs and PDFs. Furthermore, the ectopic expression of c-Myc in pig fibroblasts disrupted the stress fiber network, suppressed the formation of filopodia and lamellipodia, and resulted in RhoA/Rock pathway inactivation, which finally participates in epithelial-like morphological conversion. Taken together, these findings demonstrate, for the first time, that the enforced expression of c-Myc in fibroblasts can trigger MET, to which cytoskeleton depolymerization and RhoA/Rock pathway inactivation contribute.

Therapeutic Activity of Anti-AXL Antibody against Triple-Negative Breast Cancer Patient-Derived Xenografts and Metastasis.

PubMed

Leconet, Wilhem; Chentouf, Myriam; du Manoir, Stanislas; Chevalier, Clément; Sirvent, Audrey; Aït-Arsa, Imade; Busson, Muriel; Jarlier, Marta; Radosevic-Robin, Nina; Theillet, Charles; Chalbos, Dany; Pasquet, Jean-Max; Pèlegrin, André; Larbouret, Christel; Robert, Bruno

2017-06-01

Purpose: AXL receptor tyrosine kinase has been described as a relevant molecular marker and a key player in invasiveness, especially in triple-negative breast cancer (TNBC). Experimental Design: We evaluate the antitumor efficacy of the anti-AXL monoclonal antibody 20G7-D9 in several TNBC cell xenografts or patient-derived xenograft (PDX) models and decipher the underlying mechanisms. In a dataset of 254 basal-like breast cancer samples, genes correlated with AXL expression are enriched in EMT, migration, and invasion signaling pathways. Results: Treatment with 20G7-D9 inhibited tumor growth and bone metastasis formation in AXL-positive TNBC cell xenografts or PDX, but not in AXL-negative PDX, highlighting AXL role in cancer growth and invasion. In vitro stimulation of AXL-positive cancer cells by its ligand GAS6 induced the expression of several EMT-associated genes ( SNAIL, SLUG , and VIM ) through an intracellular signaling implicating the transcription factor FRA-1, important in cell invasion and plasticity, and increased their migration/invasion capacity. 20G7-D9 induced AXL degradation and inhibited all AXL/GAS6-dependent cell signaling implicated in EMT and in cell migration/invasion. Conclusions: The anti-AXL antibody 20G7-D9 represents a promising therapeutic strategy in TNBC with mesenchymal features by inhibiting AXL-dependent EMT, tumor growth, and metastasis formation. Clin Cancer Res; 23(11); 2806-16. ©2016 AACR . ©2016 American Association for Cancer Research.

FBXO11 promotes ubiquitination of the Snail family of transcription factors in cancer progression and epidermal development.

PubMed

Jin, Yue; Shenoy, Anitha K; Doernberg, Samuel; Chen, Hao; Luo, Huacheng; Shen, Huangxuan; Lin, Tong; Tarrash, Miriam; Cai, Qingsong; Hu, Xin; Fiske, Ryan; Chen, Ting; Wu, Lizi; Mohammed, Kamal A; Rottiers, Veerle; Lee, Siu Sylvia; Lu, Jianrong

2015-06-28

The Snail family of transcription factors are core inducers of epithelial-to-mesenchymal transition (EMT). Here we show that the F-box protein FBXO11 recognizes and promotes ubiquitin-mediated degradation of multiple Snail family members including Scratch. The association between FBXO11 and Snai1 in vitro is independent of Snai1 phosphorylation. Overexpression of FBXO11 in mesenchymal cells reduces Snail protein abundance and cellular invasiveness. Conversely, depletion of endogenous FBXO11 in epithelial cancer cells causes Snail protein accumulation, EMT, and tumor invasion, as well as loss of estrogen receptor expression in breast cancer cells. Expression of FBXO11 is downregulated by EMT-inducing signals TGFβ and nickel. In human cancer, high FBXO11 levels correlate with expression of epithelial markers and favorable prognosis. The results suggest that FBXO11 sustains the epithelial state and inhibits cancer progression. Inactivation of FBXO11 in mice leads to neonatal lethality, epidermal thickening, and increased Snail protein levels in epidermis, validating that FBXO11 is a physiological ubiquitin ligase of Snail. Moreover, in C. elegans, the FBXO11 mutant phenotype is attributed to the Snail factors as it is suppressed by inactivation/depletion of Snail homologs. Collectively, these findings suggest that the FBXO11-Snail regulatory axis is evolutionarily conserved and critically governs carcinoma progression and mammalian epidermal development. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Ephrin reverse signaling mediates palatal fusion and epithelial-to-mesenchymal transition independently of Tgfß3.

PubMed

Serrano, Maria J; Liu, Jingpeng; Svoboda, Kathy K H; Nawshad, Ali; Benson, M Douglas

2015-12-01

The mammalian secondary palate forms from shelves of epithelia-covered mesenchyme that meet at midline and fuse. The midline epithelial seam (MES) is thought to degrade by apoptosis, epithelial-to-mesenchymal transition (EMT), or both. Failure to degrade the MES blocks fusion and causes cleft palate. It was previously thought that transforming growth factor ß3 (Tgfß3) is required to initiate fusion. Members of the Eph tyrosine kinase receptor family and their membrane-bound ephrin ligands are expressed on the MES. We demonstrated that treatment of mouse palates with recombinant EphB2/Fc to activate ephrin reverse signaling (where the ephrin acts as a receptor and transduces signals from its cytodomain) was sufficient to cause mouse palatal fusion when Tgfß3 signaling was blocked by an antibody against Tgfß3 or by an inhibitor of the TgfßrI serine/threonine receptor kinase. Cultured palatal epithelial cells traded their expression of epithelial cell markers for that of mesenchymal cells and became motile after treatment with EphB2/Fc. They concurrently increased their expression of the EMT-associated transcription factors Snail, Sip1, and Twist1. EphB2/Fc did not cause apoptosis in these cells. These data reveal that ephrin reverse signaling directs palatal fusion in mammals through a mechanism that involves EMT but not apoptosis and activates a gene expression program not previously associated with ephrin reverse signaling. © 2015 Wiley Periodicals, Inc.

A 3D in vitro model to explore the inter-conversion between epithelial and mesenchymal states during EMT and its reversion.

PubMed

Bidarra, S J; Oliveira, P; Rocha, S; Saraiva, D P; Oliveira, C; Barrias, C C

2016-06-03

Epithelial-to-mesenchymal transitions (EMT) are strongly implicated in cancer dissemination. Intermediate states, arising from inter-conversion between epithelial (E) and mesenchymal (M) states, are characterized by phenotypic heterogeneity combining E and M features and increased plasticity. Hybrid EMT states are highly relevant in metastatic contexts, but have been largely neglected, partially due to the lack of physiologically-relevant 3D platforms to study them. Here we propose a new in vitro model, combining mammary E cells with a bioengineered 3D matrix, to explore phenotypic and functional properties of cells in transition between E and M states. Optimized alginate-based 3D matrices provided adequate 3D microenvironments, where normal epithelial morphogenesis was recapitulated, with formation of acini-like structures, similar to those found in native mammary tissue. TGFβ1-driven EMT in 3D could be successfully promoted, generating M-like cells. TGFβ1 removal resulted in phenotypic switching to an intermediate state (RE cells), a hybrid cell population expressing both E and M markers at gene/protein levels. RE cells exhibited increased proliferative/clonogenic activity, as compared to M cells, being able to form large colonies containing cells with front-back polarity, suggesting a more aggressive phenotype. Our 3D model provides a powerful tool to investigate the role of the microenvironment on metastable EMT stages.

The Anoikis Effector Bit1 Inhibits EMT through Attenuation of TLE1-Mediated Repression of E-Cadherin in Lung Cancer Cells

PubMed Central

Yao, Xin; Pham, Tri; Temple, Brandi; Gray, Selena; Cannon, Cornita; Chen, Renwei; Abdel-Mageed, Asim B.; Biliran, Hector

2016-01-01

The mitochondrial Bcl-2 inhibitor of transcription 1 (Bit1) protein is part of an anoikis-regulating pathway that is selectively dependent on integrins. We previously demonstrated that the caspase-independent apoptotic effector Bit1 exerts tumor suppressive function in lung cancer in part by inhibiting anoikis resistance and anchorage-independent growth in vitro and tumorigenicity in vivo. Herein we show a novel function of Bit1 as an inhibitor cell migration and epithelial–mesenchymal transition (EMT) in the human lung adenocarcinoma A549 cell line. Suppression of endogenous Bit1 expression via siRNA and shRNA strategies promoted mesenchymal phenotypes, including enhanced fibroblastoid morphology and cell migratory potential with concomitant downregulation of the epithelial marker E-cadherin expression. Conversely, ectopic Bit1 expression in A549 cells promoted epithelial transition characterized by cuboidal-like epithelial cell phenotype, reduced cell motility, and upregulated E-cadherin expression. Specific downregulation of E-cadherin in Bit1-transfected cells was sufficient to block Bit1-mediated inhibition of cell motility while forced expression of E-cadherin alone attenuated the enhanced migration of Bit1 knockdown cells, indicating that E-cadherin is a downstream target of Bit1 in regulating cell motility. Furthermore, quantitative real-time PCR and reporter analyses revealed that Bit1 upregulates E-cadherin expression at the transcriptional level through the transcriptional regulator Amino-terminal Enhancer of Split (AES) protein. Importantly, the Bit1/AES pathway induction of E-cadherin expression involves inhibition of the TLE1-mediated repression of E-cadherin, by decreasing TLE1 corepressor occupancy at the E-cadherin promoter as revealed by chromatin immunoprecipitation assays. Consistent with its EMT inhibitory function, exogenous Bit1 expression significantly suppressed the formation of lung metastases of A549 cells in an in vivo experimental metastasis model. Taken together, our studies indicate Bit1 is an inhibitor of EMT and metastasis in lung cancer and hence can serve as a molecular target in curbing lung cancer aggressiveness. PMID:27655370

Detection and characterization of invasive circulating tumor cells derived from men with metastatic castration-resistant prostate cancer.

PubMed

Friedlander, Terence W; Ngo, Vy T; Dong, Huan; Premasekharan, Gayatri; Weinberg, Vivian; Doty, Shaun; Zhao, Qiang; Gilbert, Elizabeth G; Ryan, Charles J; Chen, Wen-Tien; Paris, Pamela L

2014-05-15

The Vitatex cell-adhesion matrix (CAM) platform allows for isolation of invasive circulating tumor cells (iCTCs). Here we sought to determine the utility of prostate-specific membrane antigen (PSMA) as a metastatic castration-resistant prostate cancer (mCRPC) iCTC biomarker, to identify solitary cells and clusters of iCTCs expressing either epithelial, mesenchymal, or stem cell markers, and to explore the feasibility of iCTC epigenomic analysis. CTCs were isolated and enumerated simultaneously using the Vitatex and CellSearch platforms in 23 men with mCRPC. CAM-avid iCTCs were identified as nucleated cells capable of CAM uptake, but without detectable expression of hematopoietic lineage (HL) markers including CD45. iCTCs were enumerated immunocytochemically (ICC) and by flow cytometry. Whole-genome methylation status was determined for iCTCs using the Illumina HumanMethylation27 BeadChip. Thirty-four samples were collected for iCTC analysis. A median of 27 (range 0-800) and 23 (range 2-390) iCTCs/mL were detected by ICC and flow, respectively. In a subset of 20 samples, a median of seven CTCs/mL (range 0-85) were detected by the CellSearch platform compared to 26 by the CAM platform. iCTC clusters were observed in 17% of samples. iCTCs expressing PSMA as well as markers of EMT and stemness were detectable. The iCTC methylation profile highly resembled mCRPC. More CTCs were recovered using the CAM platform than the CellSearch platform, and the CAM platform allowed for the detection of iCTC clusters, iCTCs expressing EMT and stem-cell markers, and characterization of the iCTC methylome. Correlation with clinical data in future studies may yield further insight into the functional significance of these findings. © 2013 UICC.

Cancer-associated fibroblasts promote epithelial-mesenchymal transition and EGFR-TKI resistance of non-small cell lung cancers via HGF/IGF-1/ANXA2 signaling.

PubMed

Yi, Yanmei; Zeng, Shanshan; Wang, Zhaotong; Wu, Minhua; Ma, Yuanhuan; Ye, Xiaoxia; Zhang, Biao; Liu, Hao

2018-03-01

The involvement of the tumor stromal cells in acquired resistance of non-small cell lung cancers (NSCLCs) to tyrosine kinase inhibitors (TKIs) has previously been reported, but the precise mechanism remains unclear. In the present study, we investigated the role and mechanism underlying Cancer-associated fibroblasts (CAFs) in TKI resistance of NSCLCs. In vitro and in vivo experiments showed that HCC827 and PC9 cells, non-small cell lung cancer cells with EGFR-activating mutations, became resistant to the EGFR-TKI gefitinib when cultured with CAFs isolated from NSCLC tissues. Moreover, we showed that CAFs could induce epithelial-mesenchymal transition (EMT) phenotype of HCC827 and PC9 cells, with an associated change in the expression of epithelial to mesenchymal transition markers. Using proteomics-based method, we identified that CAFs significantly increased the expression of the Annexin A2 (ANXA2). More importantly, knockdown of ANXA2 completely reversed EMT phenotype and gefitinib resistance induced by CAFs. Furthermore, we found that CAFs increased the expression and phosphorylation of ANXA2 by secretion of growth factors HGF and IGF-1 and by activation of the corresponding receptors c-met and IGF-1R. Dual inhibition of HGF/c-met and IGF-1/IGF-1R pathways could significantly suppress ANXA2, and markedly reduced CAFs-induced EMT and gefitinib resistance. Taken together, these findings indicate that CAFs promote EGFR-TKIs resistance through HGF/IGF-1/ANXA2/EMT signaling and may be an ideal therapeutic target in NSCLCs with EGFR-activating mutations. Copyright © 2017 Elsevier B.V. All rights reserved.

Curcumin targets fibroblast–tumor cell interactions in oral squamous cell carcinoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dudás, József, E-mail: jozsef.dudas@i-med.ac.at; Fullár, Alexandra, E-mail: fullarsz@gmail.com; 1st Department of Pathology and Experimental Cancer Research, Semmelweis University, Üllői út 26, 1085 Budapest

Co-culture of periodontal ligament fibroblasts (PDLs) and SCC-25 oral squamous carcinoma cells (OSCC) results in conversion of PDLs into carcinoma-associated fibroblasts (CAFs) and induces epithelial-to mesenchymal transition (EMT) of OSCC tumor cells. We hypothesized that Curcumin targets this dynamic mutual interaction between CAFs and tumor cells. Normal and 2 μM Curcumin-treated co-culture were performed for 4 days, followed by analysis of tumor cell invasivity, mRNA/protein expression of EMT-markers and mediators, activity measure of matrix metalloproteinase 9 (MMP-9), and western blot analysis of signal transduction in tumor cells and fibroblasts. In Curcumin-treated co-culture, in tumor cells, the levels of nuclear factormore » κB (NFκBα) and early response kinase (ERK)—decreased, in fibroblasts, integrin αv protein synthesis decreased compared to corresponding cells in normal co-culture. The signal modulatory changes induced by Curcumin caused decreased release of EMT-mediators in CAFs and reversal of EMT in tumor cells, which was associated with decreased invasion. These data confirm the palliative potential of Curcumin in clinical application. - Graphical abstract: Co-culture of periodontal ligament fibroblasts (PDLs) and SCC-25 oral squamous carcinoma cells (OSCC) results in conversion of PDLs into carcinoma-associated fibroblasts (CAFs) and induces epithelial-to mesenchymal transition (EMT) of tumor cells. Curcumin targets this dynamic mutual interaction between CAFs and tumor cells by inhibiting the production of EMT mediators in CAFs and by modification of intracellular signaling in tumor cells. This causes less invasivity and reversal of EMT in tumor cells. Highlights: ► Curcumin targets tumor–fibroblast interaction in head and neck cancer. ► Curcumin suppresses mediators of epithelial–mesenchymal transition. ► Curcumin decreases the invasivity of tumor cells.« less

Telomerase reverse transcriptase coordinates with the epithelial-to-mesenchymal transition through a feedback loop to define properties of breast cancer stem cells.

PubMed

El-Badawy, Ahmed; Ghoneim, Nehal I; Nasr, Mohamed A; Elkhenany, Hoda; Ahmed, Toka A; Ahmed, Sara M; El-Badri, Nagwa

2018-06-15

Telomerase and its core component, telomerase reverse transcriptase (hTERT), are critical for stem cell compartment integrity. Normal adult stem cells have the longest telomeres in a given tissue, a property mediated by high hTERT expression and high telomerase enzymatic activity. In contrast, cancer stem cells (CSCs) have short telomeres despite high expression of hTERT, indicating that the role of hTERT in CSCs is not limited to telomere elongation and/or maintenance. The function of hTERT in CSCs remains poorly understood. Here, we knocked down hTERT expression in CSCs and observed a morphological shift to a more epithelial phenotype, suggesting a role for hTERT in the epithelial-to-mesenchymal transition (EMT) of CSCs. Therefore, in this study, we systematically explored the relationship between hTERT and EMT and identified a reciprocal, bidirectional feedback loop between hTERT and EMT in CSCs. We found that hTERT expression is mutually exclusive to the mesenchymal phenotype and that, reciprocally, loss of the mesenchymal phenotype represses hTERT expression. We also showed that hTERT plays a critical role in the expression of key CSC markers and nuclear β-catenin localization, increases the percentage of cells with side-population properties, and upregulates the CD133 expression. hTERT also promotes chemoresistance properties, tumorsphere formation and other important functional CSC properties. Subsequently, hTERT knockdown leads to the loss of the above advantages, indicating a loss of CSC properties. Our findings suggest that targeting hTERT might improve CSCs elimination by transitioning them from the aggressive mesenchymal state to a more steady epithelial state, thereby preventing cancer progression. © 2018. Published by The Company of Biologists Ltd.

α-Hederin inhibits interleukin 6-induced epithelial-to-mesenchymal transition associated with disruption of JAK2/STAT3 signaling in colon cancer cells.

PubMed

Sun, Dongdong; Shen, Weixing; Zhang, Feng; Fan, Huisen; Xu, Changliang; Li, Liu; Tan, Jiani; Miao, Yunjie; Zhang, Haibin; Yang, Ye; Cheng, Haibo

2018-05-01

Colon cancer is the third most frequently diagnosed malignancy and has high morbidity worldwide. Epithelial-mesenchymal transition (EMT) has been increasingly implicated in colon cancer progression and metastasis. The present study was aimed to evaluate the potential antitumor activity of α-hederin, a monodesmosidic triterpenoid saponin isolated from Hedera helix, in human SW620 colon cancer cells stimulated with interleukin 6 (IL-6) for mimicking the tumor inflammatory microenvironment in vivo. Cell viability assay showed that IL-6 at 6.25 ng/ml significantly enhanced viability of SW620 cells, and thus this concentration was used to stimulate SW620 cells throughout this study. We observed that α-hederin concentration-dependently inhibited cell viability, migration and invasion in IL-6-treated SW620 cells. Moreover, α-hederin significantly restored IL-6-induced decrease in E-cadherin expression and abolished IL-6-induced increase in N-cadherin, vimentin, fibronectin, twist and snail at both mRNA and protein levels in SW620 cells. These data suggested that α-hederin suppressed IL-6-indcued EMT in colon cancer cells. Further molecular examinations showed that α-hederin inhibited phosphorylation of Janus Kinase 2 (JAK2) and Signal Transducer and Activator of Transcription 3(STAT3), and halted the nuclear translocation of phosphorylated STAT3 in IL-6-treated SW620 cells. In addition, JAK2/STAT3 signaling inhibitor AG490 not only produced similar inhibitory effects on EMT markers as α-hederin, but also synergistically enhanced α-hederin's inhibitory effects on EMT markers in IL-6-treated SW620 cells. Altogether, we demonstrated that α-hederin suppressed IL-6-induced EMT associated with disruption of JAK2/STAT3 signaling in colon cancer cells. Our data strongly suggested α-hederin as a promising candidate for intervention of colon cancer and metastasis. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

miR-300 inhibits epithelial to mesenchymal transition and metastasis by targeting Twist in human epithelial cancer

PubMed Central

2014-01-01

Background Epithelial-to-mesenchymal transition (EMT) is a key step of the progression of tumor cell metastasis. Recent work has demonstrated some miRNAs play critical roles in EMT. In this study, we focused on the roles of miR-300 in regulating EMT. Methods The expression levels of miR-300 were examined in epithelial carcinoma cells that underwent an EMT using quantitative reverse transcription-PCR. The role of miR-300 in EMT was investigated by transfection of the miR-300 mimic or inhibitor in natural epithelial-mesenchymal phenotype cell line pairs and in transforming growth factor (TGF) beta-induced EMT cell models. A luciferase reporter assay and a rescue experiment were conducted to confirm the target gene of miR-300. The efficacy of miR-300 against tumor invasion and metastasis was evaluated both in vitro and in vivo. Correlation analysis between miR-300 expression and the expression levels of its target gene, as well as tumor metastasis was performed in specimens from patients with head and neck squamous cell carcinoma (HNSCC). Results MiR-300 was found down-regulated in the HNSCC cells and breast cancer cells that underwent EMT. Ectopic expression of miR-300 effectively blocked TGF-beta-induced EMT and reversed the phenotype of EMT in HN-12 and MDA-MB-231 cells, but inhibition of miR-300 in the epithelial phenotype cells, HN-4 and MCF-7 cells, could induce EMT. The luciferase reporter assay and the rescue assay results showed that miR-300 directly targets the 3′UTR of Twist. Enforced miR-300 expression suppressed cell invasion in vitro and experimental metastasis in vivo. Clinically, miR-300 expression was found inversely correlated with Twist expression and reduced miR-300 was associated with metastasis in patient specimens. Conclusions Down-regulation of miR-300 is required for EMT initiation and maintenance. MiR-300 may negatively regulate EMT by direct targeting Twist and therefore inhibit cancer cell invasion and metastasis, which implicates miR-300 as an attractive candidate for cancer therapy. PMID:24885626

miR-300 inhibits epithelial to mesenchymal transition and metastasis by targeting Twist in human epithelial cancer.

PubMed

Yu, Jingshuang; Xie, Furong; Bao, Xin; Chen, Wantao; Xu, Qin

2014-05-24

Epithelial-to-mesenchymal transition (EMT) is a key step of the progression of tumor cell metastasis. Recent work has demonstrated some miRNAs play critical roles in EMT. In this study, we focused on the roles of miR-300 in regulating EMT. The expression levels of miR-300 were examined in epithelial carcinoma cells that underwent an EMT using quantitative reverse transcription-PCR. The role of miR-300 in EMT was investigated by transfection of the miR-300 mimic or inhibitor in natural epithelial-mesenchymal phenotype cell line pairs and in transforming growth factor (TGF) beta-induced EMT cell models. A luciferase reporter assay and a rescue experiment were conducted to confirm the target gene of miR-300. The efficacy of miR-300 against tumor invasion and metastasis was evaluated both in vitro and in vivo. Correlation analysis between miR-300 expression and the expression levels of its target gene, as well as tumor metastasis was performed in specimens from patients with head and neck squamous cell carcinoma (HNSCC). MiR-300 was found down-regulated in the HNSCC cells and breast cancer cells that underwent EMT. Ectopic expression of miR-300 effectively blocked TGF-beta-induced EMT and reversed the phenotype of EMT in HN-12 and MDA-MB-231 cells, but inhibition of miR-300 in the epithelial phenotype cells, HN-4 and MCF-7 cells, could induce EMT. The luciferase reporter assay and the rescue assay results showed that miR-300 directly targets the 3'UTR of Twist. Enforced miR-300 expression suppressed cell invasion in vitro and experimental metastasis in vivo. Clinically, miR-300 expression was found inversely correlated with Twist expression and reduced miR-300 was associated with metastasis in patient specimens. Down-regulation of miR-300 is required for EMT initiation and maintenance. MiR-300 may negatively regulate EMT by direct targeting Twist and therefore inhibit cancer cell invasion and metastasis, which implicates miR-300 as an attractive candidate for cancer therapy.

Zipper-interacting protein kinase promotes epithelial-mesenchymal transition, invasion and metastasis through AKT and NF-κB signaling and is associated with metastasis and poor prognosis in gastric cancer patients

PubMed Central

Wang, Zhu; Wang, Dong; Zhang, Longjuan; Su, Qiao; Lai, Yingrong; Li, Bin; Luo, Zexing; Chen, Xu; Chen, Yu; Huang, Xiaohui; Ma, Jieyi; Wang, Wenjian; Bi, Jiong; Guan, Xinyuan

2015-01-01

Zipper-interacting Protein Kinase (ZIPK) belongs to the death-associated protein kinase family. ZIPK has been characterized as a tumor suppressor in various tumors, including gastric cancer. On the other hand, ZIPK also promotes cell survival. In this study, both in vitro and in vivo assays indicated that ZIPK promoted cell growth, proliferation, migration, invasion, tumor formation and metastasis in nude mice. ZIPK induced epithelial-mesenchymal transition (EMT) with increasing expression of β-catenin, mesenchymal markers, Snail and Slug, and with decreasing expression of E-cadherin. Furthermore, ZIPK activated the AKT/IκB/NF-κB pathway, which can promote EMT and metastasis. Additionally, ZIPK expression was detected in human primary gastric cancer and their matched metastatic lymph node samples by immunohistochemistry. Increased expression of ZIPK in lymph node metastases was significantly associated with stage VI and abdominal organ invasion. Survival analysis revealed that patients with increased ZIPK expression in metastatic lymph nodes had poor disease-specific survival. Taken together, our study reveals that ZIPK is a pro-oncogenic factor, which promotes cancer metastasis. PMID:25831050

Osteopontin splice variants expression is involved on docetaxel resistance in PC3 prostate cancer cells.

PubMed

Nakamura, K D M; Tilli, T M; Wanderley, J L; Palumbo, A; Mattos, R M; Ferreira, A C; Klumb, C E; Nasciutti, L E; Gimba, E R

2016-02-01

Osteopontin (OPN) is a phosphoprotein that activates several aspects of tumor progression. Alternative splicing of the OPN primary transcript generates three splicing isoforms, OPNa, OPNb and OPNc. In this report, we investigated some cellular mechanisms by which OPN splice variants could mediate PC3 prostate cancer (PCa) cell survival and growth in response to docetaxel (DXT)-induced cell death. Cell survival before and after DXT treatment was analyzed by phase-contrast microscopy and crystal-violet staining assays. Quantitative real-time PCR and immunocytochemical staining assays were used to evaluate the putative involvement of epithelial-mesenchymal transition (EMT) and OPN isoforms on mediating PC3 cell survival. Upon DXT treatment, PC3 cells overexpressing OPNb or OPNc isoforms showed higher cell densities, compared to cells overexpressing OPNa and controls. Notably, cells overexpressing OPNb or OPNc isoforms showed a downregulated pattern of EMT epithelial cell markers, while mesenchymal markers were mostly upregulated in these experimental conditions. We concluded that OPNc or OPNb overexpression in PC3 cells can mediate resistance and cell survival features in response to DXT-induced cell death. Our data also provide evidence the EMT program could be one of the molecular mechanisms mediating survival in OPNb- or OPNc-overexpressing cells in response to DXT treatment. These data could further contribute to a better understanding of the mechanisms by which PCa cells acquire resistance to DXT treatment.

BCORL1 is an independent prognostic marker and contributes to cell migration and invasion in human hepatocellular carcinoma.

PubMed

Yin, Guozhi; Liu, Zhikui; Wang, Yufeng; Dou, Changwei; Li, Chao; Yang, Wei; Yao, Yingmin; Liu, Qingguang; Tu, Kangsheng

2016-02-15

The deregulation of E-cadherin has been considered as a leading cause of hepatocellular carcinoma (HCC) metastasis. BCL6 corepressor-like 1 (BCORL1) is a transcriptional corepressor and contributes to the repression of E-cadherin. However, the clinical significance of BCORL1 and its role in the metastasis of HCC remain unknown. Differentially expressed BCORL1 between HCC and matched tumor-adjacent tissues, HCC cell lines and normal hepatic cell line were detected by Western blot. The expression of BCORL1 was altered by siRNAs or lentivirus-mediated vectors. Transwell assays were performed to determine HCC cell invasion and migration. Increased expression of BCORL1 protein was detected in HCC specimens and cell lines. Clinical association analysis showed that BCORL1 protein was expressed at significant higher levels in HCC patients with multiple tumor nodes, venous infiltration and advanced TNM tumor stage. Survival analysis indicated that high expression of BCORL1 protein conferred shorter overall survival (OS) and recurrence-free survival (RFS) of HCC patients. Multivariate Cox regression analysis disclosed that BCORL1 expression was an independent prognostic marker for predicting survival of HCC patients. Our in vitro studies demonstrated that BCORL1 prominently promoted HCC cell migration and invasion. Otherwise, an inverse correlation between BCORL1 and E-cadherin expression was observed in HCC tissues. BCORL1 inversely regulated E-cadherin abundance and subsequently facilitated epithelial-mesenchymal transition (EMT) in HCC cells. Notably, the effect of BCORL1 knockdown on HCC cells was abrogated by E-cadherin silencing. BCORL1 may be a novel prognostic factor and promotes cell migration and invasion through E-cadherin repression-induced EMT in HCC.

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

PubMed

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.

Isochlorogenic Acid C Reverses Epithelial-Mesenchymal Transition via Down-regulation of EGFR Pathway in MDA-MB-231 cells.

PubMed

Yu, Ji-Kuen; Yue, Chia-Herng; Pan, Ying-Ru; Chiu, Yung-Wei; Liu, Jer-Yuh; Lin, Kun-I; Lee, Chia-Jen

2018-04-01

Epidermal growth factor receptor (EGFR) has been suggested to play an important role in survival, proliferation, migration, differentiation, and tumorigenesis of many cell types. Breast cancer patients with high EGFR expression have a poor prognosis. In this study, we investigated the molecular mechanism of the inhibitory effect of isochlorogenic acid c (ICAC) extracted from Lonicera japonica on elevated EGFR levels of the triple-negative breast cancer (TNBC) cell line, MDA-MB-231. The cell viability and cell-cycle analysis were evaluated using 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) assay and flow cytometry, respectively. The migration ability and invasiveness of ICAC-treated MDA-MB-231 were examined by migration and Matrigel invasion assay. The epithelial-mesenchymal-transition (EMT)-related protein expression was examined by western blotting and reverse transcriptase-polymerase chain reaction (RT-PCR). ICAC led to significant morphological changes and suppressed migration and invasion capacities of highly metastatic MDA-MB-231 cells. Western blot analysis for EGFR/EMT-associated proteins suggested that ICAC attenuated the mesenchymal traits as observed by up-regulation of epithelial markers and down-regulation of mesenchymal markers as well as decreased activities of matrix metalloproteinase-9 (MMP-9). These results suggested that the inhibitory effects of ICAC against EGFR-induced EMT and MDA-MB-231 cell invasion were dependent on the EGFR/ phospholipase Cγ (PLCγ)/extracellular regulated protein kinase ½ (ERK½)/slug signaling pathway. Therefore, the obtained results could provide us clues for the next therapeutic strategy in the treatment of TNBC. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

IL-6, IL-8, MMP-2, MMP-9 are overexpressed in Fanconi anemia cells through a NF-κB/TNF-α dependent mechanism.

PubMed

Epanchintsev, Alexey; Shyamsunder, Pavithra; Verma, Rama S; Lyakhovich, Alex

2015-12-01

Fanconi anemia (FA) is a rare autosomal recessive genetic disorder associated with a bone-marrow failure, genome instability, hypersensitivity to DNA crosslinking agents and a predisposition to cancer. Mutations have been documented in 16 FA genes that participate in the FA-BRCA DNA repair pathway, a fundamental pathway in the development of the disease and the presentation of its symptoms. FA cells have been characterized by an overproduction of cytokines, MAPKs, and Interleukins. Through this study we have identified the overexpression of additional secretory factors such as IL-6, IL-8, MMP-2, and MMP-9 in FA cells and in cells depleted of FANCA or FANCC and proved that their expression is under the control of NF-κB/TNF-α signaling pathways. We also demonstrated that these overexpressed secretory factors were effective in promoting the proliferation, migration, and invasion of surrounding tumor cells a fundamental event in the process of epithelial mesenchymal transition (EMT) and that they also modulated the expression of EMT markers such as E-cadherin and SNAIL. Overall our data suggest that the upregulation of EMT promoting factors in FA may contribute to predisposing FA patients to cancer, thereby providing new insights into possible therapeutic interventions. © 2014 Wiley Periodicals, Inc.

Slug/SNAI2 regulates cell proliferation and invasiveness of metastatic prostate cancer cell lines.

PubMed

Emadi Baygi, Modjtaba; Soheili, Zahra-Soheila; Essmann, Frank; Deezagi, Abdolkhaleg; Engers, Rainer; Goering, Wolfgang; Schulz, Wolfgang A

2010-08-01

Many metastatic cancers recapitulate the epithelial-to-mesenchymal transition (EMT) resulting in enhanced cell motility and invasiveness. The EMT is regulated by several transcription factors, including the zinc finger protein SNAI2, also named Slug, which appears to exert additional functions during development and cancer progression. We have studied the function of SNAI2 in prostate cancer cells. Quantitative RT-PCR analysis showed strong SNAI2 expression particularly in the PC-3 and PC3-16 prostate carcinoma cell lines. Knockdown of SNAI2 by specific siRNA induced changes in EMT markers and inhibited invasion of both cell lines into a matrigel matrix. SNAI2 siRNA-treated cells did not tolerate detachment from the culture plates, likely at least in part due to downregulation of integrin alpha6beta4. SNAI2 knockdown disturbed the microtubular and actin cytoskeletons, especially severely in PC-3 cells, resulting in grossly enlarged, flattened, and sometimes multinuclear cells. Knockdown also decreased cell proliferation, with a prominent G0/G1 arrest in PC3-16. Together, our data imply that SNAI2 exerts strong effects on the cytoskeleton and adhesion of those prostate cancer cells that express it and is necessary for their proliferation and invasiveness.

In vitro biomimetic platforms featuring a perfusion system and 3D spheroid culture promote the construction of tissue-engineered corneal endothelial layers.

PubMed

Li, Shanyi; Han, Yuting; Lei, Hao; Zeng, Yingxin; Cui, Zekai; Zeng, Qiaolang; Zhu, Deliang; Lian, Ruiling; Zhang, Jun; Chen, Zhe; Chen, Jiansu

2017-04-10

Corneal endothelial cells (CECs) are very important for the maintenance of corneal transparency. However, in vitro, CECs display limited proliferation and loss of phenotype via endothelial to mesenchymal transformation (EMT) and cellular senescence. In this study, we demonstrate that continuous supplementary nutrition using a perfusion culture bioreactor and three-dimensional (3D) spheroid culture can be used to improve CEC expansion in culture and to construct a tissue-engineered CEC layer. Compared with static culture, perfusion-derived CECs exhibited an increased proliferative ability as well as formed close cell-cell contact junctions and numerous surface microvilli. We also demonstrated that the CEC spheroid culture significantly down-regulated gene expression of the proliferation marker Ki67 and EMT-related markers Vimentin and α-SMA, whereas the gene expression level of the CEC marker ATP1A1 was significantly up-regulated. Furthermore, use of the perfusion system in conjunction with a spheroid culture on decellularized corneal scaffolds and collagen sheets promoted the generation of CEC monolayers as well as neo-synthesized ECM formation. This study also confirmed that a CEC spheroid culture on a curved collagen sheet with controlled physiological intraocular pressure could generate a CEC monolayer. Thus, our results show that the use of a perfusion system and 3D spheroid culture can promote CEC expansion and the construction of tissue-engineered corneal endothelial layers in vitro.

Molecular Innovations Toward Theranostics of Aggressive Prostate Cancer

DTIC Science & Technology

2017-11-01

burden to Department of Defense, Washington Headquarters Services , Directorate for Information Operations and Reports (0704-0188), 1215 Jefferson Davis...nearly real-time manner should offer a safe and effective treatment. This project carried out by a team of chemist, radiologist, and molecular tumor...compounds containing DAB2IP protein domain are the good candidates. Although, based on EMT markers, only CS-IV-81 was able to inhibit Vimentin expression

The A2B Adenosine Receptor Modulates the Epithelial– Mesenchymal Transition through the Balance of cAMP/PKA and MAPK/ERK Pathway Activation in Human Epithelial Lung Cells

PubMed Central

Giacomelli, Chiara; Daniele, Simona; Romei, Chiara; Tavanti, Laura; Neri, Tommaso; Piano, Ilaria; Celi, Alessandro; Martini, Claudia; Trincavelli, Maria L.

2018-01-01

The epithelial-mesenchymal transition (EMT) is a complex process in which cell phenotype switches from the epithelial to mesenchymal one. The deregulations of this process have been related with the occurrence of different diseases such as lung cancer and fibrosis. In the last decade, several efforts have been devoted in understanding the mechanisms that trigger and sustain this transition process. Adenosine is a purinergic signaling molecule that has been involved in the onset and progression of chronic lung diseases and cancer through the A2B adenosine receptor subtype activation, too. However, the relationship between A2BAR and EMT has not been investigated, yet. Herein, the A2BAR characterization was carried out in human epithelial lung cells. Moreover, the effects of receptor activation on EMT were investigated in the absence and presence of transforming growth factor-beta (TGF-β1), which has been known to promote the transition. The A2BAR activation alone decreased and increased the expression of epithelial markers (E-cadherin) and the mesenchymal one (Vimentin, N-cadherin), respectively, nevertheless a complete EMT was not observed. Surprisingly, the receptor activation counteracted the EMT induced by TGF-β1. Several intracellular pathways regulate the EMT: high levels of cAMP and ERK1/2 phosphorylation has been demonstrated to counteract and promote the transition, respectively. The A2BAR stimulation was able to modulated these two pathways, cAMP/PKA and MAPK/ERK, shifting the fine balance toward activation or inhibition of EMT. In fact, using a selective PKA inhibitor, which blocks the cAMP pathway, the A2BAR-mediated EMT promotion were exacerbated, and conversely the selective inhibition of MAPK/ERK counteracted the receptor-induced transition. These results highlighted the A2BAR as one of the receptors involved in the modulation of EMT process. Nevertheless, its activation is not enough to trigger a complete transition, its ability to affect different intracellular pathways could represent a mechanism at the basis of EMT maintenance/inhibition based on the extracellular microenvironment. Despite further investigations are needed, herein for the first time the A2BAR has been related to the EMT process, and therefore to the different EMT-related pathologies. PMID:29445342

Periostin activates pathways involved in epithelial-mesenchymal transition in adamantinomatous craniopharyngioma.

PubMed

Chen, Ming; Zheng, Shi-hao; Liu, Yi; Shi, Jin; Qi, Song-tao

2016-01-15

Periostin (POSTN) is an extracellular matrix protein (ECM) critical for epithelial-mesenchymal transitions (EMT) in several kinds of tumor cells. Previous studies have indicated that EMT exists in craniopharyngioma (CP), and expression of POSTN is a significant factor in the prognosis of CP. However, it has never been explored whether POSTN exists in CP, or how it activates CP's EMT. The expression of POSTN was examined in adamantinomatous craniopharyngioma (ACP) primary cells and tissues by immunohistochemistry, PCR and Western blot, respectively. The effects and mechanisms of POSTN on ACP cells' EMT were also analyzed. It was found that POSTN expression increased in ACP-associated fibroblasts. Overexpressed POSTN significantly elevated the EMT of ACP cells by regulating the expression of associated genes. More importantly, our further study revealed that the upregulated POSTN activated Akt signaling pathway to regulate the EMT. This study showed that POSTN is responsible for the EMT of ACP cells, and POSTN might be a potential molecular therapeutic target for ACP treatment in future. Copyright © 2015 Elsevier B.V. All rights reserved.

Overexpression of SDF-1 activates the NF-κB pathway to induce epithelial to mesenchymal transition and cancer stem cell-like phenotypes of breast cancer cells.

PubMed

Kong, Lingxin; Guo, Sufen; Liu, Chunfeng; Zhao, Yiling; Feng, Chong; Liu, Yunshuang; Wang, Tao; Li, Caijuan

2016-03-01

The formation of EMT and EMT-induced CSC-like phenotype is crucial for the metastasis of tumor cells. The stromal cell-derived factor-1 (SDF-1) is upregulated in various human carcinomas, which is closely associated with proliferation, migration, invasion and prognosis of malignancies. However, limited attention has been directed towards the effect of SDF-1 on epithelial to mesenchymal transition (EMT) or cancer stem cell (CSC)-like phenotype formation in breast cancer cells and the related mechanism. In the present study, we screened MCF-7 cells with low SDF-1 expression level for the purpose of evaluating whether SDF-1 is involved in EMT and CSC-like phenotype formation in MCF-7 cells. The pEGFP-N1-SDF-1 plasmid was transfected into MCF-7 cells, and the stably overexpressed SDF-1 in MCF-7 cells was confirmed by real-time PCR and western blot analysis. Colony formation assay, MTT, wound healing assay and Transwell invasion assay demonstrated that overexpression of SDF-1 significantly boosted the proliferation, migration and invasion of MCF-7 cells compared with parental (P<0.05). Flow cytometry analysis revealed a notable increase of CD44+/CD24- subpopulation in SDF-1 overexpressing MCF-7 cells (P<0.001), accompanied by the apparently elevated ALDH activity and the upregulation of the stem cell markers OCT-4, Nanog, and SOX2 compared with parental (P<0.01). Besides, western blot analysis and immunofluorescence assay observed the significant decreased expression of E-cadherin and enhanced expression of slug, fibronectin and vimentin in SDF-1 overexpressed MCF-7 cells in comparison with parental (P<0.01). Further study found that overexpression of SDF-1 induced the activation of NF-κB pathway in MCF-7 cells. Conversely, suppressing or silencing p65 expression by antagonist or RNA interference could remarkably increase the expression of E-cadherin in SDF-1 overexpressed MCF-7 cells (P<0.001). Overall, the above results indicated that overexpression of SDF-1 enhanced EMT by activating the NF-κB pathway of MCF-7 cells and further induced the formation of CSC-like phenotypes, ultimately promoting the proliferation and metastasis of MCF-7 cells. Therefore, SDF-1 may further be assessed as a potential target for gene therapy of breast cancer.

Stem Cell Conditioned Culture Media Attenuated Albumin-Induced Epithelial– Mesenchymal Transition in Renal Tubular Cells

PubMed Central

Hu, Junping; Zhu, Qing; Li, Pin-Lan; Wang, Weili; Yi, Fan; Li, Ningjun

2015-01-01

Background Proteinuria-induced epithelial-mesenchymal transition (EMT) plays an important role in progressive renal tubulointerstitial fibrosis in chronic renal disease. Stem cell therapy has been used for different diseases. Stem cell conditioned culture media (SCM) exhibits similar beneficial effects as stem cell therapy. The present study tested the hypothesis that SCM inhibits albumin-induced EMT in cultured renal tubular cells. Methods Rat renal tubular cells were treated with/without albumin (20 μmg/ml) plus SCM or control cell media (CCM). EMT markers and inflammatory factors were measured by Western blot and fluorescent images. Results Albumin induced EMT as shown by significant decreases in levels of epithelial marker E-cadherin, increases in mesenchymal markers fibroblast-specific protein 1 and α-smooth muscle actin, and elevations in collagen I. SCM inhibited all these changes. Meanwhile, albumin induced NF-κB translocation from cytosol into nucleus and that SCM blocked the nuclear translocation of NF-κB. Albumin also increased the levels of pro-inflammatory factor monocyte chemoattractant protein-1 (MCP)-1 by nearly 30 fold compared with control. SCM almost abolished albumin-induced increase of MCP-1. Conclusion These results suggest that SCM attenuated albumin-induced EMT in renal tubular cells via inhibiting activation of inflammatory factors, which may serve as a new therapeutic approach for chronic kidney diseases. PMID:25832005

TMEPAI regulates EMT in lung cancer cells by modulating the ROS and IRS-1 signaling pathways.

PubMed

Hu, Ying; He, Kai; Wang, Dongmei; Yuan, Xinwang; Liu, Yi; Ji, Hongbin; Song, Jianguo

2013-08-01

The epithelial-mesenchymal transition (EMT) has been implicated in various pathophysiological processes, including cancer cell migration and distal metastasis. Reactive oxygen species (ROS) and insulin receptor substrate-1 (IRS-1) are important in cancer progression and regulation of EMT. To explore the biological significance and regulatory mechanism of EMT, we determined the expression, the biological function and the signaling pathway of prostate transmembrane protein, androgen induced-1 (TMEPAI), during the induction of EMT and cell migration. Transforming growth factor (TGF)-β1 significantly upregulated the expression of TMEPAI during EMT in human lung adenocarcinoma. Depletion of TMEPAI abolished TGF-β1-induced downregulation of ferritin heavy chain and the subsequent generation of ROS, thus suppressing TGF-β1-induced EMT and cell migration. In addition, increased ROS production and overexpression of TMEPAI downregulated the level of IRS-1. Both the addition of H2O2 and IRS-1 small interfering RNA rescued the ability of TGF-β1 to induce EMT in TMEPAI-depleted cells. Remarkably, the levels of TMEPAI in lung tumor tissues are very high, whereas its expression in normal lung epithelium is very low. Moreover, TMEPAI expression was positively correlated with the cell mesenchymal phenotype and migration potential. Our work reveals that TMEPAI contributes to TGF-β1-induced EMT through ROS production and IRS-1 downregulation in lung cancer cells.

Protective effect of epigallocatechin-3-gallate (EGCG) via Nrf2 pathway against oxalate-induced epithelial mesenchymal transition (EMT) of renal tubular cells.

PubMed

Kanlaya, Rattiyaporn; Khamchun, Supaporn; Kapincharanon, Chompunoot; Thongboonkerd, Visith

2016-07-25

This study evaluated effect of oxalate on epithelial mesenchymal transition (EMT) and potential anti-fibrotic property of epigallocatechin-3-gallate (EGCG). MDCK renal tubular cells were incubated with 0.5 mM sodium oxalate for 24-h with/without 1-h pretreatment with 25 μM EGCG. Microscopic examination, immunoblotting and immunofluorescence staining revealed that oxalate-treated cells gained mesenchymal phenotypes by fibroblast-like morphological change and increasing expression of vimentin and fibronectin, while levels of epithelial markers (E-cadherin, occludin, cytokeratin and ZO-1) were decreased. EGCG pretreatment could prevent all these changes and molecular mechanisms underlying the prevention by EGCG were most likely due to reduced production of intracellular ROS through activation of Nrf2 signaling and increased catalase anti-oxidant enzyme. Knockdown of Nrf2 by small interfering RNA (siRNA) abrogated all the effects of EGCG, confirming that the EGCG protection against oxalate-induced EMT was mediated via Nrf2. Taken together, our data indicate that oxalate turned on EMT of renal tubular cells that could be prevented by EGCG via Nrf2 pathway. These findings also shed light onto development of novel therapeutics or preventive strategies of renal fibrosis in the future.

Epithelio-mesenchymal transitional attributes in oral sub-mucous fibrosis.

PubMed

Das, Raunak Kumar; Anura, Anji; Pal, Mousumi; Bag, Swarnendu; Majumdar, Subhadipa; Barui, Ananya; Chakraborty, Chandan; Ray, Ajoy Kumar; Sengupta, Sanghamitra; Paul, Ranjan Rashmi; Chatterjee, Jyotirmoy

2013-12-01

Evaluating molecular attributes in association with its epithelial and sub-epithelial changes of oral sub-mucous fibrosis is meaningful in exploring the plausibility of an epithelio-mesenchymal transition (EMT) and malignant potentiality of this pathosis. In this study histopathological and histochemical attributes for basement membrane and connective tissue in biopsies of oral sub-mucous fibrosis (n = 55) and normal oral mucosa (n = 16) were assessed and expressions of p63, E-cadherin, β-catenin, N-cadherin and TWIST were analyzed immunohistochemically. The p63 and its isoforms (TA and ∆N), PARD3, E-cadherin and β-catenin were also assessed transcriptomically by q-PCR and EMT players like TWIST1, ZEB1, MMP9 and micro-RNA 205 were searched in gene expression microarrays. Oral epithelium demonstrating impairment in progressive maturation in oral sub-mucous fibrosis concomitantly experienced an increase in basement membrane thickness and collagen deposition along with alteration in target molecular expressions. In comparison to non-dysplastic conditions dysplastic stages exhibited significant increase in p63 and p63∆N expressions whereas, E-cadherin and β-catenin exhibited loss from the membrane with concurrent increase in cytoplasm. Further the N-cadherin and TWIST were gained remarkably along with the appearance of nuclear accumulation features of β-catenin. The microarray search had noticed the up-regulation of TWIST1, ZEB1 and MMP9 along with down regulation of micro-RNA 205. The simultaneous increase in basement membrane thickness and sub-epithelial collagen deposition were the plausible indicators for increased matrix stiffness with expected impact on oral epithelial functional homoeostasis. This was corroborated with the increase in expressions of epithelial master regulator p63 and its oncogenic isoform (∆N) along with membranous loss of E-cadherin (EMT hallmark) and its associate β-catein and gain of mesenchymal markers like N-cadherin and TWIST. These also became indicative for the induction of epithelial to mesenchymal transitional mechanism in oral sub-mucous fibrosis when connoted here with the relevant modulation in expressions of EMT regulators. © 2013.

Mammalian-enabled (MENA) protein enhances oncogenic potential and cancer stem cell-like phenotype in hepatocellular carcinoma cells.

PubMed

Hu, Kunpeng; Huang, Pinzhu; Luo, Hui; Yao, Zhicheng; Wang, Qingliang; Xiong, Zhiyong; Lin, Jizong; Huang, He; Xu, Shilei; Zhang, Peng; Liu, Bo

2017-08-01

Mammalian-enabled (MENA) protein is an actin-regulatory protein that influences cell motility and adhesion. It is known to play a role in tumorigenicity of hepatocellular carcinoma (HCC) but the underlying molecular mechanism remains unknown. This study aimed to investigate the oncogenic potential of MENA and its capacity to regulate cancer stem cell (CSC)-like phenotypes in HCC cells. Real-time-PCR and western blot were used to assess mRNA and protein levels of target genes in human HCC tissue specimens and HCC cell lines, respectively. Stable MENA-overexpressing HCC cells were generated from HCC cell lines. Transwell cell migration and colony formation assays were employed to evaluate tumorigenicity. Ectopic expression of MENA significantly enhanced cell migration and colony-forming ability in HCC cells. Overexpression of MENA upregulated several hepatic progenitor/stem cell markers in HCC cells. A high MENA protein level was associated with high mRNA levels of MENA, CD133, cytokeratin 19 (CK19), and epithelial cell adhesion molecule (EpCAM) in human HCC tissues. Overexpression of MENA enhanced epithelial-to-mesenchymal transition (EMT) markers, extracellular signal-regulated kinases (ERK) phosphorylation, and the level of β-catenin in HCC cells. This study demonstrated that overexpression of MENA in HCC cells promoted stem cell markers, EMT markers, and tumorigenicity. These effects may involve, at least partially, the ERK and β-catenin signaling pathways.

Genetic Mutation and Exosome Signature of Human Papilloma Virus Associated Oropharyngeal Cancer

PubMed Central

Kannan, Anbarasu; Hertweck, Kate L.; Philley, Julie V.; Wells, Robert B.; Dasgupta, Santanu

2017-01-01

Human papilloma virus-16 (HPV-16) associated oropharyngeal cancer (HPVOPC) is increasing alarmingly in the United States. We performed whole genome sequencing of a 44 year old, male HPVOPC subject diagnosed with moderately differentiated tonsillar carcinoma. We identified new somatic mutation in MUC16 (A.k.a. CA-125), MUC12, MUC4, MUC6, MUC2, SIRPA, HLA-DRB1, HLA-A and HLA-B molecules. Increased protein expression of MUC16, SIRPA and decreased expression of HLA-DRB1 was further demonstrated in this HPVOPC subject and an additional set of 15 HPVOPC cases. Copy number gain (3 copies) was also observed for MUC2, MUC4, MUC6 and SIRPA. Enhanced expression of MUC16, SIRPA and HPV-16-E7 protein was detectable in the circulating exosomes of numerous HPVOPC subjects. Treatment of non-tumorigenic mammary epithelial cells with exosomes derived from aggressive HPVOPC cells harboring MUC16, SIRPA and HPV-16-E7 proteins augmented invasion and induced epithelial to mesenchymal transition (EMT) accompanied by an increased expression ratio of the EMT markers Vimentin/E-cadherin. Exosome based screening of key HPVOPC associated molecules could be beneficial for early cancer diagnosis, monitoring and surveillance. PMID:28383029

VEGF elicits epithelial-mesenchymal transition (EMT) in prostate intraepithelial neoplasia (PIN)-like cells via an autocrine loop

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gonzalez-Moreno, Oscar; Lecanda, Jon; Green, Jeffrey E.

2010-02-15

Vascular endothelial growth factor (VEGF) is overexpressed during the transition from prostate intraepithelial neoplasia (PIN) to invasive carcinoma. We have mimicked such a process in vitro using the PIN-like C3(1)/Tag-derived Pr-111 cell line, which expresses low levels of VEGF and exhibits very low tumorigenicity in vivo. Elevated expression of VEGF164 in Pr-111 cells led to a significant increase in tumorigenicity, invasiveness, proliferation rates and angiogenesis. Moreover, VEGF164 induced strong changes in cell morphology and cell transcriptome through an autocrine mechanism, with changes in TGF-beta1- and cytoskeleton-related pathways, among others. Further analysis of VEGF-overexpressing Pr-111 cells or following exogenous addition ofmore » recombinant VEGF shows acquisition of epithelial-mesenchymal transition (EMT) features, with an increased expression of mesenchymal markers, such as N-cadherin, Snail1, Snail2 (Slug) and vimentin, and a decrease in E-cadherin. Administration of VEGF led to changes in TGF-beta1 signaling, including reduction of Smad7 (TGF-beta inhibitory Smad), increase in TGF-betaR-II, and translocation of phospho-Smad3 to the nucleus. Our results suggest that increased expression of VEGF in malignant cells during the transition from PIN to invasive carcinoma leads to EMT through an autocrine loop, which would promote tumor cell invasion and motility. Therapeutic blockade of VEGF/TGF-beta1 in PIN lesions might impair not only tumor angiogenesis, but also the early dissemination of malignant cells outside the epithelial layer.« less

Tumour-derived alkaline phosphatase regulates tumour growth, epithelial plasticity and disease-free survival in metastatic prostate cancer

PubMed Central

Rao, S R; Snaith, A E; Marino, D; Cheng, X; Lwin, S T; Orriss, I R; Hamdy, F C; Edwards, C M

2017-01-01

Background: Recent evidence suggests that bone-related parameters are the main prognostic factors for overall survival in advanced prostate cancer (PCa), with elevated circulating levels of alkaline phosphatase (ALP) thought to reflect the dysregulated bone formation accompanying distant metastases. We have identified that PCa cells express ALPL, the gene that encodes for tissue nonspecific ALP, and hypothesised that tumour-derived ALPL may contribute to disease progression. Methods: Functional effects of ALPL inhibition were investigated in metastatic PCa cell lines. ALPL gene expression was analysed from published PCa data sets, and correlated with disease-free survival and metastasis. Results: ALPL expression was increased in PCa cells from metastatic sites. A reduction in tumour-derived ALPL expression or ALP activity increased cell death, mesenchymal-to-epithelial transition and reduced migration. Alkaline phosphatase activity was decreased by the EMT repressor Snail. In men with PCa, tumour-derived ALPL correlated with EMT markers, and high ALPL expression was associated with a significant reduction in disease-free survival. Conclusions: Our studies reveal the function of tumour-derived ALPL in regulating cell death and epithelial plasticity, and demonstrate a strong association between ALPL expression in PCa cells and metastasis or disease-free survival, thus identifying tumour-derived ALPL as a major contributor to the pathogenesis of PCa progression. PMID:28006818

Holding Tight: Cell Junctions and Cancer Spread.

PubMed

Knights, Alexander J; Funnell, Alister P W; Crossley, Merlin; Pearson, Richard C M

2012-01-01

Cell junctions are sites of intercellular adhesion that maintain the integrity of epithelial tissue and regulate signalling between cells. These adhesive junctions are comprised of protein complexes that serve to establish an intercellular cytoskeletal network for anchoring cells, in addition to regulating cell polarity, molecular transport and communication. The expression of cell adhesion molecules is tightly controlled and their downregulation is essential for epithelial-mesenchymal transition (EMT), a process that facilitates the generation of morphologically and functionally diverse cell types during embryogenesis. The characteristics of EMT are a loss of cell adhesion and increased cellular mobility. Hence, in addition to its normal role in development, dysregulated EMT has been linked to cancer progression and metastasis, the process whereby primary tumors migrate to invasive secondary sites in the body. This paper will review the current understanding of cell junctions and their role in cancer, with reference to the abnormal regulation of junction protein genes. The potential use of cell junction molecules as diagnostic and prognostic markers will also be discussed, as well as possible therapies for adhesive dysregulation.

Col1A1 Production and Apoptotic Resistance in TGF-β1-Induced Epithelial-to-Mesenchymal Transition-Like Phenotype of 603B Cells

PubMed Central

Liu, Jun; Eischeid, Alex N.; Chen, Xian-Ming

2012-01-01

Recent studies have suggested that proliferating cholangiocytes have an important role in the induction of fibrosis, either directly via epithelial-to-mesenchymal transition (EMT), or indirectly via activation of other liver cell types. Transforming growth factor beta 1 (TGF-β1), a critical fibrotic cytokine for hepatic fibrosis, is a potent EMT inducer. This study aimed to clarify the potential contributions of TGF-β1-induced EMT-like cholangiocyte phenotype to collagen production and cell survival of cholangiocytes in vitro. Mouse cholangiocytes (603B cells) were treated with TGF-β1 and EMT-like phenotype alterations were monitored by morphological changes and expression of EMT-associated genes. Alterations in Col1A1 gene, Col1A1-associated miR-29s, and pro-apoptotic genes were measured in TGF-β1-treated 603B cells. Snail1 knockdown was achieved using shRNA to evaluate the contribution of EMT-associated changes to Col1A1 production and cell survival. We found TGF-β1 treatment induced partial EMT-like phenotype transition in 603B cells in a Snail1-dependent manner. TGF-β1 also stimulated collagen α1(I) expression in 603B cells. However, this induction was not parallel to the EMT-like alterations and independent of Snail1 or miR-29 expression. Cells undergoing EMT-like changes showed a modest down-regulation of multiple pro-apoptotic genes and displayed resistance to TNF-α-induced apoptosis. TGF-β1-induced apoptosis resistance was attenuated in Snail1 knockdown 603B cells. TGF-β1-induced Col1A1 production seems to be independent of EMT-like transition and miR-29 expression. Nevertheless, TGF-β1-induced EMT may contribute to the increased survival capacity of cholangiocytes via modulating the expression of pro-apoptotic genes. PMID:23236489

Molecular and pathological signatures of epithelial–mesenchymal transitions at the cancer invasion front

PubMed Central

Pauwels, Patrick; De Craene, Bram; Sabbah, Michèle; Emami, Shahin; Redeuilh, Gérard; Gespach, Christian; Bracke, Marc

2008-01-01

Reduction of epithelial cell–cell adhesion via the transcriptional repression of cadherins in combination with the acquisition of mesenchymal properties are key determinants of epithelial–mesenchymal transition (EMT). EMT is associated with early stages of carcinogenesis, cancer invasion and recurrence. Furthermore, the tumor stroma dictates EMT through intensive bidirectional communication. The pathological analysis of EMT signatures is critically, especially to determine the presence of cancer cells at the resection margins of a tumor. When diffusion barriers disappear, EMT markers may be detected in sera from cancer patients. The detection of EMT signatures is not only important for diagnosis but can also be exploited to enhance classical chemotherapy treatments. In conclusion, further detailed understanding of the contextual cues and molecular mediators that control EMT will be required in order to develop diagnostic tools and small molecule inhibitors with potential clinical implications. PMID:18648847

Core epithelial-to-mesenchymal transition interactome gene-expression signature is associated with claudin-low and metaplastic breast cancer subtypes.

PubMed

Taube, Joseph H; Herschkowitz, Jason I; Komurov, Kakajan; Zhou, Alicia Y; Gupta, Supriya; Yang, Jing; Hartwell, Kimberly; Onder, Tamer T; Gupta, Piyush B; Evans, Kurt W; Hollier, Brett G; Ram, Prahlad T; Lander, Eric S; Rosen, Jeffrey M; Weinberg, Robert A; Mani, Sendurai A

2010-08-31

The epithelial-to-mesenchymal transition (EMT) produces cancer cells that are invasive, migratory, and exhibit stem cell characteristics, hallmarks of cells that have the potential to generate metastases. Inducers of the EMT include several transcription factors (TFs), such as Goosecoid, Snail, and Twist, as well as the secreted TGF-beta1. Each of these factors is capable, on its own, of inducing an EMT in the human mammary epithelial (HMLE) cell line. However, the interactions between these regulators are poorly understood. Overexpression of each of the above EMT inducers up-regulates a subset of other EMT-inducing TFs, with Twist, Zeb1, Zeb2, TGF-beta1, and FOXC2 being commonly induced. Up-regulation of Slug and FOXC2 by either Snail or Twist does not depend on TGF-beta1 signaling. Gene expression signatures (GESs) derived by overexpressing EMT-inducing TFs reveal that the Twist GES and Snail GES are the most similar, although the Goosecoid GES is the least similar to the others. An EMT core signature was derived from the changes in gene expression shared by up-regulation of Gsc, Snail, Twist, and TGF-beta1 and by down-regulation of E-cadherin, loss of which can also trigger an EMT in certain cell types. The EMT core signature associates closely with the claudin-low and metaplastic breast cancer subtypes and correlates negatively with pathological complete response. Additionally, the expression level of FOXC1, another EMT inducer, correlates strongly with poor survival of breast cancer patients.

Core epithelial-to-mesenchymal transition interactome gene-expression signature is associated with claudin-low and metaplastic breast cancer subtypes

PubMed Central

Taube, Joseph H.; Herschkowitz, Jason I.; Komurov, Kakajan; Zhou, Alicia Y.; Gupta, Supriya; Yang, Jing; Hartwell, Kimberly; Onder, Tamer T.; Gupta, Piyush B.; Evans, Kurt W.; Hollier, Brett G.; Ram, Prahlad T.; Lander, Eric S.; Rosen, Jeffrey M.; Weinberg, Robert A.; Mani, Sendurai A.

2010-01-01

The epithelial-to-mesenchymal transition (EMT) produces cancer cells that are invasive, migratory, and exhibit stem cell characteristics, hallmarks of cells that have the potential to generate metastases. Inducers of the EMT include several transcription factors (TFs), such as Goosecoid, Snail, and Twist, as well as the secreted TGF-β1. Each of these factors is capable, on its own, of inducing an EMT in the human mammary epithelial (HMLE) cell line. However, the interactions between these regulators are poorly understood. Overexpression of each of the above EMT inducers up-regulates a subset of other EMT-inducing TFs, with Twist, Zeb1, Zeb2, TGF-β1, and FOXC2 being commonly induced. Up-regulation of Slug and FOXC2 by either Snail or Twist does not depend on TGF-β1 signaling. Gene expression signatures (GESs) derived by overexpressing EMT-inducing TFs reveal that the Twist GES and Snail GES are the most similar, although the Goosecoid GES is the least similar to the others. An EMT core signature was derived from the changes in gene expression shared by up-regulation of Gsc, Snail, Twist, and TGF-β1 and by down-regulation of E-cadherin, loss of which can also trigger an EMT in certain cell types. The EMT core signature associates closely with the claudin-low and metaplastic breast cancer subtypes and correlates negatively with pathological complete response. Additionally, the expression level of FOXC1, another EMT inducer, correlates strongly with poor survival of breast cancer patients. PMID:20713713

Inhibition of integrin-linked kinase blocks podocyte epithelial-mesenchymal transition and ameliorates proteinuria.

PubMed

Kang, Young Sun; Li, Yingjian; Dai, Chunsun; Kiss, Lawrence P; Wu, Chuanyue; Liu, Youhua

2010-08-01

Proteinuria is a primary clinical symptom of a large number of glomerular diseases that progress to end-stage renal failure. Podocyte dysfunctions play a fundamental role in defective glomerular filtration in many common forms of proteinuric kidney disorders. Since binding of these cells to the basement membrane is mediated by integrins, we determined the role of integrin-linked kinase (ILK) in podocyte dysfunction and proteinuria. ILK expression was induced in mouse podocytes by various injurious stimuli known to cause proteinuria including TGF-beta1, adriamycin, puromycin, and high ambient glucose. Podocyte ILK was also found to be upregulated in human proteinuric glomerular diseases. Ectopic expression of ILK in podocytes decreased levels of the epithelial markers nephrin and ZO-1, induced mesenchymal markers such as desmin, fibronectin, matrix metalloproteinase-9 (MMP-9), and alpha-smooth muscle actin (alpha-SMA), promoted cell migration, and increased the paracellular albumin flux across podocyte monolayers. ILK also induced Snail, a key transcription factor mediating epithelial-mesenchymal transition (EMT). Blockade of ILK activity with a highly selective small molecule inhibitor reduced Snail induction and preserved podocyte phenotypes following TGF-beta1 or adriamycin stimulation. In vivo, this ILK inhibitor ameliorated albuminuria, repressed glomerular induction of MMP-9 and alpha-SMA, and preserved nephrin expression in murine adriamycin nephropathy. Our results show that upregulation of ILK is a convergent pathway leading to podocyte EMT, migration, and dysfunction. ILK may be an attractive target for therapeutic intervention of proteinuric kidney diseases.

Epithelial-mesenchymal transition leads to crizotinib resistance in H2228 lung cancer cells with EML4-ALK translocation.

PubMed

Kim, Hyeong Ryul; Kim, Woo Sung; Choi, Yun Jung; Choi, Chang Min; Rho, Jin Kyung; Lee, Jae Cheol

2013-12-01

Epithelial-mesenchymal transition (EMT) is associated with reduced sensitivity to many chemotherapeutic drugs, including EGFR tyrosine kinase inhibitors. Here, we investigated if this reduced sensitivity also contributes to resistance to crizotinib, an ALK inhibitor of lung cancer that exhibits the EML4-ALK translocation. We established a crizotinib-resistant subline (H2228/CR), which was derived from the parental H2228 cell line by long-term exposure to increasing concentrations of crizotinib. Characteristics associated with EMT, including morphology, EMT marker proteins, and cellular mobility, were analyzed. Compared with H2228 cells, the growth of H2228/CR cells was independent of EML4-ALK, and H2228/CR cells showed cross-resistance to TAE-684 (a second-generation ALK inhibitor). Phenotypic changes to the spindle-cell shape were noted in H2228/CR cells, which were accompanied by a decrease in E-cadherin and increase in vimentin and AXL. In addition, H2228/CR cells showed increased secretion and expression of TGF-β1. Invasion and migration capabilities were dramatically increased in H2228/CR cells. Applying TGF-β1 treatment to parental H2228 cells for 72 h induced reversible EMT, leading to crizotinib resistance, but this was reversed by the removal of TGF-β1. Suppression of vimentin in H2228/CR cells by siRNA treatment restored sensitivity to crizotinib. Furthermore, these resistant cells remained highly sensitive to the Hsp90 inhibitors, similar to the parental H2228 cells. In conclusion, we suggest EMT is possibly involved in acquired resistance to crizotinib, and that HSP90 inhibitors could be a promising option for the treatment of EMT. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Prognostic Value of EMT-inducing Transcription Factors (EMT-TFs) in Metastatic Breast Cancer: A Systematic Review and Meta-analysis

PubMed Central

Imani, Saber; Hosseinifard, Hossein; Cheng, Jingliang; Wei, Chunli; Fu, Junjiang

2016-01-01

The epithelial-to-mesenchymal transition (EMT) is a vital control point in metastatic breast cancer (MBC). TWIST1, SNAIL1, SLUG, and ZEB1, as key EMT-inducing transcription factors (EMT-TFs), are involved in MBC through different signaling cascades. This updated meta-analysis was conducted to assess the correlation between the expression of EMT-TFs and prognostic value in MBC patients. A total of 3,218 MBC patients from fourteen eligible studies were evaluated. The pooled hazard ratios (HR) for EMT-TFs suggested that high EMT-TF expression was significantly associated with poor prognosis in MBC patients (HRs = 1.72; 95% confidence intervals (CIs) = 1.53–1.93; P = 0.001). In addition, the overexpression of SLUG was the most impactful on the risk of MBC compared with TWIST1 and SNAIL1, which sponsored fixed models. Strikingly, the increased risk of MBC was less associated with ZEB1 expression. However, the EMT-TF expression levels significantly increased the risk of MBC in the Asian population (HR = 2.11, 95% CI = 1.70–2.62) without any publication bias (t = 1.70, P = 0.11). These findings suggest that the overexpression of potentially TWIST1, SNAIL1 and especially SLUG play a key role in the aggregation of MBC treatment as well as in the improvement of follow-up plans in Asian MBC patients. PMID:27335258

Cancer-initiating cells derived from established cervical cell lines exhibit stem-cell markers and increased radioresistance

PubMed Central

2012-01-01

Background Cancer-initiating cells (CICs) are proposed to be responsible for the generation of metastasis and resistance to therapy. Accumulating evidences indicates CICs are found among different human cancers and cell lines derived from them. Few studies address the characteristics of CICs in cervical cancer. We identify biological features of CICs from four of the best-know human cell lines from uterine cervix tumors. (HeLa, SiHa, Ca Ski, C-4 I). Methods Cells were cultured as spheres under stem-cell conditions. Flow cytometry was used to detect expression of CD34, CD49f and CD133 antigens and Hoechst 33342 staining to identify side population (SP). Magnetic and fluorescence-activated cell sorting was applied to enrich and purify populations used to evaluate tumorigenicity in nude mice. cDNA microarray analysis and in vitro radioresistance assay were carried out under standard conditions. Results CICs, enriched as spheroids, were capable to generate reproducible tumor phenotypes in nu-nu mice and serial propagation. Injection of 1 × 103 dissociated spheroid cells induced tumors in the majority of animals, whereas injection of 1 × 105 monolayer cells remained nontumorigenic. Sphere-derived CICs expressed CD49f surface marker. Gene profiling analysis of HeLa and SiHa spheroid cells showed up-regulation of CICs markers characteristic of the female reproductive system. Importantly, epithelial to mesenchymal (EMT) transition-associated markers were found highly expressed in spheroid cells. More importantly, gene expression analysis indicated that genes required for radioresistance were also up-regulated, including components of the double-strand break (DSB) DNA repair machinery and the metabolism of reactive oxygen species (ROS). Dose-dependent radiation assay indicated indeed that CICs-enriched populations exhibit an increased resistance to ionizing radiation (IR). Conclusions We characterized a self-renewing subpopulation of CICs found among four well known human cancer-derived cell lines (HeLa, SiHa, Ca Ski and C-4 I) and found that they express characteristic markers of stem cell, EMT and radioresistance. The fact that CICs demonstrated a higher degree of resistance to radiation than differentiated cells suggests that specific detection and targeting of CICs could be highly valuable for the therapy of tumors from the uterine cervix. PMID:22284662

Suppression of triple-negative breast cancer metastasis by pan-DAC inhibitor panobinostat via inhibition of ZEB family of EMT master regulators.

PubMed

Rhodes, Lyndsay V; Tate, Chandra R; Segar, H Chris; Burks, Hope E; Phamduy, Theresa B; Hoang, Van; Elliott, Steven; Gilliam, Diari; Pounder, F Nell; Anbalagan, Muralidharan; Chrisey, Douglas B; Rowan, Brian G; Burow, Matthew E; Collins-Burow, Bridgette M

2014-06-01

Triple-negative breast cancer (TNBC) is a highly aggressive breast cancer subtype that lacks effective targeted therapies. The epithelial-to-mesenchymal transition (EMT) is a key contributor in the metastatic process. We previously showed the pan-deacetylase inhibitor LBH589 induces CDH1 expression in TNBC cells, suggesting regulation of EMT. The purpose of this study was to examine the effects of LBH589 on the metastatic qualities of TNBC cells and the role of EMT in this process. A panel of breast cancer cell lines (MCF-7, MDA-MB-231, and BT-549), drugged with LBH589, was examined for changes in cell morphology, migration, and invasion in vitro. The effect on in vivo metastasis was examined using immunofluorescent staining of lung sections. EMT gene expression profiling was used to determine LBH589-induced changes in TNBC cells. ZEB overexpression studies were conducted to validate requirement of ZEB in LBH589-mediated proliferation and tumorigenesis. Our results indicate a reversal of EMT by LBH589 as demonstrated by altered morphology and altered gene expression in TNBC. LBH589 was shown to be a more potent inhibitor of EMT than other HDAC inhibitors, SAHA and TMP269. Additionally, we found that LBH589 inhibits metastasis of MDA-MB-231 cells in vivo. These effects of LBH589 were mediated in part by inhibition of ZEB, as overexpression of ZEB1 or ZEB2 mitigated the effects of LBH589 on MDA-MB-231 EMT-associated gene expression, migration, invasion, CDH1 expression, and tumorigenesis. These data indicate therapeutic potential of LBH589 in targeting EMT and metastasis of TNBC.

Krüppel-like factor 8 involved in hypoxia promotes the invasion and metastasis of gastric cancer via epithelial to mesenchymal transition.

PubMed

Liu, Na; Wang, Yafang; Zhou, Yongan; Pang, Hailin; Zhou, Jing; Qian, Pei; Liu, Lili; Zhang, Helong

2014-12-01

Previously, we reported that hypoxia was able to induce invasion and metastasis in gastric cancer and that hypoxia-inducible factor-1 (HIF-1) is a key factor involved in this tumor type. Krüppel-like factor 8 (KLF8) as a transcriptional repressor has been suggested as a promoter of tumor metastasis in breast cancer and an inducer of the epithelial‑mesenchymal transition (EMT). KLF8 is also highly expressed in gastric cancer tissues, contributing to poor prognosis. However, the association between KLF8 and HIF-1 in regulating the progression of human gastric cancer in hypoxia is unclear. In the present study, we found that KLF8 was overexpressed in gastric cancer metastatic tissues and cells. Additionally, KLF8 siRNA significantly inhibited SGC7901 cell invasion and migration compared with SGC7901, SGC7901/Scr-si cells. Hypoxia is thus able to induce KLF8 expression and EMT in SGC7901 cells. However, following the examination of changes in cell morphology and epithelial and mesenchymal markers, it was found that KLF8 siRNA and HIF-1 siRNA strongly reversed EMT in cells undergoing hypoxia. Furthermore, hypoxia-induced KLF8 overexpression was attenuated by HIF-1 siRNA. Experiments using luciferase promoter constructs resulted in a marked increase in the activity of cells exposed to hypoxia and decreased activity in cells co-transfected with HIF-1 siRNA. The chromatin immunoprecipitation assay revealed proximal HRE at -133 is the main HIF-1 binding site in the KLF8 promoter. In conclusion, the results demonstrated that KLF8 is actively enhanced by hypoxia and is a novel HIF-1 target. KLF8 is a novel EMT regulating transcription factor that involved in the progression of gastric cancer. The specific anti-EMT drugs in combination with anti-hypoxia are new promising cancer therapies.

Ferritin heavy chain is a negative regulator of ovarian cancer stem cell expansion and epithelial to mesenchymal transition

PubMed Central

Pisanu, Maria Elena; Faniello, Maria Concetta; Jakopin, Žiga; Chiarella, Emanuela; Giovannone, Emilia Dora; Mancini, Rita; Ciliberto, Gennaro

2016-01-01

Objectives Ferritin is the major intracellular iron storage protein essential for maintaining the cellular redox status. In recent years ferritin heavy chain (FHC) has been shown to be involved also in the control of cancer cell growth. Analysis of public microarray databases in ovarian cancer revealed a correlation between low FHC expression levels and shorter survival. To better understand the role of FHC in cancer, we have silenced the FHC gene in SKOV3 cells. Results FHC-KO significantly enhanced cell viability and induced a more aggressive behaviour. FHC-silenced cells showed increased ability to form 3D spheroids and enhanced expression of NANOG, OCT4, ALDH and Vimentin. These features were accompanied by augmented expression of SCD1, a major lipid metabolism enzyme. FHC apparently orchestrates part of these changes by regulating a network of miRNAs. Methods FHC-silenced and control shScr SKOV3 cells were monitored for changes in proliferation, migration, ability to propagate as 3D spheroids and for the expression of stem cell and epithelial-to-mesenchymal-transition (EMT) markers. The expression of three miRNAs relevant to spheroid formation or EMT was assessed by q-PCR. Conclusions In this paper we uncover a new function of FHC in the control of cancer stem cells. PMID:27566559

Arctigenin Inhibits Lung Metastasis of Colorectal Cancer by Regulating Cell Viability and Metastatic Phenotypes.

PubMed

Han, Yo-Han; Kee, Ji-Ye; Kim, Dae-Seung; Mun, Jeong-Geon; Jeong, Mi-Young; Park, Sang-Hyun; Choi, Byung-Min; Park, Sung-Joo; Kim, Hyun-Jung; Um, Jae-Young; Hong, Seung-Heon

2016-08-27

Arctigenin (ARC) has been shown to have an anti-cancer effect in various cell types and tissues. However, there have been no studies concerning metastatic colorectal cancer (CRC). In this study, we investigated the anti-metastatic properties of ARC on colorectal metastasis and present a potential candidate drug. ARC induced cell cycle arrest and apoptosis in CT26 cells through the intrinsic apoptotic pathway via MAPKs signaling. In several metastatic phenotypes, ARC controlled epithelial-mesenchymal transition (EMT) through increasing the expression of epithelial marker E-cadherin and decreasing the expressions of mesenchymal markers; N-cadherin, vimentin, β-catenin, and Snail. Moreover, ARC inhibited migration and invasion through reducing of matrix metalloproteinase-2 (MMP-2) and MMP-9 expressions. In an experimental metastasis model, ARC significantly inhibited lung metastasis of CT26 cells. Taken together, our study demonstrates the inhibitory effects of ARC on colorectal metastasis.

Opposite regulation of MDM2 and MDMX expression in acquisition of mesenchymal phenotype in benign and cancer cells.

PubMed

Slabáková, Eva; Kharaishvili, Gvantsa; Smějová, Monika; Pernicová, Zuzana; Suchánková, Tereza; Remšík, Ján; Lerch, Stanislav; Straková, Nicol; Bouchal, Jan; Král, Milan; Culig, Zoran; Kozubík, Alois; Souček, Karel

2015-11-03

Plasticity of cancer cells, manifested by transitions between epithelial and mesenchymal phenotypes, represents a challenging issue in the treatment of neoplasias. Both epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET) are implicated in the processes of metastasis formation and acquisition of stem cell-like properties. Mouse double minute (MDM) 2 and MDMX are important players in cancer progression, as they act as regulators of p53, but their function in EMT and metastasis may be contradictory. Here, we show that the EMT phenotype in multiple cellular models and in clinical prostate and breast cancer samples is associated with a decrease in MDM2 and increase in MDMX expression. Modulation of EMT-accompanying changes in MDM2 expression in benign and transformed prostate epithelial cells influences their migration capacity and sensitivity to docetaxel. Analysis of putative mechanisms of MDM2 expression control demonstrates that in the context of defective p53 function, MDM2 expression is regulated by EMT-inducing transcription factors Slug and Twist. These results provide an alternative context-specific role of MDM2 in EMT, cell migration, metastasis, and therapy resistance.

Role of glycogen synthase kinase-3β and PPAR-γ on epithelial-to-mesenchymal transition in DSS-induced colorectal fibrosis.

PubMed

Di Gregorio, Jacopo; Sferra, Roberta; Speca, Silvia; Vetuschi, Antonella; Dubuquoy, Caroline; Desreumaux, Pierre; Pompili, Simona; Cristiano, Loredana; Gaudio, Eugenio; Flati, Vincenzo; Latella, Giovanni

2017-01-01

Intestinal fibrosis is characterized by abnormal production and deposition of extracellular matrix (ECM) proteins by activated myofibroblasts. The main progenitor cells of activated myofibroblasts are the fibroblasts and the epithelial cells, the latter through the epithelial-mesenchymal transition (EMT). To evaluate the action of the new PPAR-γ modulator, GED-0507-34 Levo (GED) on the expression of EMT associated and regulatory proteins such as TGF-β, Smad3, E-cadherin, Snail, ZEB1, β-catenin, and GSK-3β, in a mouse model of DSS-induced intestinal fibrosis. Chronic colitis and fibrosis were induced by oral administration of 2.5% DSS (w/v) for 6 weeks. GW9662 (GW), a selective PPAR-γ inhibitor, was also administered by intraperitoneal injection at the dose of 1 mg/kg/day combined with GED treatment. All drugs were administered at the beginning of the second cycle of DSS (day 12). 65 mice were randomly divided into five groups (H2O as controls n = 10, H2O+GED n = 10, DSS n = 15, DSS+GED n = 15, DSS+GED+GW n = 15). The colon was excised for macroscopic examination and histological and morphometric analyses. The level of expression of molecules involved in EMT and fibrosis, like TGF-β, Smad3, E-cadherin, Snail, ZEB1, β-catenin, GSK-3β and PPAR-γ, was assessed by immunohistochemistry, immunofluorescence, western blot and Real Time PCR. GED improved the DSS-induced chronic colitis and fibrosis. GED was able to reduce the expression of the main fibrosis markers (α-SMA, collagen I-III and fibronectin) as well as the pivotal pro-fibrotic molecules IL-13, TGF-β and Smad3, while it increased the anti-fibrotic PPAR-γ. All these GED effects were nullified by co-administration of GW with GED. Furthermore, GED was able to normalize the expression levels of E-cadherin and β-catenin and upregulated GSK-3β, that are all known to be involved both in EMT and fibrosis. The DSS-induced intestinal fibrosis was improved by the new PPAR-γ modulator GED-0507-34 Levo through the modulation of EMT mediators and pro-fibrotic molecules and through GSK-3β induction.

Asbestos exposure induces alveolar epithelial cell plasticity through MAPK/Erk signaling.

PubMed

Tamminen, Jenni A; Myllärniemi, Marjukka; Hyytiäinen, Marko; Keski-Oja, Jorma; Koli, Katri

2012-07-01

The inhalation of asbestos fibers is considered to be highly harmful, and lead to fibrotic and/or malignant disease. Epithelial-to-mesenchymal transition (EMT) is a common pathogenic mechanism in asbestos associated fibrotic (asbestosis) and malignant lung diseases. The characterization of molecular pathways contributing to EMT may provide new possibilities for prognostic and therapeutic applications. The role of asbestos as an inducer of EMT has not been previously characterized. We exposed cultured human lung epithelial cells to crocidolite asbestos and analyzed alterations in the expression of epithelial and mesenchymal marker proteins and cell morphology. Asbestos was found to induce downregulation of E-cadherin protein levels in A549 lung carcinoma cells in 2-dimensional (2D) and 3D cultures. Similar findings were made in primary small airway epithelial cells cultured in 3D conditions where the cells retained alveolar type II cell phenotype. A549 cells also exhibited loss of cell-cell contacts, actin reorganization and expression of α-smooth muscle actin (α-SMA) in 2D cultures. These phenotypic changes were not associated with increased transforming growth factor (TGF)-β signaling activity. MAPK/Erk signaling pathway was found to mediate asbestos-induced downregulation of E-cadherin and alterations in cell morphology. Our results suggest that asbestos can induce epithelial plasticity, which can be interfered by blocking the MAPK/Erk kinase activity. Copyright © 2012 Wiley Periodicals, Inc.

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

PubMed Central

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

Brachyury as a potential modulator of androgen receptor activity and a key player in therapy resistance in prostate cancer

PubMed Central

Pinto, Filipe; Pértega-Gomes, Nelma; Vizcaíno, José R.; Andrade, Raquel P.; Cárcano, Flavio M.; Reis, Rui Manuel

2016-01-01

Prostate cancer (PCa) is the most commonly diagnosed neoplasm and the second leading cause of cancer-related deaths in men. Acquisition of resistance to conventional therapy is a major problem for PCa patient management. Several mechanisms have been described to promote therapy resistance in PCa, such as androgen receptor (AR) activation, epithelial-to-mesenchymal transition (EMT), acquisition of stem cell properties and neuroendocrine transdifferentiation (NEtD). Recently, we identified Brachyury as a new biomarker of PCa aggressiveness and poor prognosis. In the present study we aimed to assess the role of Brachyury in PCa therapy resistance. We showed that Brachyury overexpression in prostate cancer cells lines increased resistance to docetaxel and cabazitaxel drugs, whereas Brachyury abrogation induced decrease in therapy resistance. Through ChiP-qPCR assays we further demonstrated that Brachyury is a direct regulator of AR expression as well as of the biomarker AMACR and the mesenchymal markers Snail and Fibronectin. Furthermore, in vitro Brachyury was also able to increase EMT and stem properties. By in silico analysis, clinically human Brachyury-positive PCa samples were associated with biomarkers of PCa aggressiveness and therapy resistance, including PTEN loss, and expression of NEtD markers, ERG and Bcl-2. Taken together, our results indicate that Brachyury contributes to tumor chemotherapy resistance, constituting an attractive target for advanced PCa patients. PMID:27049720

A link between premenopausal iron deficiency and breast cancer malignancy

PubMed Central

2013-01-01

Background Young breast cancer (BC) patients less than 45 years old are at higher risk of dying from the disease when compared to their older counterparts. However, specific risk factors leading to this poorer outcome have not been identified. Methods One candidate is iron deficiency, as this is common in young women and a clinical feature of young age. In the present study, we used immuno-competent and immuno-deficient mouse xenograft models as well as hemoglobin as a marker of iron status in young BC patients to demonstrate whether host iron deficiency plays a pro-metastatic role. Results We showed that mice fed an iron-deficient diet had significantly higher tumor volumes and lung metastasis compared to those fed normal iron diets. Iron deficiency mainly altered Notch but not TGF-β and Wnt signaling in the primary tumor, leading to the activation of epithelial mesenchymal transition (EMT). This was revealed by increased expression of Snai1 and decreased expression of E-cadherin. Importantly, correcting iron deficiency by iron therapy reduced primary tumor volume, lung metastasis, and reversed EMT markers in mice. Furthermore, we found that mild iron deficiency was significantly associated with lymph node invasion in young BC patients (p<0.002). Conclusions Together, our finding indicates that host iron deficiency could be a contributor of poor prognosis in young BC patients. PMID:23800380

Peroxiredoxin 5 promotes the epithelial-mesenchymal transition in colon cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ahn, Hye-Mi; Yoo, Jin-Woo; College of Natural Sciences, Kyungpook National University, Daegu

Globally, colorectal cancer (CRC) is common cause of cancer-related deaths. The high mortality rate of patients with colon cancer is due to cancer cell invasion and metastasis. Initiation of the epithelial-to-mesenchymal transition (EMT) is essential for the tumorigenesis. Peroxiredoinxs (PRX1-6) have been reported to be overexpressed in various tumor tissues, and involved to be responsible for tumor progression. However, the exact role of PRX5 in colon cancer remains to be investigated enhancing proliferation and promoting EMT properties. In this study, we constructed stably overexpressing PRX5 and suppressed PRX5 expression in CRC cells. Our results revealed that PRX5 overexpression significantly enhancedmore » CRC cell proliferation, migration, and invasion. On the other hand, PRX5 suppression markedly inhibited these EMT properties. PRX5 was also demonstrated to regulate the expression of two hallmark EMT proteins, E-cadherin and Vimentin, and the EMT-inducing transcription factors, Snail and Slug. Moreover, in the xenograft mouse model, showed that PRX5 overexpression enhances tumor growth of CRC cells. Thus, our findings first provide evidence in CRC that PRX5 promotes EMT properties by inducing the expression of EMT-inducing transcription factors. Therefore, PRX5 can be used as a predictive biomarker and serves as a putative therapeutic target for the development of clinical treatments for human CRC. - Highlights: • PRX5 promoted colorectal cancer cell proliferation. • PRX5 enhanced EMT properties in colorectal cancer. • PRX5 mediated the EMT by inducing the expression of Snail and Slug. • PRX5 promoted tumor growth of colorectal cancer cells.« less

ZEB1 induces LOXL2-mediated collagen stabilization and deposition in the extracellular matrix to drive lung cancer invasion and metastasis.

PubMed

Peng, D H; Ungewiss, C; Tong, P; Byers, L A; Wang, J; Canales, J R; Villalobos, P A; Uraoka, N; Mino, B; Behrens, C; Wistuba, I I; Han, R I; Wanna, C A; Fahrenholtz, M; Grande-Allen, K J; Creighton, C J; Gibbons, D L

2017-04-06

Lung cancer is the leading cause of cancer-related deaths, primarily due to distant metastatic disease. Metastatic lung cancer cells can undergo an epithelial-to-mesenchymal transition (EMT) regulated by various transcription factors, including a double-negative feedback loop between the microRNA-200 (miR-200) family and ZEB1, but the precise mechanisms by which ZEB1-dependent EMT promotes malignancy remain largely undefined. Although the cell-intrinsic effects of EMT are important for tumor progression, the reciprocal dynamic crosstalk between mesenchymal cancer cells and the extracellular matrix (ECM) is equally critical in regulating invasion and metastasis. Investigating the collaborative effect of EMT and ECM in the metastatic process reveals increased collagen deposition in metastatic tumor tissues as a direct consequence of amplified collagen gene expression in ZEB1-activated mesenchymal lung cancer cells. In addition, collagen fibers in metastatic lung tumors exhibit greater linearity and organization as a result of collagen crosslinking by the lysyl oxidase (LOX) family of enzymes. Expression of the LOX and LOXL2 isoforms is directly regulated by miR-200 and ZEB1, respectively, and their upregulation in metastatic tumors and mesenchymal cell lines is coordinated to that of collagen. Functionally, LOXL2, as opposed to LOX, is the principal isoform that crosslinks and stabilizes insoluble collagen deposition in tumor tissues. In turn, focal adhesion formation and FAK/SRC signaling is activated in mesenchymal tumor cells by crosslinked collagen in the ECM. Our study is the first to validate direct regulation of LOX and LOXL2 by the miR-200/ZEB1 axis, defines a novel mechanism driving tumor metastasis, delineates collagen as a prognostic marker, and identifies LOXL2 as a potential therapeutic target against tumor progression.

ZEB1 Induces LOXL2-Mediated Collagen Stabilization and Deposition in the Extracellular Matrix to Drive Lung Cancer Invasion and Metastasis

PubMed Central

Peng, David H.; Ungewiss, Christin; Tong, Pan; Byers, Lauren A.; Wang, Jing; Canales, Jaime Rodriguez; Villalobos, Pamela A.; Uraoka, Naohiro; Mino, Barbara; Behrens, Carmen; Wistuba, Ignacio I.; Han, Richard I; Wanna, Charles A.; Fahrenholtz, Monica; Grande-Allen, Kathryn Jane; Creighton, Chad J.; Gibbons, Don L.

2016-01-01

Lung cancer is the leading cause of cancer-related death, primarily due to distant metastatic disease. Metastatic lung cancer cells can undergo an epithelial-to-mesenchymal transition (EMT) regulated by many transcription factors, including double-negative feedback loop between the microRNA-200 (miR-200) family and ZEB1, but the precise mechanisms by which ZEB1-dependent EMT promotes malignancy remain largely undefined. While the cell-intrinsic effects of EMT are important for tumor progression, the reciprocal dynamic crosstalk between mesenchymal cancer cells and the extracellular matrix (ECM) is equally critical in regulating invasion and metastasis. Investigating the collaborative effect of EMT and ECM in the metastatic process reveals increased collagen deposition in metastatic tumor tissues as a direct consequence of amplified collagen gene expression in ZEB1-activated mesenchymal lung cancer cells. Additionally, collagen fibers in metastatic lung tumors exhibit greater linearity and organization as a result of collagen crosslinking by the lysyl oxidase (LOX) family of enzymes. Expression of the LOX and LOXL2 isoforms is directly regulated by miR-200 and ZEB1, respectively, and their upregulation in metastatic tumors and mesenchymal cell lines is coordinated to that of collagen. Functionally, LOXL2, as opposed to LOX, is the principle isoform that crosslinks and stabilizes insoluble collagen deposition in tumor tissues. In turn, focal adhesion formation and FAK/SRC signaling is activated in mesenchymal tumor cells by crosslinked collagen in the ECM. Our study is the first to validate direct regulation of LOX and LOXL2 by the miR-200/ZEB1 axis, defines a novel mechanism driving tumor metastasis, delineates collagen as a prognostic marker, and identifies LOXL2 as a potential therapeutic target against tumor progression. PMID:27694892

HER2 in Breast Cancer Stemness: A Negative Feedback Loop towards Trastuzumab Resistance

PubMed Central

Nami, Babak; Wang, Zhixiang

2017-01-01

HER2 receptor tyrosine kinase that is overexpressed in approximately 20% of all breast cancers (BCs) is a poor prognosis factor and a precious target for BC therapy. Trastuzumab is approved by FDA to specifically target HER2 for treating HER2+ BC. However, about 60% of patients with HER2+ breast tumor develop de novo resistance to trastuzumab, partially due to the loss of expression of HER2 extracellular domain on their tumor cells. This is due to shedding/cleavage of HER2 by metalloproteinases (ADAMs and MMPs). HER2 shedding results in the accumulation of intracellular carboxyl-terminal HER2 (p95HER2), which is a common phenomenon in trastuzumab-resistant tumors and is suggested as a predictive marker for trastuzumab resistance. Up-regulation of the metalloproteinases is a poor prognosis factor and is commonly seen in mesenchymal-like cancer stem cells that are risen during epithelial to mesenchymal transition (EMT) of tumor cells. HER2 cleavage during EMT can explain why secondary metastatic tumors with high percentage of mesenchymal-like cancer stem cells are mostly resistant to trastuzumab but still sensitive to lapatinib. Importantly, many studies report HER2 interaction with oncogenic/stemness signaling pathways including TGF-β/Smad, Wnt/β-catenin, Notch, JAK/STAT and Hedgehog. HER2 overexpression promotes EMT and the emergence of cancer stem cell properties in BC. Increased expression and activation of metalloproteinases during EMT leads to proteolytic cleavage and shedding of HER2 receptor, which downregulates HER2 extracellular domain and eventually increases trastuzumab resistance. Here, we review the hypothesis that a negative feedback loop between HER2 and stemness signaling drives resistance of BC to trastuzumab. PMID:28445439

SPHK1 (sphingosine kinase 1) induces epithelial-mesenchymal transition by promoting the autophagy-linked lysosomal degradation of CDH1/E-cadherin in hepatoma cells.

PubMed

Liu, Hong; Ma, Yan; He, Hong-Wei; Zhao, Wu-Li; Shao, Rong-Guang

2017-05-04

SPHK1 (sphingosine kinase 1), a regulator of sphingolipid metabolites, plays a causal role in the development of hepatocellular carcinoma (HCC) through augmenting HCC invasion and metastasis. However, the mechanism by which SPHK1 signaling promotes invasion and metastasis in HCC remains to be clarified. Here, we reported that SPHK1 induced the epithelial-mesenchymal transition (EMT) by accelerating CDH1/E-cadherin lysosomal degradation and facilitating the invasion and metastasis of HepG2 cells. Initially, we found that SPHK1 promoted cell migration and invasion and induced the EMT process through decreasing the expression of CDH1, which is an epithelial marker. Furthermore, SPHK1 accelerated the lysosomal degradation of CDH1 to induce EMT, which depended on TRAF2 (TNF receptor associated factor 2)-mediated macroautophagy/autophagy activation. In addition, the inhibition of autophagy recovered CDH1 expression and reduced cell migration and invasion through delaying the degradation of CDH1 in SPHK1-overexpressing cells. Moreover, the overexpression of SPHK1 produced intracellular sphingosine-1-phosphate (S1P). In response to S1P stimulation, TRAF2 bound to BECN1/Beclin 1 and catalyzed the lysine 63-linked ubiquitination of BECN1 for triggering autophagy. The deletion of the RING domain of TRAF2 inhibited autophagy and the interaction of BECN1 and TRAF2. Our findings define a novel mechanism responsible for the regulation of the EMT via SPHK1-TRAF2-BECN1-CDH1 signal cascades in HCC cells. Our work indicates that the blockage of SPHK1 activity to attenuate autophagy may be a promising strategy for the prevention and treatment of HCC.

SIRT1 overexpression protects non-small cell lung cancer cells against osteopontin-induced epithelial-mesenchymal transition by suppressing NF-κB signaling.

PubMed

Li, Xuejiao; Jiang, Zhongxiu; Li, Xiangmin; Zhang, Xiaoye

2018-01-01

Osteopontin (OPN) is a promoter for tumor progression. It has been reported to promote non-small cell lung cancer (NSCLC) progression via the activation of nuclear factor-κB (NF-κB) signaling. As the increased acetylation of NF-κB p65 is linked to NF-κB activation, the regulation of NF-κB p65 acetylation could be a potential treatment target for OPN-induced NSCLC progression. Sirtuin 1 (SIRT1) is a deacetylase, and the role of SIRT1 in tumor progression is still controversial. The effect and mechanism of SIRT1 on OPN-induced tumor progression remains unknown. The results presented in this research demonstrated that OPN inhibited SIRT1 expression and promoted NF-κB p65 acetylation in NSCLC cell lines (A549 and NCI-H358). In this article, overexpression of SIRT1 was induced by infection of SIRT1-overexpressing lentiviral vectors. The overexpression of SIRT1 protected NSCLC cells against OPN-induced NF-κB p65 acetylation and epithelial-mesenchymal transition (EMT), as indicated by the reduction of OPN-induced changes in the expression levels of EMT-related markers and cellular morphology. Furthermore, SIRT1 overexpression significantly attenuated OPN-induced cell proliferation, migration and invasion. Moreover, overexpression of SIRT1 inhibited OPN-induced NF-κB activation. As OPN induced NSCLC cell EMT through activation of NF-κB signaling, OPN-induced SIRT1 downregulation may play an important role in NSCLC cell EMT via NF-κB signaling. The results suggest that SIRT1 could be a tumor suppressor to attenuate OPN-induced NSCLC progression through the regulation of NF-κB signaling.

Investigating the Link between Molecular Subtypes of Glioblastoma, Epithelial-Mesenchymal Transition, and CD133 Cell Surface Protein

PubMed Central

Zarkoob, Hadi; Taube, Joseph H.; Singh, Sheila K.; Mani, Sendurai A.; Kohandel, Mohammad

2013-01-01

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. PMID:23734191

Low-GDP peritoneal dialysis fluid ('balance') has less impact in vitro and ex vivo on epithelial-to-mesenchymal transition (EMT) of mesothelial cells than a standard fluid.

PubMed

Bajo, María Auxiliadora; Pérez-Lozano, María Luisa; Albar-Vizcaino, Patricia; del Peso, Gloria; Castro, María-José; Gonzalez-Mateo, Guadalupe; Fernández-Perpén, Antonio; Aguilera, Abelardo; Sánchez-Villanueva, Rafael; Sánchez-Tomero, J Antonio; López-Cabrera, Manuel; Peter, Mirjam E; Passlick-Deetjen, Jutta; Selgas, Rafael

2011-01-01

Peritoneal membrane deterioration during peritoneal dialysis (PD) is associated with epithelial-to-mesenchymal transition (EMT) of mesothelial cells (MC), which is believed to be mainly due to glucose degradation products (GDPs) present in PD solutions. Here we investigate the impact of GDPs in PD solutions on the EMT of MC in vitro and ex vivo. For in vitro studies, omentum-derived MC were incubated with standard PD fluid or low-GDP solution diluted 1:1 with culture medium. For ex vivo studies, 33 patients, who were distributed at random to either the 'standard' or the 'low GDP' groups, were followed over 24 months. Effluents were collected every 6 months to determine EMT markers in effluent MC. Exposure of MC to standard fluid in vitro resulted in morphological change into a non-epitheloid shape, down-regulation of E-cadherin, indicative of EMT, and in a strong induction of vascular endothelial growth factor (VEGF) expression. In contrast, in vitro exposure of MC to low-GDP solution did not lead to these phenotype changes. This could be confirmed ex vivo, as the prevalence of non-epitheloid phenotype of MC in the standard group was significantly higher with increasing PD duration and MC isolated from this group showed significantly higher levels of EMT-associated molecules including fibronectin, collagen I, VEGF, IL-8 and TGF-β levels when compared with the low-GDP group. Over time, the expression of E-cadherin also decreased in the standard but increased in the low-GDP group. In addition, the levels of EMT-associated molecules (fibronectin, VEGF and IL-8) increased in the standard but decreased in the low-GDP group. A similar trend was also observed for collagen I and for TGF-β (for the first year), but did not reach global statistical significance. Accordingly, effluent MC with non-epitheloid morphology showed significantly lower levels of E-cadherin and greater levels of fibronectin, collagen I, VEGF and IL 8 when compared with MC with epitheloid phenotype. The incidence of peritonitis did not significantly influence these results. Drop-out due to technique failure was less in the 'balance' group. The functional, renal and peritoneal evaluation of patients being treated with either standard or 'balance' fluid did not show any significant difference over time. MC from PD effluent of patients treated with a PD fluid containing low GDP levels show fewer signs of EMT and the respective molecules than MC from patients treated with standard fluid, indicating a better preservation of the peritoneal membrane structure and a favourable outcome in patients using low-GDP fluid. It also confirms the hypothesis that the protection of EMT by GDP-reduced fluids is also present in vivo.

High expression of fructose-bisphosphate aldolase A induces progression of renal cell carcinoma.

PubMed

Huang, Zhengkai; Hua, Yibo; Tian, Ye; Qin, Chao; Qian, Jian; Bao, Meiling; Liu, Yiyang; Wang, Shangqian; Cao, Qiang; Ju, Xiaobing; Wang, Zengjun; Gu, Min

2018-06-01

Aldolase A (fructose-bisphosphate aldolase A, ALDOA) is a glycolytic enzyme that catalyzes reversible conversion of fructose‑1,6-bisphosphate to glyceraldehyde 3-phosphate and dihydroxyacetone phosphate. ALDOA has been revealed to be related with many carcinomas, but its expression and function in renal cell carcinoma (RCC) remain unknown. This study aimed to detect expression of ALDOA in human RCC tissue samples and to explore its function in RCC cell lines. Reverse transcription-polymerase chain reaction was used to quantify ALDOA in human RCC samples. A total of 139 RCC tissue samples obtained after surgery were analyzed in tissue microarray for ALDOA immunohistochemistry-based protein expression. Assays for cell cycle, viability, migration, and invasion were performed to assess phenotypic changes in RCC cells after ALDOA knockdown by small interfering RNA-mediated gene silencing approach and ALDOA upregulation by overexpression plasmids. Western blot analysis was used to identify alterations in markers for epithelial-mesenchymal transition (EMT), which affects metastasis and the Wnt/β‑catenin signaling pathway that influences RCC cell growth. ALDOA was upregulated in RCC samples and RCC cell lines (P<0.01). Expression of ALDOA was significantly associated with metastasis (P=0.020) and survival (P=0.0341). Downregulation of ALDOA suppressed proliferation (P<0.05) by triggering G0/G1 cell cycle arrest (P<0.05) and also inhibited migration (P<0.05) and invasion (P<0.01). Upregulation of ALDOA promoted proliferation (P<0.05) and enhanced migration (P<0.001) and invasion (P<0.001). Low expression of ALDOA could reverse EMT and inactivate the Wnt/β‑catenin signaling pathway. Our data revealed that ALDOA functions as a tumor promoter, plays a prominent role in proliferation, migration, and invasion of RCC cells with high expression, and may promote EMT and activate the Wnt/β‑catenin signaling pathway.

An Adaptogen: Withaferin A Ameliorates in Vitro and in Vivo Pulmonary Fibrosis by Modulating the Interplay of Fibrotic, Matricelluar Proteins, and Cytokines.

PubMed

Bale, Swarna; Venkatesh, Pooladanda; Sunkoju, Manoj; Godugu, Chandraiah

2018-01-01

Pulmonary fibrosis (PF) is chronic lung disease with only two FDA approved clinically available drugs, with limited safety profile. Inadequate therapy motivated us to explore the effect of vimentin inhibitor Withaferin A, as an anti-fibrotic agent against TGF-β1-induced in vitro fibrotic events and Bleomycin induced in vivo fibrosis with an emphasis on epithelial to mesenchymal transition (EMT), extracellular matrix deposition (ECM), inflammation, and angiogenesis. In vitro EMT and fibrotic events were induced by TGF-β1 in alveolar epithelial cells and human fetal lung fibroblasts followed by treatment with Withaferin A (0.25, 0.5, and 1 μM concentrations) to explore its anti-fibrotic effects. In vivo potential of Withaferin A (2 and 4 mg/kg) was assessed in murine model of Bleomycin induced PF. All the parameters and molecular studies related to PF were performed at the end of treatment period. Withaferin A treatment reduced the progression of PF by modulating the EMT related cell markers both in vivo and in vitro. Withaferin A ameliorated the expression of inflammatory cytokines including NF-κB p65, IL-1β and TNF-α, as well as attenuated the expression of pro-fibrotic proteins including CTGF, collagen 1A2, collagen 3A1, and fibronectin. Expression of angiogenic factors like VEGF, FAK, p38 MAPK, and PLC-γ1 were also inhibited by Withaferin A. Phosphorylation of Smad 2/3 induced by TGF-β1 and Bleomycin were significantly inhibited. Withaferin A suppressed expression of pro-inflammatory, pro-fibrotic, and pro-angiogenic mediators and also reduced the ECM deposition. In a nutshell, Withaferin A could probably prove as an efficient and potential therapeutic against PF.

An Adaptogen: Withaferin A Ameliorates in Vitro and in Vivo Pulmonary Fibrosis by Modulating the Interplay of Fibrotic, Matricelluar Proteins, and Cytokines

PubMed Central

Bale, Swarna; Venkatesh, Pooladanda; Sunkoju, Manoj; Godugu, Chandraiah

2018-01-01

Pulmonary fibrosis (PF) is chronic lung disease with only two FDA approved clinically available drugs, with limited safety profile. Inadequate therapy motivated us to explore the effect of vimentin inhibitor Withaferin A, as an anti-fibrotic agent against TGF-β1-induced in vitro fibrotic events and Bleomycin induced in vivo fibrosis with an emphasis on epithelial to mesenchymal transition (EMT), extracellular matrix deposition (ECM), inflammation, and angiogenesis. In vitro EMT and fibrotic events were induced by TGF-β1 in alveolar epithelial cells and human fetal lung fibroblasts followed by treatment with Withaferin A (0.25, 0.5, and 1 μM concentrations) to explore its anti-fibrotic effects. In vivo potential of Withaferin A (2 and 4 mg/kg) was assessed in murine model of Bleomycin induced PF. All the parameters and molecular studies related to PF were performed at the end of treatment period. Withaferin A treatment reduced the progression of PF by modulating the EMT related cell markers both in vivo and in vitro. Withaferin A ameliorated the expression of inflammatory cytokines including NF-κB p65, IL-1β and TNF-α, as well as attenuated the expression of pro-fibrotic proteins including CTGF, collagen 1A2, collagen 3A1, and fibronectin. Expression of angiogenic factors like VEGF, FAK, p38 MAPK, and PLC-γ1 were also inhibited by Withaferin A. Phosphorylation of Smad 2/3 induced by TGF-β1 and Bleomycin were significantly inhibited. Withaferin A suppressed expression of pro-inflammatory, pro-fibrotic, and pro-angiogenic mediators and also reduced the ECM deposition. In a nutshell, Withaferin A could probably prove as an efficient and potential therapeutic against PF. PMID:29623041

Rhinovirus Delays Cell Repolarization in a Model of Injured/Regenerating Human Airway Epithelium

PubMed Central

Faris, Andrea N.; Ganesan, Shyamala; Chattoraj, Asamanja; Chattoraj, Sangbrita S.; Comstock, Adam T.; Unger, Benjamin L.; Hershenson, Marc B.

2016-01-01

Rhinovirus (RV), which causes exacerbation in patients with chronic airway diseases, readily infects injured airway epithelium and has been reported to delay wound closure. In this study, we examined the effects of RV on cell repolarization and differentiation in a model of injured/regenerating airway epithelium (polarized, undifferentiated cells). RV causes only a transient barrier disruption in a model of normal (mucociliary-differentiated) airway epithelium. However, in the injury/regeneration model, RV prolongs barrier dysfunction and alters the differentiation of cells. The prolonged barrier dysfunction caused by RV was not a result of excessive cell death but was instead associated with epithelial-to-mesenchymal transition (EMT)-like features, such as reduced expression of the apicolateral junction and polarity complex proteins, E-cadherin, occludin, ZO-1, claudins 1 and 4, and Crumbs3 and increased expression of vimentin, a mesenchymal cell marker. The expression of Snail, a transcriptional repressor of tight and adherence junctions, was also up-regulated in RV-infected injured/regenerating airway epithelium, and inhibition of Snail reversed RV-induced EMT-like features. In addition, compared with sham-infected cells, the RV-infected injured/regenerating airway epithelium showed more goblet cells and fewer ciliated cells. Inhibition of epithelial growth factor receptor promoted repolarization of cells by inhibiting Snail and enhancing expression of E-cadherin, occludin, and Crumbs3 proteins, reduced the number of goblet cells, and increased the number of ciliated cells. Together, these results suggest that RV not only disrupts barrier function, but also interferes with normal renewal of injured/regenerating airway epithelium by inducing EMT-like features and subsequent goblet cell hyperplasia. PMID:27119973

Robo signaling regulates the production of cranial neural crest cells.

PubMed

Li, Yan; Zhang, Xiao-Tan; Wang, Xiao-Yu; Wang, Guang; Chuai, Manli; Münsterberg, Andrea; Yang, Xuesong

2017-12-01

Slit/Robo signaling plays an important role in the guidance of developing neurons in developing embryos. However, it remains obscure whether and how Slit/Robo signaling is involved in the production of cranial neural crest cells. In this study, we examined Robo1 deficient mice to reveal developmental defects of mouse cranial frontal and parietal bones, which are derivatives of cranial neural crest cells. Therefore, we determined the production of HNK1 + cranial neural crest cells in early chick embryo development after knock-down (KD) of Robo1 expression. Detection of markers for pre-migratory and migratory neural crest cells, PAX7 and AP-2α, showed that production of both was affected by Robo1 KD. In addition, we found that the transcription factor slug is responsible for the aberrant delamination/EMT of cranial neural crest cells induced by Robo1 KD, which also led to elevated expression of E- and N-Cadherin. N-Cadherin expression was enhanced when blocking FGF signaling with dominant-negative FGFR1 in half of the neural tube. Taken together, we show that Slit/Robo signaling influences the delamination/EMT of cranial neural crest cells, which is required for cranial bone development. Copyright © 2017. Published by Elsevier Inc.

Cellular Migration and Invasion Uncoupled: Increased Migration Is Not an Inexorable Consequence of Epithelial-to-Mesenchymal Transition

PubMed Central

Schaeffer, Daneen; Somarelli, Jason A.; Hanna, Gabi; Palmer, Gregory M.

2014-01-01

Metastatic dissemination requires carcinoma cells to detach from the primary tumor and invade through the basement membrane. To acquire these characteristics, epithelial tumor cells undergo epithelial-to-mesenchymal transitions (EMT), whereby cells lose polarity and E-cadherin-mediated cell-cell adhesion. Post-EMT cells have also been shown, or assumed, to be more migratory; however, there have been contradictory reports on an immortalized human mammary epithelial cell line (HMLE) that underwent EMT. In the context of carcinoma-associated EMT, it is not yet clear whether the change in migration and invasion must be positively correlated during EMT or whether enhanced migration is a necessary consequence of having undergone EMT. Here, we report that pre-EMT rat prostate cancer (PC) and HMLE cells are more migratory than their post-EMT counterparts. To determine a mechanism for increased epithelial cell migration, gene expression analysis was performed and revealed an increase in epidermal growth factor receptor (EGFR) expression in pre-EMT cells. Indeed, inhibition of EGFR in PC epithelial cells slowed migration. Importantly, while post-EMT PC and HMLE cell lines are less migratory, both remain invasive in vitro and, for PC cells, in vivo. Our study demonstrates that enhanced migration is not a phenotypic requirement of EMT, and migration and invasion can be uncoupled during carcinoma-associated EMT. PMID:25002532

Foxm1 transcription factor is required for lung fibrosis and epithelial-to-mesenchymal transition

PubMed Central

Balli, David; Ustiyan, Vladimir; Zhang, Yufang; Wang, I-Ching; Masino, Alex J; Ren, Xiaomeng; Whitsett, Jeffrey A; Kalinichenko, Vladimir V; Kalin, Tanya V

2013-01-01

Alveolar epithelial cells (AECs) participate in the pathogenesis of pulmonary fibrosis, producing pro-inflammatory mediators and undergoing epithelial-to-mesenchymal transition (EMT). Herein, we demonstrated the critical role of Forkhead Box M1 (Foxm1) transcription factor in radiation-induced pulmonary fibrosis. Foxm1 was induced in AECs following lung irradiation. Transgenic expression of an activated Foxm1 transcript in AECs enhanced radiation-induced pneumonitis and pulmonary fibrosis, and increased the expression of IL-1β, Ccl2, Cxcl5, Snail1, Zeb1, Zeb2 and Foxf1. Conditional deletion of Foxm1 from respiratory epithelial cells decreased radiation-induced pulmonary fibrosis and prevented the increase in EMT-associated gene expression. siRNA-mediated inhibition of Foxm1 prevented TGF-β-induced EMT in vitro. Foxm1 bound to and increased promoter activity of the Snail1 gene, a critical transcriptional regulator of EMT. Expression of Snail1 restored TGF-β-induced loss of E-cadherin in Foxm1-deficient cells in vitro. Lineage-tracing studies demonstrated that Foxm1 increased EMT during radiation-induced pulmonary fibrosis in vivo. Foxm1 is required for radiation-induced pulmonary fibrosis by enhancing the expression of genes critical for lung inflammation and EMT. PMID:23288041

Activation-induced cytidine deaminase (AID) is necessary for the epithelial–mesenchymal transition in mammary epithelial cells

PubMed Central

Muñoz, Denise P.; Lee, Elbert L.; Takayama, Sachiko; Coppé, Jean-Philippe; Heo, Seok-Jin; Boffelli, Dario; Di Noia, Javier M.; Martin, David I. K.

2013-01-01

Activation-induced cytidine deaminase (AID), which functions in antibody diversification, is also expressed in a variety of germ and somatic cells. Evidence that AID promotes DNA demethylation in epigenetic reprogramming phenomena, and that it is induced by inflammatory signals, led us to investigate its role in the epithelial–mesenchymal transition (EMT), a critical process in normal morphogenesis and tumor metastasis. We find that expression of AID is induced by inflammatory signals that induce the EMT in nontransformed mammary epithelial cells and in ZR75.1 breast cancer cells. shRNA–mediated knockdown of AID blocks induction of the EMT and prevents cells from acquiring invasive properties. Knockdown of AID suppresses expression of several key EMT transcriptional regulators and is associated with increased methylation of CpG islands proximal to the promoters of these genes; furthermore, the DNA demethylating agent 5 aza-2'deoxycytidine (5-Aza-dC) antagonizes the effects of AID knockdown on the expression of EMT factors. We conclude that AID is necessary for the EMT in this breast cancer cell model and in nontransformed mammary epithelial cells. Our results suggest that AID may act near the apex of a hierarchy of regulatory steps that drive the EMT, and are consistent with this effect being mediated by cytosine demethylation. This evidence links our findings to other reports of a role for AID in epigenetic reprogramming and control of gene expression. PMID:23882083

MicroRNA-29b regulates TGF-β1-mediated epithelial–mesenchymal transition of retinal pigment epithelial cells by targeting AKT2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Min; Li, Hui; Liu, Xiaoqiang

2016-07-15

The role of microRNA (miRNA) in proliferative vitreoretinopathy (PVR) progression has not been studied extensively, especially in retinal pigment epithelial–mesenchymal transition (EMT) which is the main reason for formation of PVR. In this study, we first investigated the miRNA expression profile in transforming growth factor beta 1 (TGF-β1) mediated EMT of ARPE-19 cells. Among the five changed miRNAs, miR-29b showed the most significant downregulation. Enhanced expression of miR-29b could reverse TGF-β1 induced EMT through targeting Akt2. Akt2 downregulation could inhibit TGF-β1-induced EMT. Furthermore, inhibition of miR-29b in ARPE-19 cells directly triggered EMT process, which characterized by the phenotypic transition andmore » the upregulation of α-smooth muscle actin (α-SMA) and downregulation of E-cadherin and zona occludin-1 (ZO-1) with increased cell migration. Akt2-shRNA also inhibited miR-29 inhibitor-induced EMT process. These data indicate that miR-29b plays an important role in TGF-β1-mediated EMT in ARPE-19 cells by targeting Akt2. - Highlights: • MiR-29b expression is decreased in TGF-β1-induced EMT of ARPE-19 cells. • MiR-29b inhibits TGF-β1-induced EMT in ARPE-19 cells. • MiR-29b inhibitor induces EMT in ARPE-19 cells. • Akt2 is the target for miR-29b. • Downregulation of Akt2 prevents TGF-β1-induced EMT of ARPE-19 cells.« less

Small interfering RNA targeting ILK inhibits metastasis in human tongue cancer cells through repression of epithelial-to-mesenchymal transition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xing, Yu; Laboratory of Forensic Medicine and Biomedical Information, Chongqing Medical University, Chongqing; Qi, Jin

2013-08-01

Integrin-linked kinase (ILK) is a multifunctional serine/threonine kinase. Accumulating evidences suggest that ILK are involved in cell–matrix interactions, cell proliferation, invasion, migration, angiogenesis and Epithelial–mesenchymal transition (EMT). However, the underlying mechanisms remain largely unknown. EMT has been postulated as a prerequisite for metastasis. The reports have demonstrated that EMT was implicated in metastasis of oral squamous cell carcinomas. Therefore, here we further postulate that ILK might participate in EMT of tongue cancer. We showed that ILK siRNA inhibited EMT with low N-cadherin, Vimentin, Snail, Slug and Twist as well as high E-cadherin expression in vivo and in vitro. We foundmore » that knockdown of ILK inhibited cell proliferation, migration and invasion as well as changed cell morphology. We also demonstrated that ILK siRNA inhibited phosphorylation of downstream signaling targets Akt and GSK3β as well as reduced expression of MMP2 and MMP9. Furthermore, we found that the tongue tumor with high metastasis capability showed higher ILK, Vimentin, Snail, Slug and Twist as well as lower E-cadherin expression in clinical specimens. Finally, ILK siRNA led to the suppression for tumorigenesis and metastasis in vivo. Our findings suggest that ILK could be a novel diagnostic and therapeutic target for tongue cancer. Highlights: • ILK siRNA influences cell morphology, cell cycle, migration and invasion. • ILK siRNA affects the expression of proteins associated with EMT. • ILK expression is related to EMT in clinical human tongue tumors. • ILK siRNA inhibits metastasis of the tongue cancer cells through suppressing EMT.« less

CXCR6 predicts poor prognosis in gastric cancer and promotes tumor metastasis through epithelial-mesenchymal transition.

PubMed

Jin, Jie-Jie; Dai, Fa-Xiang; Long, Zi-Wen; Cai, Hong; Liu, Xiao-Wen; Zhou, Ye; Hong, Qi; Dong, Qiong-Zhu; Wang, Ya-Nong; Huang, Hua

2017-06-01

Chemokines and their receptors have been confirmed to be involved in several types of cancer. However, little is known concerning the role of CXCL16 and its receptor CXCR6 in gastric cancer (GC) progression and metastasis. In the present study, expression of CXCL16 and CXCR6 in GC tumor and peritumoral tissues was detected by immunohistochemistry (IHC) in a cohort of 352 GC patients who underwent gastrectomy, and the correlation between CXCL16/CXCR6 expression and clinicopathological characteristics was further analyzed. To evaluate the function of CXCR6, we overexpressed and knocked down CXCR6 in GC cell lines. Results showed that expression of CXCR6, but not CXCL16, was significantly upregulated in GC tumor tissues, and was significantly correlated with lymph node and distant metastases, and advanced clinical stage in the GC patients. Survival analysis showed that large tumor size (>5 cm), elevated preoperative serum carcinoembryonic antigen (CEA) level, advanced TNM stage and high CXCR6 expression indicated worse overall survival (OS) and disease-free survival (DFS) in GC, and CXCR6 was an independent predictor for both OS and DFS in GC. In vitro experiments showed that CXCR6 overexpression induced cell migration and invasion ability, and promoted epithelial-mesenchymal transition of GC cells by upregulation of mesenchymal markers and inhibition of epithelial markers. In contrast, knockdown of CXCR6 in GC cells resulted in inhibition of cell proliferation, migration and invasion ability, and reversal of epithelial-mesenchymal transition (EMT) phenomenon. Our results demonstrated that CXCR6 is an independent prognostic factor for poor survival in GC patients, and may promote GC metastasis through EMT.

Poricoic acid ZA, a novel RAS inhibitor, attenuates tubulo-interstitial fibrosis and podocyte injury by inhibiting TGF-β/Smad signaling pathway.

PubMed

Wang, Ming; Chen, Dan-Qian; Wang, Min-Chang; Chen, Hua; Chen, Lin; Liu, Dan; Zhao, Hui; Zhao, Ying-Yong

2017-12-01

The pathogenesis of tubulo-interstitial fibrosis and glomerulosclerosisis was characterized by cellular hypertrophy, extracellular matrix accumulation and podocyte detachment. Poricoic acid ZA (PZA) is a tetracyclic triterpenoid compound extracted from the surface layer of Poria cocos (LPC), which have been used extensively for diuretic and renoprotective effects. The anti-fibrotic effect of PZA is investigated in HK-2 cells and podocytes induced by TGF-β1 and angiotensin II (ANGII). qRT-PCR, siRNA, immunofluorescence staining, co-immunoprecipitation and Western blot analyses are used to evaluate the expression of RAS signaling, TGF-β/Smad pathway, epithelial-to-mesenchymal transition (EMT) and podocyte markers. PZA restores the mRNA and protein expression of EMT in HK-2 cells. Specific TGF-β1-siRNA efficiently blocks ANGII-induced protein expression of TGF-β1 and further inhibits activated Smad signaling. PZA significantly attenuates up-regulation of angiotensinogen, renin, ACE and AT1. Further, PZA reverses up-regulation of TGFβRII and suppresses Smad proteins. Simultaneously, PZA inhibits the protein interaction of TGF-β receptor and Smads and PZA also inhibits activated RAS and TGF-β/Smad signaling cascade and up-regulates protein expression of podocyte markers and mitigates podocyte injury. This study demonstrated the beneficial role of PZA in renal fibrosis and podocyte injury. Our study highlighted that PZA inhibits RAS and further suppresses TGF-β/Smad pathway through inhibiting Smad2/3 phosphorylation via blocking Smad2/3-TGFβRI protein interaction. PZA is implicated in activation of RAS/TGF-β/Smad axis in HK-2 cells and podocytes. PZA could be considered as a novel RAS inhibitor for treating CKD. Copyright © 2017 Elsevier GmbH. All rights reserved.

Prognostic value of the chemokine receptor CXCR4 and epithelial-to-mesenchymal transition in patients with squamous cell carcinoma of the mobile tongue.

PubMed

Albert, Sébastien; Hourseau, Muriel; Halimi, Caroline; Serova, Maria; Descatoire, Véronique; Barry, Béatrix; Couvelard, Anne; Riveiro, Maria Eugenia; Tijeras-Raballand, Annemilaï; de Gramont, Armand; Raymond, Eric; Faivre, Sandrine

2012-12-01

The aim of this study was to evaluate the expression and the prognostic value of chemokine receptor 4 (CXCR4), its cognate ligand the CXCL12, and markers of epithelial-to-mesenchymal transition (EMT) in squamous cell carcinoma (SCC) of the mobile tongue. Patients with primary SCC of the mobile tongue who underwent surgery in our center were screened retrospectively. Patients without prior treatment, who had pre-surgery TNM staging and available tumor samples, were eligible. Protein expression of CXCL12, CXCR4, CA9, E-cadherin, and vimentin was determined by immunohistochemical staining, scored, and correlated with clinical and pathological parameters and overall survival. Multivariate and Cox proportional hazards analyses were performed. Among 160 patients treated and screened, 47 were analyzed. CXCR4 and CXCL12 expression was high in tumor cells. CXCR4 expression in primary tumor samples was significantly higher in patients with high-grade tumors, lymph node metastases, and microscopic nerve invasion (p ≤ 0.05). There was a non-significant trend towards a correlation between high CXCL12 expression and pathologic tumor stage (p=0.07). Tumors with high CXCR4 expression correlated with poor overall survival (hazard ratio=3.6, 95% confidence interval 1.3-9.7; p=0.011), notably in the CXCR4(high)/vimentin-positive subgroup. Vimentin-positive tumors, characterizing EMT, were associated with lower survival (hazard ratio=4.5, 95% confidence interval 1.6-12.3; p=0.0086). Multivariate analysis confirmed vimentin (but not CXCR4) expression as an independent prognostic factor of poor overall survival (p=0.016). Our results suggest that CXCR4 is a marker of tumor aggressiveness and vimentin is an important and independent prognostic factor in patients with SCC of the mobile tongue. Copyright © 2012 Elsevier Ltd. All rights reserved.

The prognostic role of the epithelial-mesenchymal transition markers E-cadherin and Slug in laryngeal squamous cell carcinoma.

PubMed

Cappellesso, Rocco; Marioni, Gino; Crescenzi, Marika; Giacomelli, Luciano; Guzzardo, Vincenza; Mussato, Alessio; Staffieri, Alberto; Martini, Alessandro; Blandamura, Stella; Fassina, Ambrogio

2015-10-01

Laryngeal squamous cell carcinoma (LSCC) prognosis is definitely related to lymph node metastasis. Epithelial-mesenchymal transition (EMT) allows neoplastic cells to gain the plasticity and motility required for tumour progression and metastasis. The aim of this study was to investigate the role of EMT in the prognosis of LSCC. Immunohistochemical analysis of E-cadherin, N-cadherin, Snail, Slug, ZEB1, and ZEB2 was performed in 37 consecutive LSCC cases. Low E-cadherin levels and high Slug levels correlated with both disease recurrence (P = 0.02 and P =0.01, respectively) and shorter disease-free survival (DFS) (P = 0.04 and P = 0.02, respectively). Relative expression levels of CDH1, SNAI2, miR-1 and the miR-200 family were also evaluated. CDH1, miR-200a and miR-200c down-regulation and SNAI2 overexpression were significantly associated with disease recurrence (P = 0.03, P = 0.02, P = 0.04, and P = 0.04, respectively). EMT increases tumour recurrence risk and shortens DFS in LSCC. E-cadherin and Slug immunohistochemical analysis could be useful for identifying patients requiring more aggressive treatment after surgery. © 2015 John Wiley & Sons Ltd.

Lysophosphatidic Acid Initiates Epithelial to Mesenchymal Transition and Induces β-Catenin-mediated Transcription in Epithelial Ovarian Carcinoma*

PubMed Central

Burkhalter, Rebecca J.; Westfall, Suzanne D.; Liu, Yueying; Stack, M. Sharon

2015-01-01

During tumor progression, epithelial ovarian cancer (EOC) cells undergo epithelial-to-mesenchymal transition (EMT), which influences metastatic success. Mutation-dependent activation of Wnt/β-catenin signaling has been implicated in gain of mesenchymal phenotype and loss of differentiation in several solid tumors; however, similar mutations are rare in most EOC histotypes. Nevertheless, evidence for activated Wnt/β-catenin signaling in EOC has been reported, and immunohistochemical analysis of human EOC tumors demonstrates nuclear staining in all histotypes. This study addresses the hypothesis that the bioactive lipid lysophosphatidic acid (LPA), prevalent in the EOC microenvironment, functions to regulate EMT in EOC. Our results demonstrate that LPA induces loss of junctional β-catenin, stimulates clustering of β1 integrins, and enhances the conformationally active population of surface β1 integrins. Furthermore, LPA treatment initiates nuclear translocation of β-catenin and transcriptional activation of Wnt/β-catenin target genes resulting in gain of mesenchymal marker expression. Together these data suggest that LPA initiates EMT in ovarian tumors through β1-integrin-dependent activation of Wnt/β-catenin signaling, providing a novel mechanism for mutation-independent activation of this pathway in EOC progression. PMID:26175151

WISP-2 in human gastric cancer and its potential metastatic suppressor role in gastric cancer cells mediated by JNK and PLC-γ pathways.

PubMed

Ji, Jiafu; Jia, Shuqin; Jia, Yongning; Ji, Ke; Hargest, Rachel; Jiang, Wen G

2015-09-15

It has recently been shown that WISP proteins (Wnt-inducted secreted proteins), a group of intra- and extra-cellular regulatory proteins, have been implicated in the initiation and progression of a variety of tumour types including colorectal and breast cancer. However, the role of WISP proteins in gastric cancer (GC) cells and their clinical implications have not yet been elucidated. The expression of WISP molecules in a cohort of GC patients was analysed using real-time quantitative PCR and immunohistochemistry. The expression of a panel of recognised epithelial-mesenchymal transition (EMT) markers was quantified using Q-PCR in paired tumour and normal tissues. WISP-2 knockdown (kd) sublines using ribozyme transgenes were created in the GC cell lines AGS and HGC27. Subsequently, several biological functions, including cell growth, adhesion, migration and invasion, were studied. Potential pathways for the interaction of EMT, extracellular matrix and MMP were evaluated. Overexpression of WISP-2 was detected in GC and significantly correlated with early tumour node-metastasis staging, differentiation status and positively correlated with overall survival and disease-free survival of the patients. WISP-2 expression was inversely correlated with that of Twist and Slug in paired samples. Kd of WISP-2 expression promoted the proliferation, migration and invasion of GC cells. WISP-2 suppressed GC cell metastasis through reversing EMT and suppressing the expression and activity of MMP9 and MMP2 via JNK and ERK. Cell motility analysis indicated that WISP-2 kd contributed to GC cells' motility and can be attenuated by PLC-γ and JNK small inhibitors. Increased expression of WISP-2 in GC is positively correlated with favourable clinical features and the survival of patients with GC and is a negative regulator of growth, migration and invasion in GC cells. These findings suggest that WISP-2 is a potential tumour suppressor in GC.

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

PubMed Central

Gawronska-Kozak, Barbara; Grabowska, Anna; Kur-Piotrowska, Anna; Kopcewicz, Marta

2016-01-01

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. PMID:26938103

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.

PubMed

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 own CAFs via EMT in this model.

Epithelial-mesenchymal transition (EMT) induced by TNF-α requires AKT/GSK-3β-mediated stabilization of snail in colorectal cancer.

PubMed

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.

Phoyunnanin E inhibits migration of non-small cell lung cancer cells via suppression of epithelial-to-mesenchymal transition and integrin αv and integrin β3.

PubMed

Petpiroon, Nareerat; Sritularak, Boonchoo; Chanvorachote, Pithi

2017-12-29

The conversion of the epithelial phenotype of cancer cells into cells with a mesenchymal phenotype-so-called epithelial-mesenchymal transition (EMT)-has been shown to enhance the capacity of the cells to disseminate throughout the body. EMT is therefore becoming a potential target for anti-cancer drug discovery. Here, we showed that phoyunnanin E, a compound isolated from Dendrobium venustum, possesses anti-migration activity and addressed its mechanism of action. The cytotoxic and proliferative effects of phoyunnanin E on human non-small cell lung cancer-derived H460, H292, and A549 cells and human keratinocyte HaCaT cells were investigated by MTT assay. The effect of phoyunnanin E on EMT was evaluated by determining the colony formation and EMT markers. The migration and invasion of H460, H292, A549 and HaCaT cells was evaluated by wound healing assay and transwell invasion assay, respectively. EMT markers, integrins and migration-associated proteins were examined by western blot analysis. Phoyunnanin E at the concentrations of 5 and 10 μM, which are non-toxic to H460, H292, A549 and HaCaT cells showed good potential to inhibit the migratory activity of three types of human lung cancer cells. The anti-migration effect of phoyunnanin E was shown to relate to the suppressed EMT phenotypes, including growth in anchorage-independent condition, cell motility, and EMT-specific protein markers (N-cadherin, vimentin, slug, and snail). In addition to EMT suppression, we found that phoyunnanin E treatment with 5 and 10 μM could decrease the cellular level of integrin αv and integrin β3, these integrins are frequently up-regulated in highly metastatic tumor cells. We further characterized the regulatory proteins in cell migration and found that the cells treated with phoyunnanin E exhibited a significantly lower level of phosphorylated focal adhesion kinase (p-FAK) and phosphorylated ATP-dependent tyrosine kinase (p-AKT), and their downstream effectors (including Ras-related C3 botulinum (Rac-GTP); Cell division cycle 42 (Cdc42); and Ras homolog gene family, member A (Rho-GTP)) in comparison to those of the non-treated control. We have determined for the first time that phoyunnanin E could inhibit the motility of lung cancer cells via the suppression of EMT and metastasis-related integrins. This new information could support further development of this compound for anti-metastasis approaches.

Sip1 mediates an E-cadherin-to-N-cadherin switch during cranial neural crest EMT

PubMed Central

Rogers, Crystal D.; Saxena, Ankur

2013-01-01

The neural crest, an embryonic stem cell population, initially resides within the dorsal neural tube but subsequently undergoes an epithelial-to-mesenchymal transition (EMT) to commence migration. Although neural crest and cancer EMTs are morphologically similar, little is known regarding conservation of their underlying molecular mechanisms. We report that Sip1, which is involved in cancer EMT, plays a critical role in promoting the neural crest cell transition to a mesenchymal state. Sip1 transcripts are expressed in premigratory/migrating crest cells. After Sip1 loss, the neural crest specifier gene FoxD3 was abnormally retained in the dorsal neuroepithelium, whereas Sox10, which is normally required for emigration, was diminished. Subsequently, clumps of adherent neural crest cells remained adjacent to the neural tube and aberrantly expressed E-cadherin while lacking N-cadherin. These findings demonstrate two distinct phases of neural crest EMT, detachment and mesenchymalization, with the latter involving a novel requirement for Sip1 in regulation of cadherin expression during completion of neural crest EMT. PMID:24297751

Inactivation of p53 in pterygium influence miR-200a expression resulting in ZEB1/ZEB2 up-regulation and EMT processing.

PubMed

Wu, Chueh-Wei; Peng, Mei-Ling; Yeh, Ken-Tu; Tsai, Yi-Yu; Chiang, Chun-Chi; Cheng, Ya-Wen

2016-05-01

Loss of p53 function has been linked to progression of pterygium. MiR-200a is known to be controlled by p53. Here, we hypothesize that expression of miR-200a and downstream ZEB1/ZEB2 genes are regulated epithelial-mesenchymal transition (EMT) involved in the pathogenesis and recurrence of pterygium. For this study, 120 primary pterygial samples were collected. Immunohistochemistry and real-time RT-PCR were performed to determine the expression of p53, p53 down-stream EMT associated protein and miR-200a. The molecular correlation of p53, miR-200a and downstream genes were confirmed using primary pterygium cells (PECs). Expression of miR-200a in pterygium tissues was significantly lower than in conjunctiva controls (p = 0.015). Up-regulated miR-200a levels were positively correlated with and p53 protein expression (p < 0.001). The miR-200a downstream ZEB1/ZEB1 protein expression were negative correlated with miR-200a expression. Cell model studies demonstrated that miR-200a controlled the EMT of PECs through up-regulated ZEB1, ZEB2 and Snail gene expression. Our study demonstrated that inactivation of p53 in pterygium may influence miR-200a, resulting in ZEB1/ZEB2 up-regulation and EMT processing of pterygium. Therefore, we suggest that expression of miR-200a play an important role in EMT processing and recurrence of pterygium. Copyright © 2016 Elsevier Ltd. All rights reserved.

TWIST1-induced microRNA-424 reversibly drives mesenchymal programming while inhibiting tumor initiation

PubMed Central

Drasin, David J.; Guarnieri, Anna L.; Neelakantan, Deepika; Kim, Jihye; Cabrera, Joshua H.; Wang, Chu-An; Zaberezhnyy, Vadym; Gasparini, Pierluigi; Cascione, Luciano; Huebner, Kay; Tan, Aik-Choon; Ford, Heide L.

2015-01-01

Epithelial-to-mesenchymal transition (EMT) is a dynamic process that relies on cellular plasticity. Recently, the process of an oncogenic EMT, followed by a reverse mesenchymal-to-epithelial transition (MET), has been implicated as critical in the metastatic colonization of carcinomas. Unlike governance of epithelial programming, regulation of mesenchymal programming is not well understood in EMT. Here, we describe and characterize the first microRNA that enhances exclusively mesenchymal programming. We demonstrate that microRNA-424 is upregulated early during a TWIST1 or SNAI1-induced EMT, and that it causes cells to express mesenchymal genes without affecting epithelial genes, resulting in a mixed/intermediate EMT. Furthermore, microRNA-424 increases motility, decreases adhesion and induces a growth arrest, changes associated with a complete EMT, that can be reversed when microRNA-424 expression is lowered, concomitant with an MET-like process. Breast cancer patient microRNA-424 levels positively associate with TWIST1/2 and EMT-like gene signatures, and miR-424 is increased in primary tumors versus matched normal breast. However, microRNA-424 is downregulated in patient metastases versus matched primary tumors. Correspondingly, microRNA-424 decreases tumor initiation and is post-transcriptionally downregulated in macrometastases in mice, suggesting the need for biphasic expression of miR-424 to transit the EMT-MET axis. Next-generation RNA sequencing revealed microRNA-424 regulates numerous EMT and cancer stemness-associated genes, including TGFBR3, whose downregulation promotes mesenchymal phenotypes, but not tumor-initiating phenotypes. Instead, we demonstrate that increased MAPK/ERK signaling is critical for miR-424-mediated decreases in tumor-initiating phenotypes. These findings suggest microRNA-424 plays distinct roles in tumor progression, potentially facilitating earlier, but repressing later, stages of metastasis by regulating an EMT-MET axis. PMID:25716682

Tg737 regulates epithelial-mesenchymal transition and cancer stem cell properties via a negative feedback circuit between Snail and HNF4α during liver stem cell malignant transformation.

PubMed

Huang, Qike; Pu, Meng; Zhao, Ge; Dai, Bin; Bian, Zhenyuan; Tang, Haili; Chen, Chong; Liu, Wei; Qu, Xuan; Shen, Liangliang; Tao, Kaishan

2017-08-28

Determining the origin of liver cancer stem cells is important for treating hepatocellular carcinoma. Tg737 deficiency plays an important role in the malignant transformation of liver stem cells, but the underlying mechanism remains unclear. Here we established a chemical-induced mouse hepatoma model and found that Tg737 and hepatocyte nuclear factor 4-alpha (HNF4α) expression decreased and epithelial-mesenchymal transition (EMT)-related marker expression increased during liver cancer development. To investigate the underlying mechanism, we knocked down Tg737 in WB-F344 (WB) rat hepatic oval cells. Loss of Tg737 resulted in nuclear β-catenin accumulation and activation of the Wnt/β-catenin pathway, which further promoted EMT and the malignant phenotype. XAV939, a β-catenin inhibitor, attenuated WB cell malignant transformation due to Tg737 knockdown. To clarify the relationships of Tg737, the β-catenin pathway, and HNF4α, we inhibited Snail and overexpressed HNF4α after Tg737 knockdown in WB cells and found that Snail and HNF4α comprise a negative feedback circuit. Taken together, the results showed that Tg737 regulates a Wnt/β-catenin/Snail-HNF4α negative feedback circuit, thereby blocking EMT and the malignant transformation of liver stem cells to liver cancer stem cells. Copyright © 2017 Elsevier B.V. All rights reserved.

3-bromopyruvate and buthionine sulfoximine effectively kill anoikis-resistant hepatocellular carcinoma cells

PubMed Central

Lee, Minjong; Jo, Ara; Lee, Seulki; Kim, Jong Bin; Chang, Young; Nam, Joon Yeul; Cho, Hyeki; Cho, Young Youn; Cho, Eun Ju; Lee, Jeong-Hoon; Yu, Su Jong; Yoon, Jung-Hwan

2017-01-01

Background & aims Acquisition of anoikis resistance is a prerequisite for metastasis in hepatocellular carcinoma (HCC). However, little is known about how energy metabolism and antioxidant systems are altered in anoikis-resistant (AR) HCC cells. We evaluated anti-tumor effects of a combination treatment of 3-bromopyruvate (3-BP) and buthionine sulfoximine (BSO) in AR HCC cells. Methods We compared glycolysis, reactive oxygen species (ROS) production, and chemoresistance among Huh-BAT, HepG2 HCC cells, and the corresponding AR cells. Expression of hexokinase II, gamma-glutamylcysteine synthetase (rGCS), and epithelial–mesenchymal transition (EMT) markers in AR cells was assessed. Anti-tumor effects of a combination treatment of 3-BP and BSO were evaluated in AR cells and an HCC xenograft mouse model. Results AR HCC cells showed significantly higher chemoresistance, glycolysis and lower ROS production than attached cells. Expression of hexokinase II, rGCS, and EMT markers was higher in AR HCC cells than attached cells. A combination treatment of 3-BP/BSO effectively suppressed proliferation of AR HCC cells through apoptosis by blocking glycolysis and enhancing ROS levels. In xenograft mouse models, tumor growth derived from AR HCC cells was significantly suppressed in the group treated with 3-BP/BSO compared to the group treated with 3-BP or sorafenib. Conclusions These results demonstrated that a combination treatment of 3-BP/BSO had a synergistic anti-tumor effect in an AR HCC model. This strategy may be an effective adjuvant therapy for patients with sorafenib-resistant HCC. PMID:28362858

3-bromopyruvate and buthionine sulfoximine effectively kill anoikis-resistant hepatocellular carcinoma cells.

PubMed

Lee, Minjong; Jo, Ara; Lee, Seulki; Kim, Jong Bin; Chang, Young; Nam, Joon Yeul; Cho, Hyeki; Cho, Young Youn; Cho, Eun Ju; Lee, Jeong-Hoon; Yu, Su Jong; Yoon, Jung-Hwan; Kim, Yoon Jun

2017-01-01

Acquisition of anoikis resistance is a prerequisite for metastasis in hepatocellular carcinoma (HCC). However, little is known about how energy metabolism and antioxidant systems are altered in anoikis-resistant (AR) HCC cells. We evaluated anti-tumor effects of a combination treatment of 3-bromopyruvate (3-BP) and buthionine sulfoximine (BSO) in AR HCC cells. We compared glycolysis, reactive oxygen species (ROS) production, and chemoresistance among Huh-BAT, HepG2 HCC cells, and the corresponding AR cells. Expression of hexokinase II, gamma-glutamylcysteine synthetase (rGCS), and epithelial-mesenchymal transition (EMT) markers in AR cells was assessed. Anti-tumor effects of a combination treatment of 3-BP and BSO were evaluated in AR cells and an HCC xenograft mouse model. AR HCC cells showed significantly higher chemoresistance, glycolysis and lower ROS production than attached cells. Expression of hexokinase II, rGCS, and EMT markers was higher in AR HCC cells than attached cells. A combination treatment of 3-BP/BSO effectively suppressed proliferation of AR HCC cells through apoptosis by blocking glycolysis and enhancing ROS levels. In xenograft mouse models, tumor growth derived from AR HCC cells was significantly suppressed in the group treated with 3-BP/BSO compared to the group treated with 3-BP or sorafenib. These results demonstrated that a combination treatment of 3-BP/BSO had a synergistic anti-tumor effect in an AR HCC model. This strategy may be an effective adjuvant therapy for patients with sorafenib-resistant HCC.

ROCK inhibitor primes human induced pluripotent stem cells to selectively differentiate towards mesendodermal lineage via epithelial-mesenchymal transition-like modulation.

PubMed

Maldonado, Maricela; Luu, Rebeccah J; Ramos, Michael E P; Nam, Jin

2016-09-01

Robust control of human induced pluripotent stem cell (hIPSC) differentiation is essential to realize its patient-tailored therapeutic potential. Here, we demonstrate a novel application of Y-27632, a small molecule Rho-associated protein kinase (ROCK) inhibitor, to significantly influence the differentiation of hIPSCs in a lineage-specific manner. The application of Y-27632 to hIPSCs resulted in a decrease in actin bundling and disruption of colony formation in a concentration and time-dependent manner. Such changes in cell and colony morphology were associated with decreased expression of E-cadherin, a cell-cell junctional protein, proportional to the increased exposure to Y-27632. Interestingly, gene and protein expression of pluripotency markers such as NANOG and OCT4 were not downregulated by an exposure to Y-27632 up to 36h. Simultaneously, epithelial-to-mesenchymal (EMT) transition markers were upregulated with an exposure to Y-27632. These EMT-like changes in the cells with longer exposure to Y-27632 resulted in a significant increase in the subsequent differentiation efficiency towards mesendodermal lineage. In contrast, an inhibitory effect was observed when cells were subjected to ectodermal differentiation after prolonged exposure to Y-27632. Collectively, these results present a novel method for priming hIPSCs to modulate their differentiation potential with a simple application of Y-27632. Copyright © 2016 Helmholtz Zentrum München. Published by Elsevier B.V. All rights reserved.

Octamer-binding protein 4 affects the cell biology and phenotypic transition of lung cancer cells involving β-catenin/E-cadherin complex degradation.

PubMed

Chen, Zhong-Shu; Ling, Dong-Jin; Zhang, Yang-De; Feng, Jian-Xiong; Zhang, Xue-Yu; Shi, Tian-Sheng

2015-03-01

Clinical studies have reported evidence for the involvement of octamer‑binding protein 4 (Oct4) in the tumorigenicity and progression of lung cancer; however, the role of Oct4 in lung cancer cell biology in vitro and its mechanism of action remain to be elucidated. Mortality among lung cancer patients is more frequently due to metastasis rather than their primary tumors. Epithelial‑mesenchymal transition (EMT) is a prominent biological event for the induction of epithelial cancer metastasis. The aim of the present study was to investigate whether Oct4 had the capacity to induce lung cancer cell metastasis via the promoting the EMT in vitro. Moreover, the effect of Oct4 on the β‑catenin/E‑cadherin complex, associated with EMT, was examined using immunofluorescence and immunoprecipitation assays as well as western blot analysis. The results demonstrated that Oct4 enhanced cell invasion and adhesion accompanied by the downregulation of epithelial marker cytokeratin, and upregulation of the mesenchymal markers vimentin and N‑cadherin. Furthermore, Oct4 induced EMT of lung cancer cells by promoting β‑catenin/E‑cadherin complex degradation and regulating nuclear localization of β‑catenin. In conclusion, the present study indicated that Oct4 affected the cell biology of lung cancer cells in vitro through promoting lung cancer cell metastasis via EMT; in addition, the results suggested that the association and degradation of the β‑catenin/E‑cadherin complex was regulated by Oct4 during the process of EMT.

Helicobacter pylori potentiates epithelial:mesenchymal transition in gastric cancer: links to soluble HB-EGF, gastrin and matrix metalloproteinase-7

PubMed Central

Yin, Yinfei; Grabowska, Anna M; Clarke, Philip A; Whelband, Elisabeth; Robinson, Karen; Argent, Richard H; Tobias, Amanda; Kumari, Rajendra; Atherton, John C

2010-01-01

Background and aims Helicobacter pylori (H pylori) infection is a major risk factor in the development of distal gastric adenocarcinoma. Development of the invasive phenotype is associated with the phenomenon of epithelial:mesenchymal transition (EMT). Soluble heparin-binding epidermal growth factor (HB-EGF) has been implicated in this process. A study was undertaken to investigate the possibility that matrix metalloproteinase (MMP)-7 is upregulated in H pylori infection as a result of hypergastrinaemia, which may enhance shedding of HB-EGF and contribute towards EMT in gastric adenocarcinoma cell lines. Methods Three gastric epithelial cell lines (AGS, MGLVA1 and ST16) were co-cultured with the pathogenic H pylori strain 60190 and non-pathogenic strain Tx30a in an in vitro infection model. Gene expression was quantified by real-time PCR, HB-EGF shedding by ELISA and protein expression by immunofluorescence or immunohistochemistry. The INS-GAS mouse, a transgenic mouse model of gastric carcinogenesis which overexpresses amidated gastrin, was used to investigate the in vivo relationship between HB-EGF, MMP-7, gastrin and EMT. Results The pathogenic strain of H pylori significantly upregulated EMT-associated genes Snail, Slug and vimentin in all three gastric cell lines to a greater degree than the non-pathogenic strain. Pathogenic H pylori also upregulated HB-EGF shedding, a factor implicated in EMT, which was partially dependent on both gastrin and MMP-7 expression. Gastrin and MMP-7 siRNAs and MMP-7 neutralising antibody significantly reduced upregulation of HB-EGF shedding in H pylori infected gastric cell lines and reduced EMT gene expression. The effect of H pylori on EMT was also reversed by gastrin siRNA. Neutralisation of gastrin in the INS-GAS mouse model reduced expression of MMP-7, HB-EGF and key EMT proteins. Conclusion The upregulation of MMP-7 by pathogenic H pylori is partially dependent on gastrin and may have a role in the development of gastric cancer, potentially through EMT, by indirectly increasing levels of soluble HB-EGF. PMID:20584780

Metformin inhibits 17β-estradiol-induced epithelial-to-mesenchymal transition via βKlotho-related ERK1/2 signaling and AMPKα signaling in endometrial adenocarcinoma cells.

PubMed

Liu, Zhao; Qi, Shasha; Zhao, Xingbo; Li, Mingjiang; Ding, Sentai; Lu, Jiaju; Zhang, Hui

2016-04-19

The potential role of metformin in treating endometrial cancer remains to be explored. The current study investigated the role of metformin in 17β-estradiol-induced epithelial-mesenchymal transition (EMT) in endometrial adenocarcinoma cells. We found that 17β-estradiol promoted proliferation and migration, attenuated apoptosis in both estrogen receptor (ER) positive and ER negative endometrial adenocarcinoma cells (Ishikawa and KLE cells, respectively). Metformin abolished 17β-estradiol-induced cell proliferation and reversed 17β-estradiol-induced EMT in Ishikawa cells. In addition, metformin increased the expression of βKlotho, a fibroblast growth factors (FGFs) coreceptor, and decreased ERK1/2 phosphorylation in both Ishikawa and KLE cells. Decreased expression of βKlotho was noted in human endometrial adenocarcinomas, and plasmid-driven expression of βKlotho in Ishikawa cells abolished 17β-estradiol-induced EMT via inhibiting ERK1/2 signaling. βKlotho expression and metformin show synergetic effects on the proliferation and the EMT in Ishikawa cells. Furthermore, we demonstrated that the anti-EMT effects of metformin could be partly abolished by introducing Compound C, a specific AMPKα signaling inhibitor. In conclusion, metformin abolishes 17β-estradiol-induced cell proliferation and EMT in endometrial adenocarcinoma cells by upregulating βKlotho expression, inhibiting ERK1/2 signaling, and activating AMPKα signaling. Our study provides novel mechanistic insight into the anti-tumor effects of metformin.

Metformin inhibits 17β-estradiol-induced epithelial-to-mesenchymal transition via βKlotho-related ERK1/2 signaling and AMPKα signaling in endometrial adenocarcinoma cells

PubMed Central

Liu, Zhao; Qi, Shasha; Zhao, Xingbo; Li, Mingjiang; Ding, Sentai; Lu, Jiaju; Zhang, Hui

2016-01-01

The potential role of metformin in treating endometrial cancer remains to be explored. The current study investigated the role of metformin in 17β-estradiol-induced epithelial-mesenchymal transition (EMT) in endometrial adenocarcinoma cells. We found that 17β-estradiol promoted proliferation and migration, attenuated apoptosis in both estrogen receptor (ER) positive and ER negative endometrial adenocarcinoma cells (Ishikawa and KLE cells, respectively). Metformin abolished 17β-estradiol-induced cell proliferation and reversed 17β-estradiol-induced EMT in Ishikawa cells. In addition, metformin increased the expression of βKlotho, a fibroblast growth factors (FGFs) coreceptor, and decreased ERK1/2 phosphorylation in both Ishikawa and KLE cells. Decreased expression of βKlotho was noted in human endometrial adenocarcinomas, and plasmid-driven expression of βKlotho in Ishikawa cells abolished 17β-estradiol-induced EMT via inhibiting ERK1/2 signaling. βKlotho expression and metformin show synergetic effects on the proliferation and the EMT in Ishikawa cells. Furthermore, we demonstrated that the anti-EMT effects of metformin could be partly abolished by introducing Compound C, a specific AMPKα signaling inhibitor. In conclusion, metformin abolishes 17β-estradiol-induced cell proliferation and EMT in endometrial adenocarcinoma cells by upregulating βKlotho expression, inhibiting ERK1/2 signaling, and activating AMPKα signaling. Our study provides novel mechanistic insight into the anti-tumor effects of metformin. PMID:26824324

The potential role of Brachyury in inducing epithelial-to-mesenchymal transition (EMT) and HIF-1α expression in breast cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shao, Chao; Zhang, Jingjing, E-mail: jingjingzhangzs@163.com; Fu, Jianhua

One of transcription factors of the T-box family, Brachyury has been implicated in tumorigenesis of many types of cancers, regulating cancer cell proliferation, metastasis, invasion and epithelial-to-mesenchymal transition (EMT). However, the role of Brachyury in breast cancer cells has been scarcely reported. The present study aimed to investigate the expression and role of Brachyury in breast cancer. Brachyury expression was analyzed by qRT-PCR and Western blot. The correlations between Brachyury expression and clinicopathological factors of breast cancer were determined. Involvement of EMT stimulation and hypoxia-inducible factor-1α (HIF-1α) expression induction by Brachyury was also evaluated. Moreover, the effect of Brachyury onmore » tumor growth and metastasis in vivo was examined in a breast tumor xenograft model. Brachyury expression was enhanced in primary breast cancer tissues and Brachyury expression was correlated with tumor stage and lymph node metastasis. Hypoxia enhanced Brachyury expression, the silencing of which blocked the modulation effect of hypoxia on E-cadherin and vimentin expression. Brachyury significantly augmented HIF-1alpha expression via PTEN/Akt signaling as well as accelerated cell proliferation and migration in vitro. Additionally, Brachyury accelerated breast tumor xenograft growth and increased lung metastasis in nude mice. In summary, our data confirmed that Brachyury might contribute to hypoxia-induced EMT of breast cancer and trigger HIF-1alpha expression via PTEN/Akt signaling. - Highlights: • Brachyury expression was correlated with tumor stage and lymph node metastasis. • Hypoxia enhanced Brachyury expression, which contributes to hypoxia-induced EMT. • Brachyury significantly augmented HIF-1alpha expression via PTEN/Akt signaling. • Brachyury accelerated tumor xenograft growth and increased lung metastasis.« less

Upregulation of microRNA-137 expression by Slug promotes tumor invasion and metastasis of non-small cell lung cancer cells through suppression of TFAP2C.

PubMed

Chang, Tzu-Hua; Tsai, Meng-Feng; Gow, Chien-Hung; Wu, Shang-Gin; Liu, Yi-Nan; Chang, Yih-Leong; Yu, Sung-Liang; Tsai, Hsing-Chen; Lin, Shih-Wen; Chen, Yen-Wei; Kuo, Po-Yen; Yang, Pan-Chyr; Shih, Jin-Yuan

2017-08-28

The epithelial-mesenchymal transition (EMT) regulator, Slug, plays multifaceted roles in controlling lung cancer progression, but its downstream targets and mechanisms in promoting lung cancer progression have not been well defined. In particular, the miRNAs downstream of Slug in non-small cell lung cancer (NSCLC) remain undetermined. Here, we report that miR-137 is downstream of the EMT regulator, Slug, in lung cancer cells. Slug binds directly to the E-box of the miR-137 promoter and up-regulates its expression in lung cancer cells. Knockdown of miR-137 abolished Slug-induced cancer invasion and migration, whereas upregulation of miR-137 was found to trigger lung cancer cell invasion and progression by direct suppressing TFAP2C (transcription factor AP-2 gamma). Clinical data showed that lung adenocarcinoma patients with low-level expression of Slug and miR-137 but high-level expression of TFAP2C experienced significantly better survival. miR-137 is a Slug-induced miRNA that relays the pro-metastatic effects of Slug by targeting TFAP2C. Our findings add new components to the Slug-mediated regulatory network in lung cancer, and suggest that Slug, miR-137, and TFAP2C may be useful prognostic markers in lung adenocarcinoma. Copyright © 2017 Elsevier B.V. All rights reserved.

Epithelial-to-Mesenchymal Transition Antagonizes Response to Targeted Therapies in Lung Cancer by Suppressing BIM.

PubMed

Song, Kyung-A; Niederst, Matthew J; Lochmann, Timothy L; Hata, Aaron N; Kitai, Hidenori; Ham, Jungoh; Floros, Konstantinos V; Hicks, Mark A; Hu, Haichuan; Mulvey, Hillary E; Drier, Yotam; Heisey, Daniel A R; Hughes, Mark T; Patel, Neha U; Lockerman, Elizabeth L; Garcia, Angel; Gillepsie, Shawn; Archibald, Hannah L; Gomez-Caraballo, Maria; Nulton, Tara J; Windle, Brad E; Piotrowska, Zofia; Sahingur, Sinem E; Taylor, Shirley M; Dozmorov, Mikhail; Sequist, Lecia V; Bernstein, Bradley; Ebi, Hiromichi; Engelman, Jeffrey A; Faber, Anthony C

2018-01-01

Purpose: Epithelial-to-mesenchymal transition (EMT) confers resistance to a number of targeted therapies and chemotherapies. However, it has been unclear why EMT promotes resistance, thereby impairing progress to overcome it. Experimental Design: We have developed several models of EMT-mediated resistance to EGFR inhibitors (EGFRi) in EGFR -mutant lung cancers to evaluate a novel mechanism of EMT-mediated resistance. Results: We observed that mesenchymal EGFR -mutant lung cancers are resistant to EGFRi-induced apoptosis via insufficient expression of BIM, preventing cell death despite potent suppression of oncogenic signaling following EGFRi treatment. Mechanistically, we observed that the EMT transcription factor ZEB1 inhibits BIM expression by binding directly to the BIM promoter and repressing transcription. Derepression of BIM expression by depletion of ZEB1 or treatment with the BH3 mimetic ABT-263 to enhance "free" cellular BIM levels both led to resensitization of mesenchymal EGFR -mutant cancers to EGFRi. This relationship between EMT and loss of BIM is not restricted to EGFR -mutant lung cancers, as it was also observed in KRAS -mutant lung cancers and large datasets, including different cancer subtypes. Conclusions: Altogether, these data reveal a novel mechanistic link between EMT and resistance to lung cancer targeted therapies. Clin Cancer Res; 24(1); 197-208. ©2017 AACR . ©2017 American Association for Cancer Research.

Oncogenic ALK regulates EMT in non-small cell lung carcinoma through repression of the epithelial splicing regulatory protein 1.

PubMed

Voena, Claudia; Varesio, Lydia M; Zhang, Liye; Menotti, Matteo; Poggio, Teresa; Panizza, Elena; Wang, Qi; Minero, Valerio G; Fagoonee, Sharmila; Compagno, Mara; Altruda, Fiorella; Monti, Stefano; Chiarle, Roberto

2016-05-31

A subset of Non-Small Cell Lung Carcinoma (NSCLC) carries chromosomal rearrangements involving the Anaplastic Lymphoma Kinase (ALK) gene. ALK-rearranged NSCLC are typically adenocarcinoma characterized by a solid signet-ring cell pattern that is frequently associated with a metastatic phenotype. Recent reports linked the presence of ALK rearrangement to an epithelial-mesenchymal transition (EMT) phenotype in NSCLC, but the extent and the mechanisms of an ALK-mediated EMT in ALK-rearranged NSCLC are largely unknown. We found that the ALK-rearranged H2228 and DFCI032, but not the H3122, cell lines displayed a mesenchymal phenotype. In these cell lines, oncogenic ALK activity dictated an EMT phenotype by directly suppressing E-cadherin and up-regulating vimentin expression, as well as expression of other genes involved in EMT. We found that the epithelial splicing regulatory protein 1 (ESRP1), a key regulator of the splicing switch during EMT, was repressed by EML4-ALK activity. The treatment of NSCLC cells with ALK tyrosine kinase inhibitors (TKIs) led to up-regulation of ESRP1 and E-cadherin, thus reverting the phenotype from mesenchymal to epithelial (MET). Consistently, ESRP1 knock-down impaired E-cadherin up-regulation upon ALK inhibition, whereas enforced expression of ESRP1 was sufficient to increase E-cadherin expression. These findings demonstrate an ALK oncogenic activity in the regulation of an EMT phenotype in a subset of NSCLC with potential implications for the biology of ALK-rearranged NSCLC in terms of metastatic propensity and resistance to therapy.

Oncogenic ALK regulates EMT in non-small cell lung carcinoma through repression of the epithelial splicing regulatory protein 1

PubMed Central

Menotti, Matteo; Poggio, Teresa; Panizza, Elena; Wang, Qi; Minero, Valerio G.; Fagoonee, Sharmila; Compagno, Mara; Altruda, Fiorella; Monti, Stefano; Chiarle, Roberto

2016-01-01

A subset of Non-Small Cell Lung Carcinoma (NSCLC) carries chromosomal rearrangements involving the Anaplastic Lymphoma Kinase (ALK) gene. ALK-rearranged NSCLC are typically adenocarcinoma characterized by a solid signet-ring cell pattern that is frequently associated with a metastatic phenotype. Recent reports linked the presence of ALK rearrangement to an epithelial-mesenchymal transition (EMT) phenotype in NSCLC, but the extent and the mechanisms of an ALK-mediated EMT in ALK-rearranged NSCLC are largely unknown. We found that the ALK-rearranged H2228 and DFCI032, but not the H3122, cell lines displayed a mesenchymal phenotype. In these cell lines, oncogenic ALK activity dictated an EMT phenotype by directly suppressing E-cadherin and up-regulating vimentin expression, as well as expression of other genes involved in EMT. We found that the epithelial splicing regulatory protein 1 (ESRP1), a key regulator of the splicing switch during EMT, was repressed by EML4-ALK activity. The treatment of NSCLC cells with ALK tyrosine kinase inhibitors (TKIs) led to up-regulation of ESRP1 and E-cadherin, thus reverting the phenotype from mesenchymal to epithelial (MET). Consistently, ESRP1 knock-down impaired E-cadherin up-regulation upon ALK inhibition, whereas enforced expression of ESRP1 was sufficient to increase E-cadherin expression. These findings demonstrate an ALK oncogenic activity in the regulation of an EMT phenotype in a subset of NSCLC with potential implications for the biology of ALK-rearranged NSCLC in terms of metastatic propensity and resistance to therapy. PMID:27119231

Tongxinluo ameliorates renal structure and function by regulating miR-21-induced epithelial-to-mesenchymal transition in diabetic nephropathy.

PubMed

Wang, Jin-yang; Gao, Yan-bin; Zhang, Na; Zou, Da-wei; Xu, Li-ping; Zhu, Zhi-yao; Li, Jiao-yang; Zhou, Sheng-nan; Cui, Fang-qiang; Zeng, Xiang-jun; Geng, Jian-guo; Yang, Jin-kui

2014-03-01

Diabetic nephropathy (DN) is one of the most important diabetic microangiopathies. The epithelial-to-mesenchymal transition (EMT) plays an important role in DN. The physiological role of microRNA-21 (miR-21) was closely linked to EMT. However, it remained elusive whether tongxinluo (TXL) ameliorated renal structure and function by regulating miR-21-induced EMT in DN. This study aimed to determine the effect of TXL on miR-21-induced renal tubular EMT and to explore the relationship between miR-21 and TGF-β1/smads signals. Real-time RT-PCR, cell transfection, in situ hybridization (ISH), and laser confocal microscopy were used, respectively. Here, we revealed that TXL dose dependently lowered miR-21 expression in tissue, serum, and cells. Overexpression of miR-21 can enhance α-smooth muscle actin (SMA) expression and decrease E-cadherin expression by upregulating smad3/p-smad3 expression and downregulating smad7 expression. Interestingly, TXL also increased E-cadherin expression and decreased α-SMA expression by regulating miR-21 expression. More importantly, TXL decreased collagen IV, fibronectin, glomerular basement membrane, glomerular area, and the albumin/creatinine ratio, whereas it increased the creatinine clearance ratio. The results demonstrated that TXL ameliorated renal structure and function by regulating miR-21-induced EMT, which was one of the mechanisms to protect against DN, and that miR-21 may be one of the therapeutic targets for TXL in DN.

IGF-1 induces the epithelial-mesenchymal transition via Stat5 in hepatocellular carcinoma.

PubMed

Zhao, Chuanzong; Wang, Qian; Wang, Ben; Sun, Qi; He, Zhaobin; Hong, Jianguo; Kuehn, Florian; Liu, Enyu; Zhang, Zongli

2017-12-19

It has been reported that the epithelial-mesenchymal transition (EMT) plays an important role in hepatocellular carcinoma (HCC). However, the relationship between the insulin-like growth factor-1 (IGF-1) and EMT of HCC was not fully elucidated. In the present work, we found that the expression of N-cadherin, Vimentin, Snail1, Snail2, and Twist1 was positively associated with IGF-1R expression, while E-cadherin expression was negatively associated with IGF-1 expression in human HCC samples. Furthermore, we observed that IGF-1 up-regulated the expression of N-cadherin, Vimentin, Snail1, Snail2 and Twist1, and down-regulated the expression of E-cadherin. In addition, Stat5 was induced in IGF-1-treated HepG2 and Hep3B cells, and Stat5 inhibition or siRNA significantly affected IGF-1-induced EMT in HepG2 and Hep3B cells. In conclusion, IGF-1 induces EMT of HCC via Stat5 signaling pathway. Thus, IGF-1/Stat5 can be recommended as a potential and novel therapeutic strategy for HCC patients.

The expression of cancer stem cell markers in human colorectal carcinoma cells in a microenvironment dependent manner.

PubMed

Stankevicius, Vaidotas; Kunigenas, Linas; Stankunas, Edvinas; Kuodyte, Karolina; Strainiene, Egle; Cicenas, Jonas; Samalavicius, Narimantas E; Suziedelis, Kestutis

2017-03-18

Numerous lines of evidence support the hierarchical model of cancer development and tumor initiation. According to the theory, cancer stem cells play a crucial role in the formation of the tumor and should be targeted for more effective anticancer treatment. However, cancer stem cells quickly loose their characteristics when propagated as 2D cell culture, indicating that the 2D cell culture does not provide the appropriate settings to maintain an in vivo environment. In this study we have investigated the expression of self-renewal, cancer stem cell and epithelial to mesenchymal transition markers after the transfer of human colorectal carcinoma cell DLD1 and HT29 lines from 2D cell cultures to scaffold-attached laminin rich extracellular matrix and scaffold-free multicellular spheroid 3D culture models. Based on the up-regulated expression of multipotency, CSC and EMT markers, our data suggests that human colorectal carcinoma cells grown in 3D exhibit enhanced cancer stem cell characteristics. Therefore, in order to design more efficient targeted therapies, we suggest that 3D cell culture models should be employed in cancer stem cell research. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Loss of corepressor PER2 under hypoxia up-regulates OCT1-mediated EMT gene expression and enhances tumor malignancy

PubMed Central

Hwang-Verslues, Wendy W.; Chang, Po-Hao; Jeng, Yung-Ming; Kuo, Wen-Hung; Chiang, Pei-Hsun; Chang, Yi-Cheng; Hsieh, Tsung-Han; Su, Fang-Yi; Lin, Liu-Chen; Abbondante, Serena; Yang, Cheng-Yuan; Hsu, Huan-Ming; Yu, Jyh-Cherng; Chang, King-Jen; Shew, Jin-Yuh; Lee, Eva Y.-H. P.; Lee, Wen-Hwa

2013-01-01

The circadian clock gene Period2 (PER2) has been suggested to be a tumor suppressor. However, detailed mechanistic evidence has not been provided to support this hypothesis. We found that loss of PER2 enhanced invasion and activated expression of epithelial-mesenchymal transition (EMT) genes including TWIST1, SLUG, and SNAIL. This finding was corroborated by clinical observation that PER2 down-regulation was associated with poor prognosis in breast cancer patients. We further demonstrated that PER2 served as a transcriptional corepressor, which recruited polycomb proteins EZH2 and SUZ12 as well as HDAC2 to octamer transcription factor 1 (OCT1) (POU2F1) binding sites of the TWIST1 and SLUG promoters to repress expression of these EMT genes. Hypoxia, a condition commonly observed in tumors, caused PER2 degradation and disrupted the PER2 repressor complex, leading to activation of EMT gene expression. This result was further supported by clinical data showing a significant negative correlation between hypoxia and PER2. Thus, our findings clearly demonstrate the tumor suppression function of PER2 and elucidate a pathway by which hypoxia promotes EMT via degradation of PER2. PMID:23836662

Exosomes as mediators of platinum resistance in ovarian cancer.

PubMed

Crow, Jennifer; Atay, Safinur; Banskota, Samagya; Artale, Brittany; Schmitt, Sarah; Godwin, Andrew K

2017-02-14

Exosomes have been implicated in the cell-cell transfer of oncogenic proteins and genetic material. We speculated this may be one mechanism by which an intrinsically platinum-resistant population of epithelial ovarian cancer (EOC) cells imparts its influence on surrounding tumor cells. To explore this possibility we utilized a platinum-sensitive cell line, A2780 and exosomes derived from its resistant subclones, and an unselected, platinum-resistant EOC line, OVCAR10. A2780 cells demonstrate a ~2-fold increase in viability upon treatment with carboplatin when pre-exposed to exosomes from platinum-resistant cells as compared to controls. This coincided with increased epithelial to mesenchymal transition (EMT). DNA sequencing of EOC cell lines revealed previously unreported somatic mutations in the Mothers Against Decapentaplegic Homolog 4 (SMAD4) within platinum-resistant cells. A2780 cells engineered to exogenously express these SMAD4 mutations demonstrate up-regulation of EMT markers following carboplatin treatment, are more resistant to carboplatin, and release exosomes which impart a ~1.7-fold increase in resistance in naive A2780 recipient cells as compared to controls. These studies provide the first evidence that acquired SMAD4 mutations enhance the chemo-resistance profile of EOC and present a novel mechanism in which exchange of tumor-derived exosomes perpetuates an EMT phenotype, leading to the development of subpopulations of platinum-refractory cells.

Smad4 and epithelial-mesenchymal transition proteins in colorectal carcinoma: an immunohistochemical study.

PubMed

Ioannou, M; Kouvaras, E; Papamichali, R; Samara, M; Chiotoglou, I; Koukoulis, G

2018-06-01

Epithelial-mesenchymal transition (EMT) plays an important role in cancer metastasis. During EMT, tumor cells acquire the capacity to migrate and invade the stroma. Activation of the transforming growth factor-b (TGF-b) signaling pathway is of major importance for the initiation of EMT. Smad4, an essential protein of this pathway, is known to complex with multiple transcription factors (e.g. Snail-1, Slug, Twist-1), in various types of cancer, promoting the repression or activation of target genes. The role of Smad4 in colorectal cancer (CRC) is not straightforward so far. In the present study forty eight resected CRC tumor specimens were immunohistochemically examined in order to assess the expression of Smad4 and its association with E-cadherin, Snail-1, Slug, Twist-1 protein expression and with various pathological parameters. Smad4 was found to be positively correlated with Snail-1, Slug and Twist-1 expression (p < 0.001). On the other hand it was negatively correlated with the expression of E-cadherin (p < 0.001). Furthermore, lymphatic invasion could be clearly associated with Smad4 expression, a finding complying with the metastatic ability of EMT cells. In conclusion, Smad4 could be considered as a central component of EMT transition in human colorectal cancer that combines with transcriptional factors to reduce E-cadherin and alter the expression of the epithelial phenotype.

The fibroblast surface markers FAP, anti-fibroblast, and FSP are expressed by cells of epithelial origin and may be altered during epithelial-to-mesenchymal transition.

PubMed

Kahounová, Zuzana; Kurfürstová, Daniela; Bouchal, Jan; Kharaishvili, Gvantsa; Navrátil, Jiří; Remšík, Ján; Šimečková, Šárka; Študent, Vladimír; Kozubík, Alois; Souček, Karel

2017-04-06

The identification of fibroblasts and cancer-associated fibroblasts from human cancer tissue using surface markers is difficult, especially because the markers used currently are usually not expressed solely by fibroblasts, and the identification of fibroblast-specific surface molecules is still under investigation. It was aimed to compare three commercially available antibodies in the detection of different surface epitopes of fibroblasts (anti-fibroblast, fibroblast activation protein α, and fibroblast surface protein). The specificity of their expression, employing fibroblast cell lines and tumor-derived fibroblasts from breast and prostate tissues was investigated. Both the established fibroblast cell line HFF-1 and ex vivo primary fibroblasts isolated from breast and prostate cancer tissues expressed the tested surface markers to different degrees. Surprisingly, those markers were expressed also by permanent cell lines of epithelial origin, both benign and cancer-derived (breast-cell lines MCF 10A, HMLE and prostate-cell lines BPH-1, DU 145, and PC-3). The expression of fibroblast activation protein α increased on the surface of previously described models of epithelial cells undergoing epithelial-to-mesenchymal transition in response to treatment with TGF-β1. To prove the co-expression of the fibroblast markers on cells of epithelial origin, we used freshly dissociated human prostate and breast cancer tissues. The results confirmed the co-expression of anti-fibroblast and fibroblast surface protein on CD31/CD45-negative/EpCAM-positive epithelial cells. In summary, our data support the findings that the tested fibroblast markers are not fibroblast specific and may be expressed also by cells of epithelial origin (e.g., cells undergoing EMT). Therefore, the expression of these markers should be interpreted with caution, and the combination of several epitopes for both positive (anti-fibroblast or fibroblast activation protein α) and negative (EpCAM) identification of fibroblasts from breast and prostate tumor tissues is advised. © 2017 International Society for Advancement of Cytometry. © 2017 International Society for Advancement of Cytometry.

LncRNA-LET inhibits cell viability, migration and EMT while induces apoptosis by up-regulation of TIMP2 in human granulosa-like tumor cell line KGN.

PubMed

Han, Qingfang; Zhang, Wenke; Meng, Jinlai; Ma, Li; Li, Aihua

2018-04-01

Polycystic ovary syndrome (PCOS) is a common endocrine disease characterized by hyperandrogenism, irregular menses, and polycystic ovaries. Several long non-coding RNAs (lncRNAs) are aberrantly expressed in PCOS patients; however, little is known about the effects of the lncRNA-low expression in tumor (lncRNA-LET) on PCOS. We aimed to explore the effects of lncRNA-LET on human granulosa-like tumor cell line, KGN. Expression of lncRNA-LET in normal IOSE80 cells and granulosa cells was determined by qRT-PCR. KGN cell viability, apoptosis and migration were measured by trypan blue exclusion method, flow cytometry assay and wound healing assay, respectively. TGF-β1 was used to induce epithelial-mesenchymal transition (EMT) process. LncRNA-LET expression and mRNA expressions of TIMP2 and EMT-related proteins were measured by qRT-PCR. Western blot analysis was used to measure the protein expression of apoptosis-related proteins, EMT-related proteins, TIMP2, and the proteins in the Wnt/β-catenin and Notch signaling pathways. lncRNA-LET was down-regulated in KGN cells, and its overexpression inhibited cell viability and migration, and promoted apoptosis in KGN cells. Overexpression of lncRNA-LET increased the expression of E-cadherin and decreased the expressions of N-cadherin and vimentin in KGN cells. These effects of lncRNA-LET on KGN cells were reversed by TIMP2 suppression. Overexpression of TIMP2 inhibited cell viability, migration and EMT process, and increased apoptosis by activating the Wnt/β-catenin and Notch pathways. Overexpression of lncRNA-LET inhibits cell viability, migration and EMT process, and increases apoptosis in KGN cells by up-regulating the expression of TIMP2 and activating the Wnt/β-catenin and notch signaling pathways. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

A supercritical-CO2 extract of Ganoderma lucidum spores inhibits cholangiocarcinoma cell migration by reversing the epithelial-mesenchymal transition.

PubMed

Li, Lian; Guo, Hui-Jun; Zhu, Ling-Yan; Zheng, Limin; Liu, Xin

2016-05-15

Ganoderma lucidum (G. lucidum) is an oriental medical mushroom that has been widely used in Asian countries for centuries to prevent and treat different diseases, including cancer. The objective of this study was to investigate the effect of A supercritical-CO2 extract of G. lucidum spores on the transforming growth factor beta 1 (TGF-β1)-induced epithelial-mesenchymal transition (EMT) of cholangiocarcinoma cells. This was an in vitro study with human cholangiocarcinoma TFK-1 cells treated with varying concentrations of G. lucidum. A supercritical-CO2 extract of G. lucidum spores (GLE) was obtained from completely sporoderm-broken germinating G. lucidum spores by supercritical fluid carbon dioxide (SCF-CO2) extraction. GLE pre-incubated with human cholangiocarcinoma TFK-1 cells prior to TGF-β1 treatment (2ng/ml) for 48h. Changes in EMT markers were analyzed by western blotting and immunofluorescence. The formation of F-actin stress fibers was assessed via immunostaining with phalloidin and examined using confocal microscopy. Additionally, the effect of the GLE on TGF-β1-induced migration was investigated by a Boyden chamber assay. TGF-β1-induced reduction in E-cadherin expression was associated with a loss of epithelial morphology and cell-cell contact. Concomitant increases in N-cadherin and Fibronectin were evident in predominantly elongated fibroblast-like cells. The GLE suppressed the TGF-β1-induced morphological changes and the changes in cadherin expression, and also inhibited the formation of F-actin stress fibers, which are a hallmark of EMT. The GLE also inhibited TGF-β1-induced migration of TFK-1 cells. Our findings provide new evidence that GLE suppress cholangiocarcinoma migration in vitro through inhibition of TGF-β1-induced EMT. The GLE may be clinically applied in the prevention and/or treatment of cancer metastasis. Copyright © 2016. Published by Elsevier GmbH.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Seung-Hwan; Kim, Dong-Young; Jing, Feifeng

Developmental endothelial locus-1 (Del-1) is an endogenous anti-inflammatory molecule that is highly expressed in the lung and the brain and limits leukocyte migration to these tissues. We previously reported that the expression of Del-1 is positively regulated by p53 in lung endothelial cells. Although several reports have implicated the altered expression of Del-1 gene in cancer patients, little is known about its role in tumor cells. We here investigated the effect of Del-1 on the features of human lung carcinoma cells. Del-1 mRNA was found to be significantly decreased in the human lung adenocarcinoma cell lines A549 (containing wild typemore » of p53), H1299 (null for p53) and EKVX (mutant p53), compared to in human normal lung epithelial BEAS-2B cells and MRC-5 fibroblasts. The decrease of Del-1 expression was dependent on the p53 activity in the cell lines, but not on the expression of p53. Neither treatment with recombinant human Del-1 protein nor the introduction of adenovirus expressing Del-1 altered the expression of the apoptosis regulators BAX, PUMA and Bcl-2. Unexpectedly, the adenovirus-mediated overexpression of Del-1 gene into the lung carcinoma cell lines promoted proliferation and invasion of the lung carcinoma cells, as revealed by BrdU incorporation and transwell invasion assays, respectively. In addition, overexpression of the Del-1 gene enhanced features of epithelial–mesenchymal transition (EMT), such as increasing vimentin while decreasing E-cadherin in A549 cells, and increases in the level of Slug, an EMT-associated transcription regulator. Our findings demonstrated for the first time that there are deleterious effects of high levels of Del-1 in lung carcinoma cells, and suggest that Del-1 may be used as a diagnostic or prognostic marker for cancer progression, and as a novel therapeutic target for lung carcinoma. - Highlights: • Developmental Endothelial Locus-1 (Del-1) expression is downregulated in human lung cancer cells. • Overexpression of the Del-1 gene potentiates proliferation and invasion of lung carcinoma cells. • Del-1 may be used as a diagnostic or prognostic marker for lung cancer progression.« less

The Significance of Epithelial-to-Mesenchymal Transition for Circulating Tumor Cells

PubMed Central

Kölbl, Alexandra C.; Jeschke, Udo; Andergassen, Ulrich

2016-01-01

Epithelial to mesenchymal transition (EMT) is a process involved in embryonic development, but it also plays a role in remote metastasis formation in tumor diseases. During this process cells lose their epithelial features and adopt characteristics of mesenchymal cells. Thereby single tumor cells, which dissolve from the primary tumor, are enabled to invade the blood vessels and travel throughout the body as so called “circulating tumor cells” (CTCs). After leaving the blood stream the reverse process of EMT, the mesenchymal to epithelial transition (MET) helps the cells to seed in different tissues, thereby generating the bud of metastasis formation. As metastasis is the main reason for tumor-associated death, CTCs and the EMT process are in the focus of research in recent years. This review summarizes what was already found out about the molecular mechanisms driving EMT, the consequences of EMT for tumor cell detection, and suitable markers for the detection of CTCs which underwent EMT. The research work done in this field could open new roads towards combating cancer. PMID:27529216

Chronic inorganic arsenic exposure in vitro induces a cancer cell phenotype in human peripheral lung epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Person, Rachel J.; Olive Ngalame, Ntube N.; Makia, Ngome L.

Inorganic arsenic is a human lung carcinogen. We studied the ability of chronic inorganic arsenic (2 μM; as sodium arsenite) exposure to induce a cancer phenotype in the immortalized, non-tumorigenic human lung peripheral epithelial cell line, HPL-1D. After 38 weeks of continuous arsenic exposure, secreted matrix metalloproteinase-2 (MMP2) activity increased to over 200% of control, levels linked to arsenic-induced cancer phenotypes in other cell lines. The invasive capacity of these chronic arsenic-treated lung epithelial (CATLE) cells increased to 320% of control and colony formation increased to 280% of control. CATLE cells showed enhanced proliferation in serum-free media indicative of autonomousmore » growth. Compared to control cells, CATLE cells showed reduced protein expression of the tumor suppressor gene PTEN (decreased to 26% of control) and the putative tumor suppressor gene SLC38A3 (14% of control). Morphological evidence of epithelial-to-mesenchymal transition (EMT) occurred in CATLE cells together with appropriate changes in expression of the EMT markers vimentin (VIM; increased to 300% of control) and e-cadherin (CDH1; decreased to 16% of control). EMT is common in carcinogenic transformation of epithelial cells. CATLE cells showed increased KRAS (291%), ERK1/2 (274%), phosphorylated ERK (p-ERK; 152%), and phosphorylated AKT1 (p-AKT1; 170%) protein expression. Increased transcript expression of metallothioneins, MT1A and MT2A and the stress response genes HMOX1 (690%) and HIF1A (247%) occurred in CATLE cells possibly in adaptation to chronic arsenic exposure. Thus, arsenic induced multiple cancer cell characteristics in human peripheral lung epithelial cells. This model may be useful to assess mechanisms of arsenic-induced lung cancer. - Highlights: • Chronic arsenic exposure transforms a human peripheral lung epithelia cell line. • Cells acquire characteristics in common with human lung adenocarcinoma cells. • These transformed cells provide a valuable model for arsenic-induced lung cancer.« less

Fibulin-3 negatively regulates ALDH1 via c-MET suppression and increases γ-radiation-induced sensitivity in some pancreatic cancer cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, In-Gyu, E-mail: igkim@kaeri.re.kr; Department of Radiation Biotechnology and Applied Radioisotope, Korea University of Science and Technology; Lee, Jae-Ha

2014-11-21

Highlights: • FBLN-3 gene was poorly expressed in some pancreatic cancer lines. • FBLN-3 promoter region was highly methylated in some pancreatic cancer cell lines. • FBLN-3 inhibited c-MET activation and expression and reduced cellular level of ALDH1. • FBLN-3/c-Met/ALDH1 axis modulates stemness and EMT in pancreatic cancer cells. - Abstract: Fibulin-3 (FBLN-3) has been postulated to be either a tumor suppressor or promoter depending on the cell type, and hypermethylation of the FBLN-3 promoter is often associated with human disease, especially cancer. We report that the promoter region of the FBLN-3 was significantly methylated (>95%) in some pancreatic cancermore » cell lines and thus FBLN-3 was poorly expressed in pancreatic cancer cell lines such as AsPC-1 and MiaPaCa-2. FBLN-3 overexpression significantly down-regulated the cellular level of c-MET and inhibited hepatocyte growth factor-induced c-MET activation, which were closely associated with γ-radiation resistance of cancer cells. Moreover, we also showed that c-MET suppression or inactivation decreased the cellular level of ALDH1 isozymes (ALDH1A1 or ALDH1A3), which serve as cancer stem cell markers, and subsequently induced inhibition of cell growth in pancreatic cancer cells. Therefore, forced overexpression of FBLN-3 sensitized cells to cytotoxic agents such as γ-radiation and strongly inhibited the stemness and epithelial to mesenchymal transition (EMT) property of pancreatic cancer cells. On the other hand, if FBLN3 was suppressed in FBLN-3-expressing BxPC3 cells, the results were opposite. This study provides the first demonstration that the FBLN-3/c-MET/ALDH1 axis in pancreatic cancer cells partially modulates stemness and EMT as well as sensitization of cells to the detrimental effects of γ-radiation.« less

HAND2 Target Gene Regulatory Networks Control Atrioventricular Canal and Cardiac Valve Development.

PubMed

Laurent, Frédéric; Girdziusaite, Ausra; Gamart, Julie; Barozzi, Iros; Osterwalder, Marco; Akiyama, Jennifer A; Lincoln, Joy; Lopez-Rios, Javier; Visel, Axel; Zuniga, Aimée; Zeller, Rolf

2017-05-23

The HAND2 transcriptional regulator controls cardiac development, and we uncover additional essential functions in the endothelial to mesenchymal transition (EMT) underlying cardiac cushion development in the atrioventricular canal (AVC). In Hand2-deficient mouse embryos, the EMT underlying AVC cardiac cushion formation is disrupted, and we combined ChIP-seq of embryonic hearts with transcriptome analysis of wild-type and mutants AVCs to identify the functionally relevant HAND2 target genes. The HAND2 target gene regulatory network (GRN) includes most genes with known functions in EMT processes and AVC cardiac cushion formation. One of these is Snai1, an EMT master regulator whose expression is lost from Hand2-deficient AVCs. Re-expression of Snai1 in mutant AVC explants partially restores this EMT and mesenchymal cell migration. Furthermore, the HAND2-interacting enhancers in the Snai1 genomic landscape are active in embryonic hearts and other Snai1-expressing tissues. These results show that HAND2 directly regulates the molecular cascades initiating AVC cardiac valve development. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

HAND2 Target Gene Regulatory Networks Control Atrioventricular Canal and Cardiac Valve Development

DOE PAGES

Laurent, Frédéric; Girdziusaite, Ausra; Gamart, Julie; ...

2017-05-23

The HAND2 transcriptional regulator controls cardiac development, and we uncover additional essential functions in the endothelial to mesenchymal transition (EMT) underlying cardiac cushion development in the atrioventricular canal (AVC). In Hand2-deficient mouse embryos, the EMT underlying AVC cardiac cushion formation is disrupted, and we combined ChIP-seq of embryonic hearts with transcriptome analysis of wild-type and mutants AVCs to identify the functionally relevant HAND2 target genes. The HAND2 target gene regulatory network (GRN) includes most genes with known functions in EMT processes and AVC cardiac cushion formation. One of these is Snai1, an EMT master regulator whose expression is lost frommore » Hand2-deficient AVCs. Re-expression of Snai1 in mutant AVC explants partially restores this EMT and mesenchymal cell migration. Furthermore, the HAND2-interacting enhancers in the Snai1 genomic landscape are active in embryonic hearts and other Snai1-expressing tissues. These results show that HAND2 directly regulates the molecular cascades initiating AVC cardiac valve development.« less

HAND2 Target Gene Regulatory Networks Control Atrioventricular Canal and Cardiac Valve Development

DOE Office of Scientific and Technical Information (OSTI.GOV)

Laurent, Frédéric; Girdziusaite, Ausra; Gamart, Julie

The HAND2 transcriptional regulator controls cardiac development, and we uncover additional essential functions in the endothelial to mesenchymal transition (EMT) underlying cardiac cushion development in the atrioventricular canal (AVC). In Hand2-deficient mouse embryos, the EMT underlying AVC cardiac cushion formation is disrupted, and we combined ChIP-seq of embryonic hearts with transcriptome analysis of wild-type and mutants AVCs to identify the functionally relevant HAND2 target genes. The HAND2 target gene regulatory network (GRN) includes most genes with known functions in EMT processes and AVC cardiac cushion formation. One of these is Snai1, an EMT master regulator whose expression is lost frommore » Hand2-deficient AVCs. Re-expression of Snai1 in mutant AVC explants partially restores this EMT and mesenchymal cell migration. Furthermore, the HAND2-interacting enhancers in the Snai1 genomic landscape are active in embryonic hearts and other Snai1-expressing tissues. These results show that HAND2 directly regulates the molecular cascades initiating AVC cardiac valve development.« less

Surprisal analysis characterizes the free energy time course of cancer cells undergoing epithelial-to-mesenchymal transition.

PubMed

Zadran, Sohila; Arumugam, Rameshkumar; Herschman, Harvey; Phelps, Michael E; Levine, R D

2014-09-09

The epithelial-to-mesenchymal transition (EMT) initiates the invasive and metastatic behavior of many epithelial cancers. Mechanisms underlying EMT are not fully known. Surprisal analysis of mRNA time course data from lung and pancreatic cancer cells stimulated to undergo TGF-β1-induced EMT identifies two phenotypes. Examination of the time course for these phenotypes reveals that EMT reprogramming is a multistep process characterized by initiation, maturation, and stabilization stages that correlate with changes in cell metabolism. Surprisal analysis characterizes the free energy time course of the expression levels throughout the transition in terms of two state variables. The landscape of the free energy changes during the EMT for the lung cancer cells shows a stable intermediate state. Existing data suggest this is the previously proposed maturation stage. Using a single-cell ATP assay, we demonstrate that the TGF-β1-induced EMT for lung cancer cells, particularly during the maturation stage, coincides with a metabolic shift resulting in increased cytosolic ATP levels. Surprisal analysis also characterizes the absolute expression levels of the mRNAs and thereby examines the homeostasis of the transcription system during EMT.

Knockdown of BAG3 induces epithelial-mesenchymal transition in thyroid cancer cells through ZEB1 activation.

PubMed

Meng, X; Kong, D-H; Li, N; Zong, Z-H; Liu, B-Q; Du, Z-X; Guan, Y; Cao, L; Wang, H-Q

2014-02-27

The process by which epithelial features are lost in favor of a mesenchymal phenotype is referred to as epithelial-mesenchymal transition (EMT). Most carcinomas use this mechanism to evade into neighboring tissues. Reduction or a loss of E-cadherin expression is a well-established hallmark of EMT. As a potent suppressor of E-cadherin, transcription factor ZEB1 is one of the key inducers of EMT, whose expression promotes tumorigenesis and metastasis of carcinomas. Bcl-2-associated athanogene 3 (BAG3) affects multifaceted cellular functions, including proliferation, apoptosis, cell adhesion and invasion, viral infection, and autophagy. Recently, we have reported a novel role of BAG3 implicated in EMT, while the mechanisms are poorly elucidated. The current study demonstrated that knockdown of BAG3 induced EMT, and increased cell migratory and invasiveness in thyroid cancer cells via transcriptional activation of ZEB1. We also found that BAG3 knockdown led to nuclear accumulation of β-catenin, which was responsible for the transcriptional activation of ZEB1. These results indicate BAG3 as a regulator of ZEB1 expression in EMT and as a regulator of metastasis in thyroid cancer cells, providing potential targets to prevent and/or treat thyroid cancer cell invasion and metastasis.

Knockdown of BAG3 induces epithelial–mesenchymal transition in thyroid cancer cells through ZEB1 activation

PubMed Central

Meng, X; Kong, D-H; Li, N; Zong, Z-H; Liu, B-Q; Du, Z-X; Guan, Y; Cao, L; Wang, H-Q

2014-01-01

The process by which epithelial features are lost in favor of a mesenchymal phenotype is referred to as epithelial–mesenchymal transition (EMT). Most carcinomas use this mechanism to evade into neighboring tissues. Reduction or a loss of E-cadherin expression is a well-established hallmark of EMT. As a potent suppressor of E-cadherin, transcription factor ZEB1 is one of the key inducers of EMT, whose expression promotes tumorigenesis and metastasis of carcinomas. Bcl-2-associated athanogene 3 (BAG3) affects multifaceted cellular functions, including proliferation, apoptosis, cell adhesion and invasion, viral infection, and autophagy. Recently, we have reported a novel role of BAG3 implicated in EMT, while the mechanisms are poorly elucidated. The current study demonstrated that knockdown of BAG3 induced EMT, and increased cell migratory and invasiveness in thyroid cancer cells via transcriptional activation of ZEB1. We also found that BAG3 knockdown led to nuclear accumulation of β-catenin, which was responsible for the transcriptional activation of ZEB1. These results indicate BAG3 as a regulator of ZEB1 expression in EMT and as a regulator of metastasis in thyroid cancer cells, providing potential targets to prevent and/or treat thyroid cancer cell invasion and metastasis. PMID:24577090

MicroRNA-34a targets epithelial to mesenchymal transition-inducing transcription factors (EMT-TFs) and inhibits breast cancer cell migration and invasion

PubMed Central

Fu, Shangyi; Yang, Luquan; Tania, Mousumi; Zhang, Xianqin; Xiao, Xiuli; Zhang, Xianning; Fu, Junjiang

2017-01-01

MicroRNA-34a (miR-34a) plays an essential role against tumorigenesis and progression of cancer metastasis. Here, we analyzed the expression, targets and functional effects of miR-34a on epithelial to mesenchymal transition-inducing transcription factors (EMT-TFs), such as TWIST1, SLUG and ZEB1/2, and an EMT-inducing protein NOTCH1 in breast cancer (BC) cell migration and invasion and its correlation with tumorigenesis and clinical outcomes. Expression of miR-34a is downregulated in human metastatic breast cancers (MBC) compared to normal breast tissues and is negatively correlated with clinicopathological features of MBC patients. Ectopic expression of miR-34a in MBC cell-line BT-549 significantly inhibits cell migration and invasion, but exhibits no clear effect on BC cell growth. We found that miR-34a is able to inactivate EMT signaling pathway with mediatory of NOTCH1, TWIST1, and ZEB1 upon 3′-UTR activity in MBC cell lines, but has no inhibitory effects on SLUG and ZEB2. Furthermore, we investigated the synergistic effects of Thymoquinone (TQ) and miR-34a together on the expression of EMT-associated proteins. Results showed that co-delivery of miR-34a and TQ is able to inactivate EMT signaling pathway by directly targeting TWIST1 and ZEB1 in BT-549 cell line, indicating that they might be a promising therapeutic combination against breast cancer metastasis. Epigenetic inactivation of the EMT-TFs/miR-34a pathway can potentially alter the equilibrium of these regulations, facilitating EMT and metastasis in BC. Altogether, our findings suggest that miR-34a alone could serve as a potential therapeutic agent for MBC, and together with TQ, their therapeutic potential is synergistically enhanced. PMID:28423483

MicroRNA-34a targets epithelial to mesenchymal transition-inducing transcription factors (EMT-TFs) and inhibits breast cancer cell migration and invasion.

PubMed

Imani, Saber; Wei, Chunli; Cheng, Jingliang; Khan, Md Asaduzzaman; Fu, Shangyi; Yang, Luquan; Tania, Mousumi; Zhang, Xianqin; Xiao, Xiuli; Zhang, Xianning; Fu, Junjiang

2017-03-28

MicroRNA-34a (miR-34a) plays an essential role against tumorigenesis and progression of cancer metastasis. Here, we analyzed the expression, targets and functional effects of miR-34a on epithelial to mesenchymal transition-inducing transcription factors (EMT-TFs), such as TWIST1, SLUG and ZEB1/2, and an EMT-inducing protein NOTCH1 in breast cancer (BC) cell migration and invasion and its correlation with tumorigenesis and clinical outcomes. Expression of miR-34a is downregulated in human metastatic breast cancers (MBC) compared to normal breast tissues and is negatively correlated with clinicopathological features of MBC patients. Ectopic expression of miR-34a in MBC cell-line BT-549 significantly inhibits cell migration and invasion, but exhibits no clear effect on BC cell growth. We found that miR-34a is able to inactivate EMT signaling pathway with mediatory of NOTCH1, TWIST1, and ZEB1 upon 3'-UTR activity in MBC cell lines, but has no inhibitory effects on SLUG and ZEB2. Furthermore, we investigated the synergistic effects of Thymoquinone (TQ) and miR-34a together on the expression of EMT-associated proteins. Results showed that co-delivery of miR-34a and TQ is able to inactivate EMT signaling pathway by directly targeting TWIST1 and ZEB1 in BT-549 cell line, indicating that they might be a promising therapeutic combination against breast cancer metastasis. Epigenetic inactivation of the EMT-TFs/miR-34a pathway can potentially alter the equilibrium of these regulations, facilitating EMT and metastasis in BC. Altogether, our findings suggest that miR-34a alone could serve as a potential therapeutic agent for MBC, and together with TQ, their therapeutic potential is synergistically enhanced.

Twist promotes tumor metastasis in basal-like breast cancer by transcriptionally upregulating ROR1.

PubMed

Cao, Jingying; Wang, Xin; Dai, Tao; Wu, Yuanzhong; Zhang, Meifang; Cao, Renxian; Zhang, Ruhua; Wang, Gang; Jiang, Rou; Zhou, Binhua P; Shi, Jian; Kang, Tiebang

2018-01-01

Rationale: Twist is a key transcription factor for induction of epithelial-mesenchymal transition (EMT), which promotes cell migration, invasion, and cancer metastasis, confers cancer cells with stem cell-like characteristics, and provides therapeutic resistance. However, the functional roles and targeted genes of Twist in EMT and cancer progression remain elusive. Methods: The potential targeted genes of Twist were identified from the global transcriptomes of T47D/Twist cells by microarray analysis. EMT phenotype was detected by western blotting and immunofluorescence of marker proteins. The dual-luciferase reporter and chromatin immunoprecipitation assays were employed to observe the direct transcriptional induction of ROR1 by Twist. A lung metastasis model was used to study the pro-metastatic role of Twist and ROR1 by injecting MDA-MB-231 cells into tail vein of nude mice. Bio-informatics analysis was utilized to measure the metastasis-free survival of breast cancer patients. Results: Twist protein was proved to directly activate the transcription of ROR1 gene, a receptor of Wnt5a in non-canonical WNT signaling pathway. Silencing of ROR1 inhibited EMT process, cell migration, invasion, and cancer metastasis of basal-like breast cancer (BLBC) cells. Knockdown of ROR1 also ameliorated the pro-metastatic effect of Twist. Furthermore, analyses of clinical specimens indicated that high expression of both ROR1 and Twist tightly correlates with poor metastasis-free survival of breast cancer patients. Conclusion: ROR1 is a targeted gene of Twist. Twist/ROR1 signaling is critical for invasion and metastasis of BLBC cells.

E-cadherin regulators are differentially expressed in the epithelium and stroma of keratocystic odontogenic tumors.

PubMed

Porto, Lia Pontes Arruda; dos Santos, Jean Nunes; Ramalho, Luciana Maria Pedreira; Figueiredo, Andreia Leal; Carneiro Júnior, Bráulio; Gurgel, Clarissa Araújo; Paiva, Katiúcia Batista Silva; Xavier, Flávia Caló Aquino

2016-04-01

The epithelial-mesenchymal transition (EMT) is the process where cells lose their epithelial features and acquire properties of typical mesenchymal cells. The dissociation of tumor cells due to changes in cell-cell adhesion is one of the key principles of tumor invasion and EMT. Thus, the knowledge of the molecular features of EMT in keratocyst odontogenic tumor (KOT) can provide useful markers to aid in the diagnosis and prognosis and perhaps contribute to an alternative therapeutic approach as it shows an aggressive clinical behavior and high recurrence rates. This study aimed to evaluate the EMT in KOT by the immunoexpression of E-cadherin, N-cadherin, Snail, and Slug and comparing to radicular cysts and dental follicles. Thirty-two KOTs, 15 radicular cysts, and 08 dental follicles were used for immunohistochemistry, evaluating the extent, intensity, labeling pattern, cellular compartment in the epithelium and stroma, and the presence of inflammation. E-cadherin was preserved in most cases of keratocystic odontogenic tumor. N-cadherin was increased in the tumor epithelium, a result that was positively correlated with the heterogeneous and nuclear immunoexpression of Slug in the epithelium; Slug also correlated with high Snail immunoexpression. N-cadherin was positively correlated with Slug in the stroma of keratocystic odontogenic tumors. The high immunoexpression of Snail and nuclear Slug in keratocystic odontogenic tumors suggests these proteins as transcription factors without necessarily participating in 'cadherin switching'. However, the knowledge of their induction of the epithelial-mesenchymal transition in odontogenic tumors is still limited. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Marker-less multi-frame motion tracking and compensation in PET-brain imaging

NASA Astrophysics Data System (ADS)

Lindsay, C.; Mukherjee, J. M.; Johnson, K.; Olivier, P.; Song, X.; Shao, L.; King, M. A.

2015-03-01

In PET brain imaging, patient motion can contribute significantly to the degradation of image quality potentially leading to diagnostic and therapeutic problems. To mitigate the image artifacts resulting from patient motion, motion must be detected and tracked then provided to a motion correction algorithm. Existing techniques to track patient motion fall into one of two categories: 1) image-derived approaches and 2) external motion tracking (EMT). Typical EMT requires patients to have markers in a known pattern on a rigid too attached to their head, which are then tracked by expensive and bulky motion tracking camera systems or stereo cameras. This has made marker-based EMT unattractive for routine clinical application. Our main contributions are the development of a marker-less motion tracking system that uses lowcost, small depth-sensing cameras which can be installed in the bore of the imaging system. Our motion tracking system does not require anything to be attached to the patient and can track the rigid transformation (6-degrees of freedom) of the patient's head at a rate 60 Hz. We show that our method can not only be used in with Multi-frame Acquisition (MAF) PET motion correction, but precise timing can be employed to determine only the necessary frames needed for correction. This can speeds up reconstruction by eliminating the unnecessary subdivision of frames.

Epithelial to mesenchymal transition in arsenic-transformed cells promotes angiogenesis through activating β-catenin–vascular endothelial growth factor pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Zhishan; Humphries, Brock; Xiao, Hua

2013-08-15

Arsenic exposure represents a major health concern increasing cancer risks, yet the mechanism of arsenic carcinogenesis has not been elucidated. We and others recently reported that cell malignant transformation by arsenic is accompanied by epithelial to mesenchymal transition (EMT). However, the role of EMT in arsenic carcinogenesis is not well understood. Although previous studies showed that short term exposure of endothelial cells to arsenic stimulated angiogenesis, it remains to be determined whether cells that were malignantly transformed by long term arsenic exposure have a pro-angiogenic effect. The objective of this study was to investigate the effect of arsenic-transformed human bronchialmore » epithelial cells that underwent EMT on angiogenesis and the underlying mechanism. It was found that the conditioned medium from arsenic-transformed cells strongly stimulated tube formation by human umbilical vein endothelial cells (HUVECs). Moreover, enhanced angiogenesis was detected in mouse xenograft tumor tissues resulting from inoculation of arsenic-transformed cells. Mechanistic studies revealed that β-catenin was activated in arsenic-transformed cells up-regulating its target gene expression including angiogenic-stimulating vascular endothelial growth factor (VEGF). Stably expressing microRNA-200b in arsenic-transformed cells that reversed EMT inhibited β-catenin activation, decreased VEGF expression and reduced tube formation by HUVECs. SiRNA knockdown β-catenin decreased VEGF expression. Adding a VEGF neutralizing antibody into the conditioned medium from arsenic-transformed cells impaired tube formation by HUVECs. Reverse transcriptase-PCR analysis revealed that the mRNA levels of canonical Wnt ligands were not increased in arsenic-transformed cells. These findings suggest that EMT in arsenic-transformed cells promotes angiogenesis through activating β-catenin–VEGF pathway. - Highlights: • Arsenic-transformed cells that underwent EMT displayed a pro-angiogenic effect. • EMT in arsenic-transformed cells activates β-catenin. • β-Catenin activation increases VEGF expression in arsenic-transformed cells. • β-Catenin activation is likely independent of canonical Wnt signaling. • EMT in arsenic-transformed cells promotes angiogenesis via β-catenin–VEGF pathway.« less

Aldolase B Overexpression is Associated with Poor Prognosis and Promotes Tumor Progression by Epithelial-Mesenchymal Transition in Colorectal Adenocarcinoma.

PubMed

Li, Qingguo; Li, Yaqi; Xu, Junyan; Wang, Sheng; Xu, Ye; Li, Xinxiang; Cai, Sanjun

2017-01-01

Glycolysis is considered to be the root of cancer development and progression, which involved a multi-step enzymatic reaction. Our study aimed at figuring out which glycolysis enzyme participates in the development of colorectal cancer and its possible mechanisms. We firstly screened out Aldolase B (ALDOB) by performing qRT-PCR arrays of glycolysis-related genes in five paired liver metastasis and primary colorectal tissues, and further detected ALDOB protein with immunohistochemistry in tissue microarray (TMA) consisting of 229 samples from stage I-III colorectal cancer patients. CRISPR-Cas9 method was adopted to create knock out colon cancer cell lines (LoVo and SW480) of ALDOB. The effect of ALDOB on cell proliferation and metastasis was examined in vitro using colony formation assay as well as transwell migration and invasion assay, respectively. In TMA, there was 64.6% of samples demonstrated strong intensity of ALDOB. High ALDOB expression were associated with poor overall survival and disease-free survival in both univariate and multivariate regression analyses (P<0.05). In vitro functional studies of CCK-8 demonstrated that silencing ALDOB expression significantly (P<0.05) inhibited proliferation, migration and invasion of colon cancer cells. Mechanically, silencing ALDOB activated epithelial markers and repressed mesenchymal markers, indicating inactivation of ALDOB may lead to inhibition of epithelial-mesenchymal transition (EMT). Upregulation of ALDOB promotes colorectal cancer metastasis by facilitating EMT and acts as a potential prognostic factor and therapeutic target in colorectal cancer. © 2017 The Author(s). Published by S. Karger AG, Basel.

Enhanced epithelial to mesenchymal transition (EMT) and upregulated MYC in ectopic lesions contribute independently to endometriosis.

PubMed

Proestling, Katharina; Birner, Peter; Gamperl, Susanne; Nirtl, Nadine; Marton, Erika; Yerlikaya, Gülen; Wenzl, Rene; Streubel, Berthold; Husslein, Heinrich

2015-07-22

Epithelial to mesenchymal transition (EMT) is a process in which epithelial cells lose polarity and cell-to-cell contacts and acquire the migratory and invasive abilities of mesenchymal cells. These abilities are thought to be prerequisites for the establishment of endometriotic lesions. A hallmark of EMT is the functional loss of E-cadherin (CDH1) expression in epithelial cells. TWIST1, a transcription factor that represses E-cadherin transcription, is among the EMT inducers. SNAIL, a zinc-finger transcription factor, and its close relative SLUG have similar properties to TWIST1 and are thus also EMT inducers. MYC, which is upregulated by estrogens in the uterus by an estrogen response cis-acting element (ERE) in its promoter, is associated with proliferation in endometriosis. The role of EMT and proliferation in the pathogenesis of endometriosis was evaluated by analyzing TWIST1, CDH1 and MYC expression. CDH1, TWIST1, SNAIL and SLUG mRNA expression was analyzed by qRT-PCR from 47 controls and 74 patients with endometriosis. Approximately 42 ectopic and 62 eutopic endometrial tissues, of which 30 were matched samples, were collected during the same surgical procedure. We evaluated TWIST1 and MYC protein expression by immunohistochemistry (IHC) in the epithelial and stromal tissue of 69 eutopic and 90 ectopic endometrium samples, of which 49 matched samples were analyzed from the same patient. Concordant expression of TWIST1/SNAIL/SLUG and CDH1 but also of TWIST1 and MYC was analyzed. We found that TWIST1, SNAIL and SLUG are overexpressed (p < 0.001, p = 0.016 and p < 0.001) in endometriosis, while CDH1 expression was concordantly reduced in these samples (p < 0.001). Similar to TWIST1, the epithelial expression of MYC was also significantly enhanced in ectopic endometrium compared to eutopic tissues (p = 0.008). We found exclusive expression of either TWIST1 or MYC in the same samples (p = 0.003). Epithelial TWIST1 is overexpressed in endometriosis and may contribute to the formation of endometriotic lesions by inducing epithelial to mesenchymal transition, as CDH1 was reduced in ectopic lesions. We found exclusive expression of either TWIST1 or MYC in the same samples, indicating that EMT and proliferation contribute independently of each other to the formation of endometriotic lesions.

Mechanisms of nitric oxide-mediated inhibition of EMT in cancer

PubMed Central

Baritaki, Stavroula; Huerta-Yepez, Sara; Sahakyan, Anna; Karagiannides, Iordanis; Bakirtzi, Kyriaki; Jazirehi, Ali R

2010-01-01

The role of nitric oxide (NO) in cancer has been controversial and is based on the levels of NO and the responsiveness of the tumor type. It remains unclear whether NO can inhibit the epithelial to mesenchymal transition (EMT) in cancer cells. EMT induction is mediated, in part, by the constitutive activation of the metastasis-inducer transcription factor, Snail and EMT can be inhibited by the metastasis-suppressors Raf-1 kinase inhibitor protein (RKIP) and E-cadherin. Snail is transcriptionally regulated by NFκB and in turn, Snail represses RKIP transcription. Hence, we hypothesized that high levels of NO, that inhibit NFκB activity, may also inhibit Snail, induce RKIP and leading to inhibition of EMT. We show that treatment of human prostate metastatic cell lines with the NO donor, DETANONOate, inhibits EMT and reverses both the mesenchymal phenotype and the cell invasive properties. Further, treatment with DETANONOate inhibits Snail expression and DNA-binding activity in parallel with the upregulation of RKIP and E-cadherin protein levels. The pivotal roles of Snail inhibition and RKIP induction in DETANONOate-mediated inhibition of EMT were corroborated by both Snail silencing by siRNA and by ectopic expression of RKIP. The in vitro findings were validated in vivo in mice bearing PC-3 xenografts treated with DETANONOate. The present findings show, for the first time, the novel role of high, yet, subtoxic concentrations of NO in the inhibition of EMT. Thus, NO donors may exert therapeutic activities in the reversal of EMT and metastasis. PMID:21150329

Phenotypic Plasticity Determines Cancer Stem Cell Therapeutic Resistance in Oral Squamous Cell Carcinoma.

PubMed

Biddle, Adrian; Gammon, Luke; Liang, Xiao; Costea, Daniela Elena; Mackenzie, Ian C

2016-02-01

Cancer stem cells (CSCs) drive tumour spread and therapeutic resistance, and can undergo epithelial-to-mesenchymal transition (EMT) and mesenchymal-to-epithelial transition (MET) to switch between epithelial and post-EMT sub-populations. Examining oral squamous cell carcinoma (OSCC), we now show that increased phenotypic plasticity, the ability to undergo EMT/MET, underlies increased CSC therapeutic resistance within both the epithelial and post-EMT sub-populations. The post-EMT CSCs that possess plasticity exhibit particularly enhanced therapeutic resistance and are defined by a CD44(high)EpCAM(low/-) CD24(+) cell surface marker profile. Treatment with TGFβ and retinoic acid (RA) enabled enrichment of this sub-population for therapeutic testing, through which the endoplasmic reticulum (ER) stressor and autophagy inhibitor Thapsigargin was shown to selectively target these cells. Demonstration of the link between phenotypic plasticity and therapeutic resistance, and development of an in vitro method for enrichment of a highly resistant CSC sub-population, provides an opportunity for the development of improved chemotherapeutic agents that can eliminate CSCs.

EGFR Signal-Network Reconstruction Demonstrates Metabolic Crosstalk in EMT

PubMed Central

Choudhary, Kumari Sonal; Rohatgi, Neha; Briem, Eirikur; Gudjonsson, Thorarinn; Gudmundsson, Steinn; Rolfsson, Ottar

2016-01-01

Epithelial to mesenchymal transition (EMT) is an important event during development and cancer metastasis. There is limited understanding of the metabolic alterations that give rise to and take place during EMT. Dysregulation of signalling pathways that impact metabolism, including epidermal growth factor receptor (EGFR), are however a hallmark of EMT and metastasis. In this study, we report the investigation into EGFR signalling and metabolic crosstalk of EMT through constraint-based modelling and analysis of the breast epithelial EMT cell model D492 and its mesenchymal counterpart D492M. We built an EGFR signalling network for EMT based on stoichiometric coefficients and constrained the network with gene expression data to build epithelial (EGFR_E) and mesenchymal (EGFR_M) networks. Metabolic alterations arising from differential expression of EGFR genes was derived from a literature review of AKT regulated metabolic genes. Signaling flux differences between EGFR_E and EGFR_M models subsequently allowed metabolism in D492 and D492M cells to be assessed. Higher flux within AKT pathway in the D492 cells compared to D492M suggested higher glycolytic activity in D492 that we confirmed experimentally through measurements of glucose uptake and lactate secretion rates. The signaling genes from the AKT, RAS/MAPK and CaM pathways were predicted to revert D492M to D492 phenotype. Follow-up analysis of EGFR signaling metabolic crosstalk in three additional breast epithelial cell lines highlighted variability in in vitro cell models of EMT. This study shows that the metabolic phenotype may be predicted by in silico analyses of gene expression data of EGFR signaling genes, but this phenomenon is cell-specific and does not follow a simple trend. PMID:27253373

EGFR Signal-Network Reconstruction Demonstrates Metabolic Crosstalk in EMT.

PubMed

Choudhary, Kumari Sonal; Rohatgi, Neha; Halldorsson, Skarphedinn; Briem, Eirikur; Gudjonsson, Thorarinn; Gudmundsson, Steinn; Rolfsson, Ottar

2016-06-01

Epithelial to mesenchymal transition (EMT) is an important event during development and cancer metastasis. There is limited understanding of the metabolic alterations that give rise to and take place during EMT. Dysregulation of signalling pathways that impact metabolism, including epidermal growth factor receptor (EGFR), are however a hallmark of EMT and metastasis. In this study, we report the investigation into EGFR signalling and metabolic crosstalk of EMT through constraint-based modelling and analysis of the breast epithelial EMT cell model D492 and its mesenchymal counterpart D492M. We built an EGFR signalling network for EMT based on stoichiometric coefficients and constrained the network with gene expression data to build epithelial (EGFR_E) and mesenchymal (EGFR_M) networks. Metabolic alterations arising from differential expression of EGFR genes was derived from a literature review of AKT regulated metabolic genes. Signaling flux differences between EGFR_E and EGFR_M models subsequently allowed metabolism in D492 and D492M cells to be assessed. Higher flux within AKT pathway in the D492 cells compared to D492M suggested higher glycolytic activity in D492 that we confirmed experimentally through measurements of glucose uptake and lactate secretion rates. The signaling genes from the AKT, RAS/MAPK and CaM pathways were predicted to revert D492M to D492 phenotype. Follow-up analysis of EGFR signaling metabolic crosstalk in three additional breast epithelial cell lines highlighted variability in in vitro cell models of EMT. This study shows that the metabolic phenotype may be predicted by in silico analyses of gene expression data of EGFR signaling genes, but this phenomenon is cell-specific and does not follow a simple trend.

Notch3 inhibits epithelial–mesenchymal transition by activating Kibra-mediated Hippo/YAP signaling in breast cancer epithelial cells

PubMed Central

Zhang, X; Liu, X; Luo, J; Xiao, W; Ye, X; Chen, M; Li, Y; Zhang, G-J

2016-01-01

Invasion, metastasis and chemoresistance are leading causes of death in breast cancer patients. A vital change of epithelial cells, epithelial–mesenchymal transition (EMT), is involved in these processes. Unfortunately, the molecular mechanisms controlling EMT remain to be elucidated. Our previous studies have shown that ectopic N3ICD expression inhibits EMT in MDA-MB-231, a triple-negative breast cancer (TNBC) epithelial cell line. To decipher the mechanism, we performed in-depth studies. Specifically, we found that overexpressing N3ICD transcriptionally upregulated the expression of Kibra, an upstream member of the Hippo pathway. Correspondingly, we also observed that phosphorylated Hippo pathway core kinases, including Lats1/2 and MST1/2, were increased and decreased by overexpressing and knocking down Notch3, respectively. Furthermore, we found that the oncogenic transcriptional coactivator yes-associated protein (YAP), which is negatively regulated by the Hippo pathway, was inhibited by overexpressing N3ICD in breast cancer epithelial cells. The ability of Kibra to inhibit EMT has been previously reported. We thus speculated that Notch3 inhibition of EMT is mediated by upregulated Kibra. To verify this hypothesis, a rescue experiment was performed. Evidently, the ability of Notch3 to inhibit EMT can be countered by knocking down Kibra expression. These data suggest that Notch3 inhibits EMT by activating the Hippo/YAP pathway by upregulating Kibra in breast cancer epithelial cells, and Kibra may be a downstream effector of Notch3. These findings deepen our understanding of EMT in both development and disease, and will undoubtedly help to provide new therapeutic strategies for interfering with cancer invasion and metastasis, especially for TNBC. PMID:27841855

Slug is upregulated during wound healing and regulates cellular phenotypes in corneal epithelial cells.

PubMed

Aomatsu, Keiichi; Arao, Tokuzo; Abe, Kosuke; Kodama, Aya; Sugioka, Koji; Matsumoto, Kazuko; Kudo, Kanae; Kimura, Hideharu; Fujita, Yoshihiko; Hayashi, Hidetoshi; Nagai, Tomoyuki; Shimomura, Yoshikazu; Nishio, Kazuto

2012-02-16

The involvement of the epithelial mesenchymal transition (EMT) in the process of corneal wound healing remains largely unclear. The purpose of the present study was to gain insight into Slug expression and corneal wound healing. Slug expression during wound healing in the murine cornea was evaluated using fluorescence staining in vivo. Slug or Snail was stably introduced into human corneal epithelial cells (HCECs). These stable transfectants were evaluated for the induction of the EMT, cellular growth, migration activity, and expression changes in differentiation-related molecules. Slug, but not Snail, was clearly expressed in the nuclei of corneal epithelial cells in basal lesion of the corneal epithelium during wound healing in vivo. The overexpression of Slug or Snail induced an EMT-like cellular morphology and cadherin switching in HCECs, indicating that these transcription factors were able to mediate the typical EMT in HCECs. The overexpression of Slug or Snail suppressed cellular proliferation but enhanced the migration activity. Furthermore, ABCG2, TP63, and keratin 19, which are known as stemness-related molecules, were downregulated in these transfectants. It was found that Slug is upregulated during corneal wound healing in vivo. The overexpression of Slug mediated a change in the cellular phenotype affecting proliferation, migration, and expression levels of differentiation-related molecules. This is the first evidence that Slug is regulated during the process of corneal wound healing in the corneal epithelium in vivo, providing a novel insight into the EMT and Slug expression in corneal wound healing.

Cross Talk Mechanism among EMT, ROS, and Histone Acetylation in Phorbol Ester-Treated Human Breast Cancer MCF-7 Cells.

PubMed

Kamiya, Tetsuro; Goto, Aki; Kurokawa, Eri; Hara, Hirokazu; Adachi, Tetsuo

2016-01-01

Epithelial-mesenchymal transition (EMT) plays a pivotal role in the progression of cancer, and some transcription factors including Slug and Snail are known to be involved in EMT processes. It has been well established that the excess production of reactive oxygen species (ROS) and epigenetics such as DNA methylation and histone modifications participate in carcinogenesis; however, the cross talk mechanism among EMT, ROS, and epigenetics remains unclear. In the present study, we demonstrated that the treatment of human breast cancer MCF-7 cells with phorbol ester (TPA), a protein kinase C activator, significantly induced cell proliferation and migration, and these were accompanied by the significant induction of Slug expression. Moreover, the TPA-elicited induction of Slug expression was regulated by histone H3 acetylation and NADPH oxidase (NOX) 2-derived ROS signaling, indicating that ROS and histone acetylation are involved in TPA-elicited EMT processes. We herein determined the cross talk mechanism among EMT, ROS, and histone acetylation, and our results provide an insight into the progression of cancer metastasis.

Metastasis is regulated via microRNA-200/ZEB1 axis control of tumour cell PD-L1 expression and intratumoral immunosuppression.

PubMed

Chen, Limo; Gibbons, Don L; Goswami, Sangeeta; Cortez, Maria Angelica; Ahn, Young-Ho; Byers, Lauren A; Zhang, Xuejun; Yi, Xiaohui; Dwyer, David; Lin, Wei; Diao, Lixia; Wang, Jing; Roybal, Jonathon; Patel, Mayuri; Ungewiss, Christin; Peng, David; Antonia, Scott; Mediavilla-Varela, Melanie; Robertson, Gordon; Suraokar, Milind; Welsh, James W; Erez, Baruch; Wistuba, Ignacio I; Chen, Lieping; Peng, Di; Wang, Shanshan; Ullrich, Stephen E; Heymach, John V; Kurie, Jonathan M; Qin, F Xiao-Feng

2014-10-28

Immunosuppression of tumour-infiltrating lymphocytes (TIL) is a common feature of advanced cancer, but its biological basis has remained obscure. We demonstrate here a molecular link between epithelial-to-mesenchymal transition (EMT) and CD8(+) TIL immunosuppression, two key drivers of cancer progression. We show that microRNA-200 (miR-200), a cell-autonomous suppressor of EMT and metastasis, targets PD-L1. Moreover, ZEB1, an EMT activator and transcriptional repressor of miR-200, relieves miR-200 repression of PD-L1 on tumour cells, leading to CD8(+) T-cell immunosuppression and metastasis. These findings are supported by robust correlations between the EMT score, miR-200 levels and PD-L1 expression in multiple human lung cancer datasets. In addition to revealing a link between EMT and T-cell dysfunction, these findings also show that ZEB1 promotes metastasis through a heretofore unappreciated cell non-autonomous mechanism, and suggest that subgroups of patients in whom malignant progression is driven by EMT activators may respond to treatment with PD-L1 antagonists.

Enterolactone modulates the ERK/NF-κB/Snail signaling pathway in triple-negative breast cancer cell line MDA-MB-231 to revert the TGF-β-induced epithelial-mesenchymal transition.

PubMed

Mali, Aniket V; Joshi, Asavari A; Hegde, Mahabaleshwar V; Kadam, Shivajirao S

2018-05-01

Triple-negative breast cancer (TNBC) is highly metastatic, and there is an urgent unmet need to develop novel therapeutic strategies leading to the new drug discoveries against metastasis. The transforming growth factor-β (TGF-β) is known to promote the invasive and migratory potential of breast cancer cells through induction of epithelial-mesenchymal transition (EMT) via the ERK/NF-κB/Snail signaling pathway, leading to breast cancer metastasis. Targeting this pathway to revert the EMT would be an attractive, novel therapeutic strategy to halt breast cancer metastasis. Effects of enterolactone (EL) on the cell cycle and apoptosis were investigated using flow cytometry and a cleaved caspase-3 enzyme-linked immunosorbent assay (ELISA), respectively. Effects of TGF-β induction and EL treatment on the functional malignancy of MDA-MB-231 breast cancer cells were investigated using migration and chemo-invasion assays. The effects of EL on EMT markers and the ERK/NF-κB/Snail signaling pathway after TGF-β induction were studied using confocal microscopy, quantitative reverse transcription polymerase chain reaction (qRT-PCR), Western blot, and flow cytometry. Herein, we report that EL exhibits a significant antimetastatic effect on MDA-MB-231 cells by almost reverting the TGF-β-induced EMT in vitro . EL downregulates the mesenchymal markers N-cadherin and vimentin, and upregulates the epithelial markers E-cadherin and occludin. It represses actin stress fiber formation via inhibition of mitogen-activated protein kinase p-38 (MAPK-p38) and cluster of differentiation 44 (CD44). EL also suppresses ERK-1/2, NF-κB, and Snail at the mRNA and protein levels. Briefly, EL was found to inhibit TGF-β-induced EMT by blocking the ERK/NF-κB/Snail signaling pathway, which is a promising target for breast cancer metastasis therapy.

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

PubMed Central

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 own CAFs via EMT in this model. PMID:25658113

High expression of SALL4 and fascin, and loss of E-cadherin expression in undifferentiated/dedifferentiated carcinomas of the endometrium

PubMed Central

Onder, Semen; Taskin, Orhun Cig; Sen, Fatma; Topuz, Samet; Kucucuk, Seden; Sozen, Hamdullah; Ilhan, Ridvan; Tuzlali, Sitki; Yavuz, Ekrem

2017-01-01

Abstract Undifferentiated/dedifferentiated endometrial carcinomas (UCE/DCEs) of the endometrium are rare tumors with poor prognosis. There are few clinicopathologic studies with detailed immunohistochemical analysis regarding UCE/DCEs. We evaluated the diagnostic value of a selected tumor stem-cell marker and epithelial-mesenchymal transition (EMT) markers, in addition to previously studied markers in identifying UCE/DCEs from other types of high-grade endometrial carcinomas. Eleven cases of UCE/DCEs with complete clinical follow-up that were diagnosed between 2006 and 2015 were included in the study. For immunohistochemical comparison, 11 clinically matched cases for each type of other high-grade endometrial carcinomas (high-grade endometrioid (F3-EC), serous [SC], and clear cell carcinoma [CCC]) were used as a control group. An immunohistochemical analysis including fascin, SALL4, E-cadherin, and β-catenin, in addition to epithelial and neuroendocrine markers was performed in each case. The majority of UCE/DCEs displayed diffuse expression of fascin (81.9%) and loss of E-cadherin expression (54.5%). SALL4 expression was detected in 36.3% of the UCE/DCE cases. SALL4 expression was significantly more frequent in UCE/DCEs than all other high-grade carcinomas (P < 0.001). Loss of E-cadherin and fascin expression was significantly more frequent in UCE/DCEs than high-grade endometrioid and clear cell adenocarcinomas (P = 0.012, 0.014 and P = 0.01, 0.003, respectively). We suggest that loss of E-cadherin expression together with fascin and SALL4 immunopositivity in addition to morphologic features have an impact in differential diagnosis of UCE/DCEs from other high-grade endometrial carcinomas. PMID:28272224

MicroRNA-218 inhibits EMT, migration and invasion by targeting SFMBT1 and DCUN1D1 in cervical cancer

PubMed Central

Li, Jing; Li, Jingyu; Lin, Xiaochun; Chen, Xing; Zhang, Jiren; Zheng, Yanfang

2016-01-01

Repeated infection with high-risk HPV is a major cause for the development and metastasis of human cervical cancer, even though the mechanism of the metastasis is still not completely understood. Here, we reported that miR-218 (microRNA-218) was downregulated in cervical cancer tissues, especially in metastatic cancer tissues. We found that miR-218 expression was associated with clinicopathological characteristics of patients with cervical cancer. MiR-218 overexpression inhibited Epithelial-Mesenchymal Transition (EMT), migration and invasiveness of cervical cancer cells in vitro. Moreover, miR-218 repressed the expression of SFMFBT1 (Scm-like with four MBT domains 1) and DCUN1D1 (defective in cullin neddylation 1, domain containing 1) by direct binding to the 3′UTRs of the mRNAs. The overexpression of SFMBT1 induced EMT and increased the migration and invasiveness of cervical cancer cells, while the overexpression of DCUN1D1 increased the migration and invasiveness of these cells, but did not induce EMT. An inverse correlation was observed between the expression of miR-218 and DCUN1D1 protein in cervical cancer tissues. Importantly, HPV16 E6 downregulated the expression of miR-218 in cervical cancer, while miR-218 rescued the promotion effect of HPV16 E6 on the expression of SFMBT1 and DCUN1D1. Taken together, our results revealed that HPV16 E6 promoted EMT and invasion in cervical cancer via the repression of miR-218, while miR-218 inhibited EMT and invasion in cervical cancer by targeting SFMBT1 and DCUN1D1. PMID:27285984

Tumor-associated macrophages induce the expression of FOXQ1 to promote epithelial-mesenchymal transition and metastasis in gastric cancer cells.

PubMed

Guo, Jian; Yan, Yan; Yan, Yu; Guo, Qinyue; Zhang, Mingxin; Zhang, Jia; Goltzman, David

2017-10-01

Gastric cancer (GC) is one of the most common malignancies, and is the second leading cause of cancer-related deaths worldwide. Macrophages infiltrated in the tumor microenvironment (TME) called tumor-associated macrophages (TAMs) are key orchestrators in TME. In GC, it has been reported that infiltration of TAMs is associated with epithelial-mesenchymal transition (EMT)-related proteins in human GC tissues, but the exactly mechanism has not been clarified. In the present study, we aimed to elucidate the underlying mechanism of TAMs on GC cells. THP-1 cells were used to investigate the effects of TAMs on GC cells. The effects of invasion and migration induced by coculture with TAMs were investigated by Transwell invasion and wound healing assays. The expression of EMT-related genes and forkhead box Q1 (FOXQ1) were examined in MKN45 and MKN74 cells after being co-cultured with TAMs. The density of TAMs and the expression of FOXQ1 were analyzed by immunohistochemistry in GC tissues. Our results revealed that, co-culture with TAMs promoted the invasion and migration of GC cells. Co-culture with TAMs induced EMT in GC cells. FOXQ1 is essential for TAM-induced EMT and metastasis in GC cells. Furthermore, silencing of FOXQ1 blocked the effect of TAM-enhanced EMT and metastasis of GC cells. High expression of CD68 was correlated with positive FOXQ1 expression (r=0.613; P<0.001) in clinical GC samples. Our data provided evidence that TAMs promote EMT, invasion and migration of GC cells via FOXQ1. Therefore, the TAM/FOXQ1 axis may represent a novel target for GC cells.

MUC1 enhances invasiveness of pancreatic cancer cells by inducing epithelial to mesenchymal transition

PubMed Central

Roy, Lopamudra Das; Sahraei, Mahnaz; Subramani, Durai B.; Besmer, Dahlia; Nath, Sritama; Tinder, Teresa L.; Bajaj, Ekta; Shanmugam, Kandavel; Lee, Yong Yook; Hwang, Sun IL; Gendler, Sandra J.; Mukherjee, Pinku

2010-01-01

Increased motility and invasiveness of pancreatic cancer cells are associated with epithelial to mesenchymal transition (EMT). Snai1 and Slug are zinc-finger transcription factors that trigger this process by repressing E-cadherin and enhancing vimentin and N-Cadherin protein expression. However, the mechanisms that regulate this activation in pancreatic tumors remain elusive. MUC1, a transmembrane mucin glycoprotein, is associated with the most invasive forms of pancreatic adenocarcinomas (PDA). In this study, we show that over expression of MUC1 in pancreatic cancer cells triggers the molecular process of EMT which translates to increased invasiveness and metastasis. EMT was significantly reduced when Muc1 was genetically deleted in a mouse model of PDA or when all seven tyrosines in the cytoplasmic tail of MUC1 were mutated to phenylalanine (mutated MUC1 CT). Using proteomics, RT-PCR, and Western blotting, we revealed a significant increase in vimentin, Slug and Snail expression with repression of E-Cadherin in MUC1-expressing cells compared to cells expressing the mutated MUC1 CT. In the cells that carried the mutated MUC1 CT, MUC1 failed to co-immunoprecipitate with β-catenin and translocate to the nucleus thereby blocking transcription of the genes associated with EMT and metastasis. Thus, functional tyrosines are critical in stimulating the interactions between MUC1 and β-catenin and their nuclear translocation to initiate the process of EMT. This study signifies the oncogenic role of MUC1 CT and is the first to identify a direct role of the MUC1 in initiating EMT during pancreatic cancer. The data may have implications in future design of MUC1-targeted therapies for pancreatic cancer. PMID:21102519

MUC1 enhances invasiveness of pancreatic cancer cells by inducing epithelial to mesenchymal transition.

PubMed

Roy, L D; Sahraei, M; Subramani, D B; Besmer, D; Nath, S; Tinder, T L; Bajaj, E; Shanmugam, K; Lee, Y Y; Hwang, S I L; Gendler, S J; Mukherjee, P

2011-03-24

Increased motility and invasiveness of pancreatic cancer cells are associated with epithelial to mesenchymal transition (EMT). Snai1 and Slug are zinc-finger transcription factors that trigger this process by repressing E-cadherin and enhancing vimentin and N-cadherin protein expression. However, the mechanisms that regulate this activation in pancreatic tumors remain elusive. MUC1, a transmembrane mucin glycoprotein, is associated with the most invasive forms of pancreatic ductal adenocarcinomas (PDA). In this study, we show that over expression of MUC1 in pancreatic cancer cells triggers the molecular process of EMT, which translates to increased invasiveness and metastasis. EMT was significantly reduced when MUC1 was genetically deleted in a mouse model of PDA or when all seven tyrosines in the cytoplasmic tail of MUC1 were mutated to phenylalanine (mutated MUC1 CT). Using proteomics, RT-PCR and western blotting, we revealed a significant increase in vimentin, Slug and Snail expression with repression of E-Cadherin in MUC1-expressing cells compared with cells expressing the mutated MUC1 CT. In the cells that carried the mutated MUC1 CT, MUC1 failed to co-immunoprecipitate with β-catenin and translocate to the nucleus, thereby blocking transcription of the genes associated with EMT and metastasis. Thus, functional tyrosines are critical in stimulating the interactions between MUC1 and β-catenin and their nuclear translocation to initiate the process of EMT. This study signifies the oncogenic role of MUC1 CT and is the first to identify a direct role of the MUC1 in initiating EMT during pancreatic cancer. The data may have implications in future design of MUC1-targeted therapies for pancreatic cancer.

Bleomycin-induced epithelial–mesenchymal transition in sclerotic skin of mice: Possible role of oxidative stress in the pathogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhou, Cheng-Fan, E-mail: zhouchengfan@sohu.com; Department of Occupational Health and Environmental Health, School of Public Health, Anhui Medical University, Hefei, Anhui 230032; Zhou, Deng-Chuan

Epithelial–mesenchymal transition (EMT) derived myofibroblasts are partly responsible for the increased collagen synthesis and deposition that occur in tissue fibrosis; however EMT occurrence in skin fibrosis and its mechanism remain unknown. The aim of this study was to investigate whether epithelial cells undergo EMT and determine the role of oxidative stress in this process. BALB/c mice were subcutaneously injected with bleomycin (BLM) or phosphate buffer saline (PBS) into the shaved back daily for 2, 3, and 4 weeks. Skin collagen deposition was evaluated by histopathology and Western blotting. EMT characteristics in the skin were determined by histopathology and immunofluorescent stainingmore » for E-cadherin and vimentin, which were further evaluated by Western blotting and reverse transcriptase polymerase chain reaction (RT-PCR). To investigate the role of oxidative stress in EMT, the antioxidant N-acetylcysteine (NAC) was intraperitoneally (100 mg/kg body weight/day) injected daily for 3 weeks. The epithelial suprabasal cells were detached from the basement membrane zone (BMZ) in the sclerotic skin treated with BLM. Immunofluorescent staining indicated vimentin-positive epithelial cells frequently occurring in the thickened epidermis of BLM-treated mice. Western blotting and RT-PCR showed that the expression of E-cadherin was significantly decreased but that of vimentin significantly increased in the skin treated with BLM. NAC attenuated BLM induced oxidative damage, changes in E-cadherin and vimentin expressions and collagen deposition in the sclerotic skin of mice. This study provides the first evidence that BLM induces the EMT of the epithelial cells superficial to the basement membrane zone in the skin fibrosis. Oxidative stress may contribute, at least in part, to BLM induced EMT and skin fibrosis in mice. - Highlights: • We provided the first evidence that EMT occurred in BLM-induced skin fibrosis. • Epithelial cells superficial to the BMZ underwent EMT in BLM-induced skin fibrosis. • NAC attenuated EMT induction in the process of BLM-induced skin fibrosis. • NAC didn't significantly inhibit TGF-β expression in BLM-induced skin fibrosis.« less

Epithelial-mesenchymal transition and tumor suppression are controlled by a reciprocal feedback loop between ZEB1 and Grainyhead-like-2

PubMed Central

Cieply, Benjamin; Farris, Joshua; Denvir, James; Ford, Heide; Frisch, Steven M.

2013-01-01

Epithelial-mesenchymal transition (EMT) in carcinoma cells enhances malignant progression by promoting invasion and survival. EMT is induced by microenvironmental factors including TGF-β and Wnt agonists, and by the E-box-binding transcription factors Twist, Snail and ZEB. Grainyhead-like-2 (GRHL2), a member of the mammalian Grainyhead family of wound healing regulatory transcription factors, suppresses EMT and restores sensitivity to anoikis by repressing ZEB1 expression and inhibiting TGF-β signaling. In this study, we elucidate the functional relationship between GRHL2 and ZEB1 in EMT/MET and tumor biology. At least three homeodomain proteins, Six1, LBX1, and HoxA5, transactivated the ZEB1 promoter, in the case of Six1, through direct protein-promoter interaction. GRHL2 altered the Six1-DNA complex, inhibiting this transactivation. Correspondingly, GRHL2 expression prevented tumor initiation in xenograft assays, sensitized breast cancer cells to paclitaxel and suppressed the emergence of CD44highCD24low cells (defining the cancer stem cell phenotype in the cell type studied). GRHL2 was down-regulated in recurrent mouse tumors that had evolved to an oncogene-independent, EMT-like state, supporting a role for GRHL2 down-regulation in this phenotypic transition, modeling disease recurrence. The combination of TGF-β and Wnt activation repressed GRHL2 expression by direct interaction of ZEB1 with the GRHL2 promoter, inducing EMT. Together, our observations indicate that a reciprocal feedback loop between GRHL2 and ZEB1 controls epithelial vs. mesenchymal phenotypes and EMT-driven tumor progression. PMID:23943797

Reciprocal regulation of the cholinic phenotype and epithelial-mesenchymal transition in glioblastoma cells

PubMed Central

Koch, Katharina; Hartmann, Rudolf; Schröter, Friederike; Suwala, Abigail Kora; Maciaczyk, Donata; Krüger, Andrea Caroline; Willbold, Dieter; Kahlert, Ulf Dietrich; Maciaczyk, Jaroslaw

2016-01-01

Glioblastoma (GBM) is the most malignant brain tumor with very limited therapeutic options. Standard multimodal treatments, including surgical resection and combined radio-chemotherapy do not target the most aggressive subtype of glioma cells, brain tumor stem cells (BTSCs). BTSCs are thought to be responsible for tumor initiation, progression, and relapse. Furthermore, they have been associated with the expression of mesenchymal features as a result of epithelial-mesenchymal transition (EMT) thereby inducing tumor dissemination and chemo resistance. Using high resolution proton nuclear magnetic resonance spectroscopy (1H NMR) on GBM cell cultures we provide evidence that the expression of well-known EMT activators of the ZEB, TWIST and SNAI families and EMT target genes N-cadherin and VIMENTIN is associated with aberrant choline metabolism. The cholinic phenotype is characterized by high intracellular levels of phosphocholine and total choline derivatives and was associated with malignancy in various cancers. Both genetic and pharmacological inhibition of the cardinal choline metabolism regulator choline kinase alpha (CHKα) significantly reduces the cell viability, invasiveness, clonogenicity, and expression of EMT associated genes in GBM cells. Moreover, in some cell lines synergetic cytotoxic effects were observed when combining the standard of care chemotherapeutic temozolomide with the CHKα inhibitor V-11-0711. Taken together, specific inhibition of the enzymatic activity of CHKα is a powerful strategy to suppress EMT which opens the possibility to target chemo-resistant BTSCs through impairing their mesenchymal transdifferentiation. Moreover, the newly identified EMT-oncometabolic network may be helpful to monitor the invasive properties of glioblastomas and the success of anti-EMT therapy. PMID:27705917

Mechanisms of nitric oxide-mediated inhibition of EMT in cancer: inhibition of the metastasis-inducer Snail and induction of the metastasis-suppressor RKIP.

PubMed

Baritaki, Stavroula; Huerta-Yepez, Sara; Sahakyan, Anna; Karagiannides, Iordanis; Bakirtzi, Kyriaki; Jazirehi, Ali; Bonavida, Benjamin

2010-12-15

The role of nitric oxide (NO) in cancer has been controversial and is based on the levels of NO and the responsiveness of the tumor type. It remains unclear whether NO can inhibit the epithelial to mesenchymal transition (EMT) in cancer cells. EMT induction is mediated, in part, by the constitutive activation of the metastasis-inducer transcription factor, Snail and EMT can be inhibited by the metastasis-suppressor Raf-1 kinase inhibitor protein (RKIP) and E-cadherin. Snail is transcriptionally regulated by NF-κB and in turn, Snail represses RKIP transcription. Hence, we hypothesized that high levels of NO, that inhibit NF-κB activity, may also inhibit Snail and induce RKIP and leading to inhibition of EMT. We show that treatment of human prostate metastatic cell lines with the NO donor, DETANONOate, inhibits EMT and reverses both the mesenchymal phenotype and the cell invasive properties. Further, treatment with DETANONOate inhibits Snail expression and DNA-binding activity in parallel with the upregulation of RKIP and E-cadherin protein levels. The pivotal roles of Snail inhibition and RKIP induction in DETANONOate-mediated inhibition of EMT were corroborated by both Snail silencing by siRNA and by ectopic expression of RKIP. The in vitro findings were validated in vivo in mice bearing PC-3 xenografts and treated with DETANONOate. The present findings show, for the first time, the novel role of high subtoxic concentrations of NO in the inhibition of EMT. Thus, NO donors may exert therapeutic activities in the reversal of EMT and metastasis.

The Nuclear Receptor, RORγ, Regulates Pathways Necessary for Breast Cancer Metastasis

PubMed Central

Oh, Tae Gyu; Wang, Shu-Ching M.; Acharya, Bipul R.; Goode, Joel M.; Graham, J. Dinny; Clarke, Christine L.; Yap, Alpha S.; Muscat, George E.O.

2016-01-01

We have previously reported that RORγ expression was decreased in ER − ve breast cancer, and increased expression improves clinical outcomes. However, the underlying RORγ dependent mechanisms that repress breast carcinogenesis have not been elucidated. Here we report that RORγ negatively regulates the oncogenic TGF-β/EMT and mammary stem cell (MaSC) pathways, whereas RORγ positively regulates DNA-repair. We demonstrate that RORγ expression is: (i) decreased in basal-like subtype cancers, and (ii) inversely correlated with histological grade and drivers of carcinogenesis in breast cancer cohorts. Furthermore, integration of RNA-seq and ChIP-chip data reveals that RORγ regulates the expression of many genes involved in TGF-β/EMT-signaling, DNA-repair and MaSC pathways (including the non-coding RNA, LINC00511). In accordance, pharmacological studies demonstrate that an RORγ agonist suppresses breast cancer cell viability, migration, the EMT transition (microsphere outgrowth) and mammosphere-growth. In contrast, RNA-seq demonstrates an RORγ inverse agonist induces TGF-β/EMT-signaling. These findings suggest pharmacological modulation of RORγ activity may have utility in breast cancer. PMID:27211549

Contextual extracellular cues promote tumor cell EMT and metastasis by regulating miR-200 family expression

PubMed Central

Gibbons, Don L.; Lin, Wei; Creighton, Chad J.; Rizvi, Zain H.; Gregory, Philip A.; Goodall, Gregory J.; Thilaganathan, Nishan; Du, Liqin; Zhang, Yiqun; Pertsemlidis, Alexander; Kurie, Jonathan M.

2009-01-01

Metastatic disease is a primary cause of cancer-related death, and factors governing tumor cell metastasis have not been fully elucidated. Here, we address this question by using tumor cell lines derived from mice that develop metastatic lung adenocarcinoma owing to expression of mutant K-ras and p53. Despite having widespread somatic genetic alterations, the metastasis-prone tumor cells retained a marked plasticity. They transited reversibly between epithelial and mesenchymal states, forming highly polarized epithelial spheres in three-dimensional culture that underwent epithelial-to-mesenchymal transition (EMT) following treatment with transforming growth factor-β or injection into syngeneic mice. This transition was entirely dependent on the microRNA (miR)-200 family, which decreased during EMT. Forced expression of miR-200 abrogated the capacity of these tumor cells to undergo EMT, invade, and metastasize, and conferred transcriptional features of metastasis-incompetent tumor cells. We conclude that tumor cell metastasis is regulated by miR-200 expression, which changes in response to contextual extracellular cues. PMID:19759262

The metastasis-associated gene MTA3, a component of the Mi-2/NuRD transcriptional repression complex, predicts prognosis of gastroesophageal junction adenocarcinoma.

PubMed

Dong, Hongmei; Guo, Hong; Xie, Liangxi; Wang, Geng; Zhong, Xueyun; Khoury, Thaer; Tan, Dongfeng; Zhang, Hao

2013-01-01

Gastroesophageal junction (GEJ) adenocarcinoma carries a poor prognosis that is largely attributable to early and frequent metastasis. The acquisition of metastatic potential in cancer involves epithelial-to-mesenchymal transition (EMT). The metastasis-associated gene MTA3, a novel component of the Mi-2/NuRD transcriptional repression complex, was identified as master regulator of EMT through inhibition of Snail to increase E-cadherin expression in breast cancer. Here, we evaluated the expression pattern of the components of MTA3 pathway and the corresponding prognostic significance in GEJ adenocarcinoma. MTA3 expression was decreased at both protein and mRNA levels in tumor tissues compared to the non-tumorous and lowed MTA3 levels were noted in tumor cell lines with stronger metastatic potential. Immunohistochemical analysis of a cohort of 128 cases exhibited that patients with lower expression of MTA3 had poorer outcomes. Combined misexpression of MTA3, Snail and E-cadherin had stronger correlation with malignant properties. Collectively, results suggest that the MTA3-regulated EMT pathway is altered to favor EMT and, therefore, disease progression and that MTA3 expression was an independent prognostic factor in patients with GEJ adenocarcinoma.

An in vivo model of epithelial to mesenchymal transition reveals a mitogenic switch

PubMed Central

Jahn, Stephan C.; Law, Mary E.; Corsino, Patrick E.; Parker, Nicole N.; Pham, Kien; Davis, Bradley J.; Lu, Jianrong; Law, Brian K.

2012-01-01

The epithelial to mesenchymal transition (EMT) is a process by which differentiated epithelial cells transition to a mesenchymal phenotype. EMT enables the escape of epithelial cells from the rigid structural constraints of the tissue architecture to a phenotype more amenable to cell migration and, therefore, invasion and metastasis. We characterized an in vivo model of EMT and discovered that marked changes in mitogenic signaling occurred during this process. DNA microarray analysis revealed that the expression of a number of genes varied significantly between post-EMT and pre-EMT breast cancer cells. Post-EMT cancer cells upregulated mRNA encoding c-Met and the PDGF and LPA receptors, and acquired increased responsiveness to HGF, PDGF, and LPA. This rendered the post-EMT cells responsive to the growth inhibitory effects of HGF, PDGF, and LPA receptor inhibitors/antagonists. Furthermore, post- EMT cells exhibited decreased basal Raf and Erk phosphorylation, and in comparison to pre-EMT cells, their proliferation was poorly inhibited by a MEK inhibitor. These studies suggest that therapies need to be designed to target both pre-EMT and post-EMT cancer cells and that signaling changes in post- EMT cells may allow them to take advantage of paracrine signaling from the stroma in vivo. PMID:22906417

Exosomes as mediators of platinum resistance in ovarian cancer

PubMed Central

Crow, Jennifer; Atay, Safinur; Banskota, Samagya; Artale, Brittany; Schmitt, Sarah; Godwin, Andrew K

2017-01-01

Exosomes have been implicated in the cell-cell transfer of oncogenic proteins and genetic material. We speculated this may be one mechanism by which an intrinsically platinum-resistant population of epithelial ovarian cancer (EOC) cells imparts its influence on surrounding tumor cells. To explore this possibility we utilized a platinum-sensitive cell line, A2780 and exosomes derived from its resistant subclones, and an unselected, platinum-resistant EOC line, OVCAR10. A2780 cells demonstrate a ~2-fold increase in viability upon treatment with carboplatin when pre-exposed to exosomes from platinum-resistant cells as compared to controls. This coincided with increased epithelial to mesenchymal transition (EMT). DNA sequencing of EOC cell lines revealed previously unreported somatic mutations in the Mothers Against Decapentaplegic Homolog 4 (SMAD4) within platinum-resistant cells. A2780 cells engineered to exogenously express these SMAD4 mutations demonstrate up-regulation of EMT markers following carboplatin treatment, are more resistant to carboplatin, and release exosomes which impart a ~1.7-fold increase in resistance in naive A2780 recipient cells as compared to controls. These studies provide the first evidence that acquired SMAD4 mutations enhance the chemo-resistance profile of EOC and present a novel mechanism in which exchange of tumor-derived exosomes perpetuates an EMT phenotype, leading to the development of subpopulations of platinum-refractory cells. PMID:28060758

Connective tissue growth factor is a positive regulator of epithelial-mesenchymal transition and promotes the adhesion with gastric cancer cells in human peritoneal mesothelial cells.

PubMed

Jiang, Cheng-Gang; Lv, Ling; Liu, Fu-Rong; Wang, Zhen-Ning; Na, Di; Li, Feng; Li, Jia-Bin; Sun, Zhe; Xu, Hui-Mian

2013-01-01

Connective tissue growth factor (CTGF) is involved in human cancer development and progression. Epithelial to mesenchymal transition (EMT) plays an important role in many biological processes. In this study, we wished to investigate the role of CTGF in EMT of peritoneal mesothelial cells and the effects of CTGF on adhesion of gastric cancer cells to mesothelial cells. Human peritoneal mesothelial cells (HPMCs) were cultured with TGF-β1 or various concentrations of CTGF for different time. The EMT process was monitored by morphology. Real-time RT-PCR and Western blot were used to evaluate the expression of vimentin, α-SMA , E-cadherin and β-catenin. RNA interference was used to achieve selective and specific knockdown of CTGF. We demonstrated that CTGF induced EMT of mesothelial cells in a dose- and time-dependent manner. HPMCs were exposed to TGF-β1 also underwent EMT which was associated with the induction of CTGF expression. Transfection with CTGF siRNA was able to reverse the EMT partially after treatment of TGF-β1. Moreover, the induced EMT of HPMCs was associated with an increased adhesion of gastric cancer cells to mesothelial cells. These findings suggest that CTGF is not only an important mediator but a potent activator of EMT in peritoneal mesothelial cells, which in turn promotes gastric cancer cell adhesion to peritoneum. Copyright © 2012 Elsevier Ltd. All rights reserved.

Elevation of adenylate energy charge by angiopoietin-like 4 enhances epithelial-mesenchymal transition by inducing 14-3-3γ expression.

PubMed

Teo, Z; Sng, M K; Chan, J S K; Lim, M M K; Li, Y; Li, L; Phua, T; Lee, J Y H; Tan, Z W; Zhu, P; Tan, N S

2017-11-16

Metastatic cancer cells acquire energy-intensive processes including increased invasiveness and chemoresistance. However, how the energy demand is met and the molecular drivers that coordinate an increase in cellular metabolic activity to drive epithelial-mesenchymal transition (EMT), the first step of metastasis, remain unclear. Using different in vitro and in vivo EMT models with clinical patient's samples, we showed that EMT is an energy-demanding process fueled by glucose metabolism-derived adenosine triphosphate (ATP). We identified angiopoietin-like 4 (ANGPTL4) as a key player that coordinates an increase in cellular energy flux crucial for EMT via an ANGPTL4/14-3-3γ signaling axis. This augmented cellular metabolic activity enhanced metastasis. ANGPTL4 knockdown suppresses an adenylate energy charge elevation, delaying EMT. Using an in vivo dual-inducible EMT model, we found that ANGPTL4 deficiency reduces cancer metastasis to the lung and liver. Unbiased kinase inhibitor screens and Ingenuity Pathway Analysis revealed that ANGPTL4 regulates the expression of 14-3-3γ adaptor protein via the phosphatidylinositol-3-kinase/AKT and mitogen-activated protein kinase signaling pathways that culminate to activation of transcription factors, CREB, cFOS and STAT3. Using a different mode of action, as compared with protein kinases, the ANGPTL4/14-3-3γ signaling axis consolidated cellular bioenergetics and stabilized critical EMT proteins to coordinate energy demand and enhanced EMT competency and metastasis, through interaction with specific phosphorylation signals on target proteins.

Metabolic reprogramming during TGFβ1-induced epithelial-to-mesenchymal transition

PubMed Central

Jiang, Lei; Xiao, Ling; Sugiura, Hidekazu; Huang, Xiumei; Ali, Aktar; Kuro-o, Makoto; Deberardinis, Ralph J.; Boothman, David A.

2014-01-01

Metastatic progression, including extravasation and micro-metastatic outgrowth, is the main cause of cancer patient death. Recent studies suggest that cancer cells reprogram their metabolism to support increased proliferation through increased glycolysis and biosynthetic activities, including lipogenesis pathways. However, metabolic changes during metastatic progression, including alterations in regulatory gene expression, remain undefined. We show that transforming growth factor beta 1 (TGFβ1) induced Epithelial-to-Mesenchymal Transition (EMT) is accompanied by coordinately reduced enzyme expression required to convert glucose into fatty acids, and concomitant enhanced respiration. Over-expressed Snail1, a transcription factor mediating TGFβ1-induced EMT, was sufficient to suppress carbohydrate-responsive-element-binding protein (ChREBP, a master lipogenic regulator), and fatty acid synthase (FASN), its effector lipogenic gene. Stable FASN knock-down was sufficient to induce EMT, stimulate migration and extravasation in vitro. FASN silencing enhanced lung metastasis and death in vivo. These data suggest that a metabolic transition that suppresses lipogenesis and favors energy production is an essential component of TGFβ1-induced EMT and metastasis. PMID:25284588

Baicalin ameliorates renal fibrosis via inhibition of transforming growth factor β1 production and downstream signal transduction

PubMed Central

Zheng, Long; Zhang, Chao; Li, Long; Hu, Chao; Hu, Mushuang; Sidikejiang, Niyazi; Wang, Xuanchuan; Lin, Miao; Rong, Ruiming

2017-01-01

Previous studies have demonstrated the potential antifibrotic effects of baicalin in vitro, via examination of 21 compounds isolated from plants. However, its biological activity and underlying mechanisms of action in vivo remain to be elucidated. The present study aimed to evaluate the effect of baicalin on renal fibrosis in vivo, and the potential signaling pathways involved. A unilateral ureteral obstruction (UUO)-induced renal fibrosis model was established using Sprague-Dawley rats. Baicalin was administrated intraperitoneally every 2 days for 10 days. The degree of renal damage and fibrosis was investigated by histological assessment, and detection of fibronectin and collagen I mRNA expression levels. Epithelial-mesenchymal transition (EMT) markers, transforming growth factor-β1 (TGF-β1) levels and downstream phosphorylation of mothers against decapentaplegic 2/3 (Smad2/3) were examined in vivo and in an NRK-52E rat renal tubular cell line in vitro. Baicalin was demonstrated to markedly ameliorate renal fibrosis and suppress EMT, as evidenced by reduced interstitial collagen accumulation, decreased fibronectin and collagen I mRNA expression levels, upregulation of N- and E-cadherin expression levels, and downregulation of α-smooth muscle actin and vimentin expression. Furthermore, baicalin decreased TGF-β1 expression levels in serum and kidney tissue following UUO, and suppressed Smad2/3 phosphorylation in rat kidney tissue. In vitro studies identified that baicalin may inhibit the phosphorylation of Smad2/3 under the same TGF-β1 concentration. In conclusion, baicalin may protect against renal fibrosis, potentially via inhibition of TGF-β1 production and its downstream signal transduction. PMID:28260014

DHEA increases epithelial markers and decreases mesenchymal proteins in breast cancer cells and reduces xenograft growth.

PubMed

Colín-Val, Zaira; González-Puertos, Viridiana Yazmín; Mendoza-Milla, Criselda; Gómez, Erika Olivia; Huesca-Gómez, Claudia; López-Marure, Rebeca

2017-10-15

Breast cancer is one of the most common neoplasias and the leading cause of cancer death in women worldwide. Its high mortality rate is linked to a great metastatic capacity associated with the epithelial-mesenchymal transition (EMT). During this process, a decrease in epithelial proteins expression and an increase of mesenchymal proteins are observed. On the other hand, it has been shown that dehydroepiandrosterone (DHEA), the most abundant steroid in human plasma, inhibits migration of breast cancer cells; however, the underlying mechanisms have not been elucidated. In this study, the in vitro effect of DHEA on the expression pattern of some EMT-related proteins, such as E-cadherin (epithelial), N-cadherin, vimentin and Snail (mesenchymal) was measured by Western blot and immunofluorescence in MDA-MB-231 breast cancer cells with invasive, metastatic and mesenchymal phenotype. Also, the in vivo effect of DHEA on xenograft tumor growth in nude mice (nu - /nu - ) and on expression of the same epithelial and mesenchymal proteins in generated tumors was evaluated. We found that DHEA increased expression of E-cadherin and decreased N-cadherin, vimentin and Snail expression both in MD-MB-231 cells and in the formed tumors, possibly by DHEA-induced reversion of mesenchymal phenotype. These results were correlated with a tumor size reduction in mouse xenografts following DHEA administration either a week earlier or concurrent with breast cancer cells inoculation. In conclusion, DHEA could be useful in the treatment of breast cancer with mesenchymal phenotype. Copyright © 2017 Elsevier Inc. All rights reserved.

1α,25-Dihydroxyvitamin D Inhibits the Metastatic Capability of MCF10CA1a and MDA-MB-231 Cells in an In Vitro Model of Breast to Bone Metastasis.

PubMed

Wilmanski, Tomasz; Barnard, Alle; Parikh, Mukti R; Kirshner, Julia; Buhman, Kimberly; Burgess, John; Teegarden, Dorothy

2016-10-01

Breast cancer metastasis to the bone continues to be a major health problem, with approximately 80% of advanced breast cancer patients expected to develop bone metastasis. Although the problem of bone metastasis persists, current treatment options for metastatic cancer patients are limited. In this study, we investigated the preventive role of the active vitamin D metabolite, 1α,25-dihydroxyvitamin D (1,25(OH)2D), against the metastatic potential of breast cancer cells using a novel three-dimensional model (rMET) recapitulating multiple steps of the bone metastatic process. Treatment of MCF10CA1a and MDA-MB-231 cells inhibited metastasis in the rMET model by 70% (±5.7%) and 21% (±6%), respectively. In addition, 1,25(OH)2D treatment decreased invasiveness (20 ± 11% of vehicle) and decreased the capability of MCF10CA1a cells to survive in the reconstructed bone environment after successful invasion through the basement membrane (69 ± 5% of vehicle). An essential step in metastasis is epithelial-mesenchymal transition (EMT). Treatment of MCF10CA1a cells with 1,25(OH)2D increased gene (2.04 ± 0.28-fold increase) and protein (1.87 ± 0.20-fold increase) expression of E-cadherin. Additionally, 1,25(OH)2D treatment decreased N-cadherin gene expression (42 ± 8% decrease), a marker for EMT. Collectively, the present study suggests that 1,25(OH)2D inhibits breast cancer cell metastatic capability as well as inhibits EMT, an essential step in the metastatic process.

ZEB1 drives epithelial-to-mesenchymal transition in lung cancer. | Office of Cancer Genomics

Cancer.gov

Increased expression of zinc finger E-box binding homeobox 1 (ZEB1) is associated with tumor grade and metastasis in lung cancer, likely due to its role as a transcription factor in epithelial-to-mesenchymal transition (EMT). Here, we modeled malignant transformation in human bronchial epithelial cells (HBECs) and determined that EMT and ZEB1 expression are early, critical events in lung cancer pathogenesis. Specific oncogenic mutations in TP53 and KRAS were required for HBECs to engage EMT machinery in response to microenvironmental (serum/TGF-β) or oncogenetic (MYC) factors.

Rubus idaeus L. reverses epithelial-to-mesenchymal transition and suppresses cell invasion and protease activities by targeting ERK1/2 and FAK pathways in human lung cancer cells.

PubMed

Hsieh, Yih-Shou; Chu, Shu-Chen; Hsu, Li-Sung; Chen, Kuo-Shuen; Lai, Ming-Tsung; Yeh, Chia-Heng; Chen, Pei-Ni

2013-12-01

Epithelial to mesenchymal transition (EMT) has been considered essential for cancer metastasis, a multistep complicated process including local invasion, intravasation, extravasation, and proliferation at distant sites. Herein we provided molecular evidence associated with the antimetastatic effect of Rubus idaeus L. extracts (RIE) by showing a nearly complete inhibition on the invasion (p<0.001) of highly metastatic A549 cells via reduced activities of matrix metalloproteinase-2 (MMP-2) and urokinasetype plasminogen activator (u-PA). We performed Western blot to find that RIE could induce up-regulation of epithelial marker such as E-cadherin and α-catenin and inhibit the mesenchymal markers such as N-cadherin, fibronectin, snail-1, and vimentin. Selective snail-1 inhibition by snail-1-specific-siRNA also showed increased E-cadherin expression in A549 cells suggesting a possible involvement of snail-1 inhibition in RIE-caused increase in E-cadherin level. RIE also inhibited p-FAK, p-paxillin and AP-1 by Western blot analysis, indicating the anti-EMT effect of RIE in human lung carcinoma. Importantly, an in vivo BALB/c nude mice xenograft model showed that RIE treatment reduced tumor growth by oral gavage, and RIE represent promising candidates for future phytochemical-based mechanistic pathway-targeted cancer prevention strategies. Copyright © 2013 Elsevier Ltd. All rights reserved.

Pretreatment microRNA Expression Impacting on Epithelial-to-Mesenchymal Transition Predicts Intrinsic Radiosensitivity in Head and Neck Cancer Cell Lines and Patients.

PubMed

de Jong, Monique C; Ten Hoeve, Jelle J; Grénman, Reidar; Wessels, Lodewyk F; Kerkhoven, Ron; Te Riele, Hein; van den Brekel, Michiel W M; Verheij, Marcel; Begg, Adrian C

2015-12-15

Predominant causes of head and neck cancer recurrence after radiotherapy are rapid repopulation, hypoxia, fraction of cancer stem cells, and intrinsic radioresistance. Currently, intrinsic radioresistance can only be assessed by ex vivo colony assays. Besides being time-consuming, colony assays do not identify causes of intrinsic resistance. We aimed to identify a biomarker for intrinsic radioresistance to be used before start of treatment and to reveal biologic processes that could be targeted to overcome intrinsic resistance. We analyzed both microRNA and mRNA expression in a large panel of head and neck squamous cell carcinoma (HNSCC) cell lines. Expression was measured on both irradiated and unirradiated samples. Results were validated using modified cell lines and a series of patients with laryngeal cancer. miRs, mRNAs, and gene sets that correlated with resistance could be identified from expression data of unirradiated cells. The presence of epithelial-to-mesenchymal transition (EMT) and low expression of miRs involved in the inhibition of EMT were important radioresistance determinants. This finding was validated in two independent cell line pairs, in which the induction of EMT reduced radiosensitivity. Moreover, low expression of the most important miR (miR-203) was shown to correlate with local disease recurrence after radiotherapy in a series of patients with laryngeal cancer. These findings indicate that EMT and low expression of EMT-inhibiting miRs, especially miR-203, measured in pretreatment material, causes intrinsic radioresistance of HNSCC, which could enable identification and treatment modification of radioresistant tumors. Clin Cancer Res; 21(24); 5630-8. ©2015 AACR. ©2015 American Association for Cancer Research.

Downregulation of 26S proteasome catalytic activity promotes epithelial-mesenchymal transition

PubMed Central

van Baarsel, Eric D.; Metz, Patrick J.; Fisch, Kathleen; Widjaja, Christella E.; Kim, Stephanie H.; Lopez, Justine; Chang, Aaron N.; Geurink, Paul P.; Florea, Bogdan I.; Overkleeft, Hermen S.; Ovaa, Huib; Bui, Jack D.; Yang, Jing; Chang, John T.

2016-01-01

The epithelial-mesenchymal transition (EMT) endows carcinoma cells with phenotypic plasticity that can facilitate the formation of cancer stem cells (CSCs) and contribute to the metastatic cascade. While there is substantial support for the role of EMT in driving cancer cell dissemination, less is known about the intracellular molecular mechanisms that govern formation of CSCs via EMT. Here we show that β2 and β5 proteasome subunit activity is downregulated during EMT in immortalized human mammary epithelial cells. Moreover, selective proteasome inhibition enabled mammary epithelial cells to acquire certain morphologic and functional characteristics reminiscent of cancer stem cells, including CD44 expression, self-renewal, and tumor formation. Transcriptomic analyses suggested that proteasome-inhibited cells share gene expression signatures with cells that have undergone EMT, in part, through modulation of the TGF-β signaling pathway. These findings suggest that selective downregulation of proteasome activity in mammary epithelial cells can initiate the EMT program and acquisition of a cancer stem cell-like phenotype. As proteasome inhibitors become increasingly used in cancer treatment, our findings highlight a potential risk of these therapeutic strategies and suggest a possible mechanism by which carcinoma cells may escape from proteasome inhibitor-based therapy. PMID:26930717

A sequential EMT-MET mechanism drives the differentiation of human embryonic stem cells towards hepatocytes.

PubMed

Li, Qiuhong; Hutchins, Andrew P; Chen, Yong; Li, Shengbiao; Shan, Yongli; Liao, Baojian; Zheng, Dejin; Shi, Xi; Li, Yinxiong; Chan, Wai-Yee; Pan, Guangjin; Wei, Shicheng; Shu, Xiaodong; Pei, Duanqing

2017-05-03

Reprogramming has been shown to involve EMT-MET; however, its role in cell differentiation is unclear. We report here that in vitro differentiation of hESCs to hepatic lineage undergoes a sequential EMT-MET with an obligatory intermediate mesenchymal phase. Gene expression analysis reveals that Activin A-induced formation of definitive endoderm (DE) accompanies a synchronous EMT mediated by autocrine TGFβ signalling followed by a MET process. Pharmacological inhibition of TGFβ signalling blocks the EMT as well as DE formation. We then identify SNAI1 as the key EMT transcriptional factor required for the specification of DE. Genetic ablation of SNAI1 in hESCs does not affect the maintenance of pluripotency or neural differentiation, but completely disrupts the formation of DE. These results reveal a critical mesenchymal phase during the acquisition of DE, highlighting a role for sequential EMT-METs in both differentiation and reprogramming.

Semaphorin 3 C drives epithelial-to-mesenchymal transition, invasiveness, and stem-like characteristics in prostate cells.

PubMed

Tam, Kevin J; Hui, Daniel H F; Lee, Wilson W; Dong, Mingshu; Tombe, Tabitha; Jiao, Ivy Z F; Khosravi, Shahram; Takeuchi, Ario; Peacock, James W; Ivanova, Larissa; Moskalev, Igor; Gleave, Martin E; Buttyan, Ralph; Cox, Michael E; Ong, Christopher J

2017-09-13

Prostate cancer (PCa) is among the most commonly-occurring cancers worldwide and a leader in cancer-related deaths. Local non-invasive PCa is highly treatable but limited treatment options exist for those with locally-advanced and metastatic forms of the disease underscoring the need to identify mechanisms mediating PCa progression. The semaphorins are a large grouping of membrane-associated or secreted signalling proteins whose normal roles reside in embryogenesis and neuronal development. In this context, semaphorins help establish chemotactic gradients and direct cell movement. Various semaphorin family members have been found to be up- and down-regulated in a number of cancers. One family member, Semaphorin 3 C (SEMA3C), has been implicated in prostate, breast, ovarian, gastric, lung, and pancreatic cancer as well as glioblastoma. Given SEMA3C's roles in development and its augmented expression in PCa, we hypothesized that SEMA3C promotes epithelial-to-mesenchymal transition (EMT) and stem-like phenotypes in prostate cells. In the present study we show that ectopic expression of SEMA3C in RWPE-1 promotes the upregulation of EMT and stem markers, heightened sphere-formation, and cell plasticity. In addition, we show that SEMA3C promotes migration and invasion in vitro and cell dissemination in vivo.

Evidence for pleural epithelial-mesenchymal transition in murine compensatory lung growth

PubMed Central

Ysasi, Alexandra B.; Wagner, Willi L.; Valenzuela, Cristian D.; Kienzle, Arne; Servais, Andrew B.; Bennett, Robert D.; Tsuda, Akira; Ackermann, Maximilian; Mentzer, Steven J.

2017-01-01

In many mammals, including rodents and humans, removal of one lung results in the compensatory growth of the remaining lung; however, the mechanism of compensatory lung growth is unknown. Here, we investigated the changes in morphology and phenotype of pleural cells after pneumonectomy. Between days 1 and 3 after pneumonectomy, cells expressing α-smooth muscle actin (SMA), a cytoplasmic marker of myofibroblasts, were significantly increased in the pleura compared to surgical controls (p < .01). Scanning electron microscopy of the pleural surface 3 days post-pneumonectomy demonstrated regions of the pleura with morphologic features consistent with epithelial-mesenchymal transition (EMT); namely, cells with disrupted intercellular junctions and an acquired mesenchymal (rounded and fusiform) morphotype. To detect the migration of the transitional pleural cells into the lung, a biotin tracer was used to label the pleural mesothelial cells at the time of surgery. By post-operative day 3, image cytometry of post-pneumonectomy subpleural alveoli demonstrated a 40-fold increase in biotin+ cells relative to pneumonectomy-plus-plombage controls (p < .01). Suggesting a similar origin in space and time, the distribution of cells expressing biotin, SMA, or vimentin demonstrated a strong spatial autocorrelation in the subpleural lung (p < .001). We conclude that post-pneumonectomy compensatory lung growth involves EMT with the migration of transitional mesothelial cells into subpleural alveoli. PMID:28542402

Epithelial-mesenchymal transition is associated with a distinct tumor microenvironment including elevation of inflammatory signals and multiple immune checkpoints in lung adenocarcinoma

PubMed Central

Lou, Yanyan; Diao, Lixia; Cuentas, Edwin Roger Parra; Denning, Warren L.; Chen, Limo; Fan, Youhong; Byers, Lauren A.; Wang, Jing; Papadimitrakopoulou, Vassiliki; Behrens, Carmen; Rodriguez, Jaime Canales; Hwu, Patrick; Wistuba, Ignacio I.; Heymach, John V.; Gibbons, Don L.

2016-01-01

Purpose Promising results in the treatment of NSCLC have been seen with agents targeting immune checkpoints, such as PD-1 or PD-L1. However, only a select group of patients respond to these interventions. The identification of biomarkers that predict clinical benefit to immune checkpoint blockade is critical to successful clinical translation of these agents. Methods We conducted an integrated analysis of three independent large datasets, including The Cancer Genome Atlas (TCGA) of lung adenocarcinoma and two datasets from MD Anderson Cancer Center, Profiling of Resistance patterns and Oncogenic Signaling Pathways in Evaluation of Cancers of the Thorax (named PROSPECT) and Biomarker-integrated Approaches of Targeted Therapy for Lung Cancer Elimination (named BATTLE-1). Comprehensive analysis of mRNA gene expression, reverse phase protein array (RPPA), immunohistochemistry and correlation with clinical data were performed. Results Epithelial-mesenchymal transition (EMT) is highly associated with an inflammatory tumor microenvironment in lung adenocarcinoma, independent of tumor mutational burden. We found immune activation co-existent with elevation of multiple targetable immune checkpoint molecules, including PD-L1, PD-L2, PD-1, TIM-3, B7-H3, BTLA and CTLA-4, along with increases in tumor infiltration by CD4+Foxp3+ regulatory T cells in lung adenocarcinomas that displayed an EMT phenotype. Furthermore, we identify B7-H3 as a prognostic marker for NSCLC. Conclusions The strong association between EMT status and an inflammatory tumor microenvironment with elevation of multiple targetable immune checkpoint molecules warrants further investigation of using EMT as a predictive biomarker for immune checkpoint blockade agents and other immunotherapies in NSCLC and possibly a broad range of other cancers. PMID:26851185

Phenotypic Plasticity Determines Cancer Stem Cell Therapeutic Resistance in Oral Squamous Cell Carcinoma

PubMed Central

Biddle, Adrian; Gammon, Luke; Liang, Xiao; Costea, Daniela Elena; Mackenzie, Ian C.

2016-01-01

Cancer stem cells (CSCs) drive tumour spread and therapeutic resistance, and can undergo epithelial-to-mesenchymal transition (EMT) and mesenchymal-to-epithelial transition (MET) to switch between epithelial and post-EMT sub-populations. Examining oral squamous cell carcinoma (OSCC), we now show that increased phenotypic plasticity, the ability to undergo EMT/MET, underlies increased CSC therapeutic resistance within both the epithelial and post-EMT sub-populations. The post-EMT CSCs that possess plasticity exhibit particularly enhanced therapeutic resistance and are defined by a CD44highEpCAMlow/− CD24+ cell surface marker profile. Treatment with TGFβ and retinoic acid (RA) enabled enrichment of this sub-population for therapeutic testing, through which the endoplasmic reticulum (ER) stressor and autophagy inhibitor Thapsigargin was shown to selectively target these cells. Demonstration of the link between phenotypic plasticity and therapeutic resistance, and development of an in vitro method for enrichment of a highly resistant CSC sub-population, provides an opportunity for the development of improved chemotherapeutic agents that can eliminate CSCs. PMID:26981578

[S100A7 promotes the metastasis and epithelial-mesenchymal transition on HeLa and CaSki cells].

PubMed

Tian, T; Hua, Z; Wang, L Z; Wang, X Y; Chen, H Y; Liu, Z H; Cui, Z M

2018-02-25

Objective: To elucidate the impact of over-expression of S100A7 on migration, invasion, proliferation, cell cycle, and epithelial-mesenchymal transition (EMT) in human cervical cancer HeLa and CaSki cells. Methods: (1) Immunohistochemistry of SP was used to examine the expression of S100A7 in 40 cases of squamous cervical cancer tissues and 20 cases of normal cervical tissues. (2) The vectors of pLVX-IRES-Neo-S100A7 and pLVX-IRES-Neo were used to transfect human cervical cancer HeLa and CaSki cells, and the positive clones were screened and identified. Next, transwell migration assay, cell counting kit-8 (CCK-8) assay and fluorescence activating cell sorter (FACS) were used to detect the effect of S100A7-overexpression on the migration, invasion, proliferation and cell cycle of cervical cancer cells. Furthermore, western blot was performed to observe the expression of epithelial marker (E-cadherin) and mesenchymal markers (N-cadherin, vimentin, and fibronectin) of EMT. Results: (1) S100A7 expression was significantly higher in cervical squamous cancer tissues (median 91.6) than that in normal cervical tissues (median 52.1; Z=- 2.948, P= 0.003) . (2) Stable S100A7-overexpressed cells were established using lentiviral-mediated gene delivery in HeLa and CaSki cells. S100A7 was detected by real-time quantitative reverse transcription PCR, S100A7 mRNA of S100A7-overexpressed cells were 119±3 and 177±16, increased significantly compared with control groups of median ( P< 0.01) . Compared with the control cells, the number of S100A7-overexpressed HeLa and CaSki cells that passed the transwell membrane assay were increased significanatly (572±51 vs 337±25, P< 0.01; 100±8 vs 41±4, P< 0.01) .Matrigel invasion assay showed that the number of S100A7-overexpressed HeLa and CaSki cells that passed the transwell membrane were respectively 441±15 and 110±14, elevated significantly compared with control cells (156±21 and 59±7; P< 0.05) . However, S100A7 overexpression didn't influence the proliferation and cell cycle progression of HeLa and CaSki cells ( P> 0.05) . Expression of E-cadherin was dramatically decreased, while N-cadherin, vimentin, and fibronectin increased in S100A7-overexpressed cells. Conclusion: S100A7 enhances the migration, invasion and EMT of HeLa cells and CaSki cells, and may be plays an important role in the development of cervical cancer.

Regulation of HIF-1-Alpha, miR-200, and Markers of Cancer Stem Cells by CDF Under Hypoxic Condition

DTIC Science & Technology

2012-04-01

tumors. It has been well recognized that cancer stem cells (CSCs) and epithelial-to- mesenchymal transition (EMT) phenotypic cells are associated with...epithelial-to- mesenchymal transition (EMT), cancer stem cell (CSC) functions, and inflammation, which contribute to radiation therapy and chemotherapy... Hypoxia induces the VEGF and IL-6 cytokine production in PCa cells and its CSC-like sphere forming cells . ● The CSC-like sphere forming

Saxagliptin Attenuates Albuminuria by Inhibiting Podocyte Epithelial- to-Mesenchymal Transition via SDF-1α in Diabetic Nephropathy.

PubMed

Chang, Yun-Peng; Sun, Bei; Han, Zhe; Han, Fei; Hu, Shao-Lan; Li, Xiao-Yu; Xue, Mei; Yang, Yang; Chen, Li; Li, Chun-Jun; Chen, Li-Ming

2017-01-01

The dipeptidyl peptidase-4 (DPP-4) inhibitor saxagliptin has been found to reduce progressive albuminuria, but the exact mechanism of inhibition is unclear. Podocyte epithelial-to-mesenchymal transition (EMT) has emerged as a potential pathway leading to proteinuria in diabetic nephropathy (DN). Stromal cell-derived factor-1α (SDF-1α), one of the substrates of DPP-4, can activate the protein kinase A pathway and subsequently inhibit its downstream effector, transforming growth factor-β1 (TGF-β1), which induces podocyte EMT. Thus, this study was designed to test the hypothesis that saxagliptin reduces progressive albuminuria by preventing podocyte EMT through inhibition of SDF-1α cleavage in DN. The results of a series of assays, including ELISA, western blotting, and immunochemistry/immunofluorescence, showed that saxagliptin treatment obviously ameliorated urinary microalbumin excretion and renal histological changes in high-fat diet/streptozotocin-induced diabetic rats. Furthermore, saxagliptin-treated diabetic rats presented with suppression of DPP-4 activity/protein expression accompanied by restoration of SDF-1α levels, which subsequently hindered NOX2 expression and podocyte EMT. In vitro , we consistently observed that saxagliptin significantly inhibited increased DPP-4 activity/expression, oxidative stress and podocyte EMT. Application of an SDF-1α receptor inhibitor (AMD3100) to cultured podocytes further confirmed the essential role of SDF-1α in podocyte EMT inhibition. In sum, we demonstrated for the first time that saxagliptin treatment plays an essential role in ameliorating progressive DN by preventing podocyte EMT through a SDF-1α-related pathway, suggesting that saxagliptin could offer renoprotection and that SDF-1α might be a potential therapeutic target for DN.

Saxagliptin Attenuates Albuminuria by Inhibiting Podocyte Epithelial- to-Mesenchymal Transition via SDF-1α in Diabetic Nephropathy

PubMed Central

Chang, Yun-peng; Sun, Bei; Han, Zhe; Han, Fei; Hu, Shao-lan; Li, Xiao-yu; Xue, Mei; Yang, Yang; Chen, Li; Li, Chun-jun; Chen, Li-ming

2017-01-01

The dipeptidyl peptidase-4 (DPP-4) inhibitor saxagliptin has been found to reduce progressive albuminuria, but the exact mechanism of inhibition is unclear. Podocyte epithelial-to-mesenchymal transition (EMT) has emerged as a potential pathway leading to proteinuria in diabetic nephropathy (DN). Stromal cell–derived factor-1α (SDF-1α), one of the substrates of DPP-4, can activate the protein kinase A pathway and subsequently inhibit its downstream effector, transforming growth factor-β1 (TGF-β1), which induces podocyte EMT. Thus, this study was designed to test the hypothesis that saxagliptin reduces progressive albuminuria by preventing podocyte EMT through inhibition of SDF-1α cleavage in DN. The results of a series of assays, including ELISA, western blotting, and immunochemistry/immunofluorescence, showed that saxagliptin treatment obviously ameliorated urinary microalbumin excretion and renal histological changes in high-fat diet/streptozotocin-induced diabetic rats. Furthermore, saxagliptin-treated diabetic rats presented with suppression of DPP-4 activity/protein expression accompanied by restoration of SDF-1α levels, which subsequently hindered NOX2 expression and podocyte EMT. In vitro, we consistently observed that saxagliptin significantly inhibited increased DPP-4 activity/expression, oxidative stress and podocyte EMT. Application of an SDF-1α receptor inhibitor (AMD3100) to cultured podocytes further confirmed the essential role of SDF-1α in podocyte EMT inhibition. In sum, we demonstrated for the first time that saxagliptin treatment plays an essential role in ameliorating progressive DN by preventing podocyte EMT through a SDF-1α-related pathway, suggesting that saxagliptin could offer renoprotection and that SDF-1α might be a potential therapeutic target for DN. PMID:29163166

Diverse Targets of β-Catenin during the Epithelial-Mesenchymal Transition Define Cancer Stem Cells and Predict Disease Relapse.

PubMed

Chang, Yi-Wen; Su, Ying-Jhen; Hsiao, Michael; Wei, Kuo-Chen; Lin, Wei-Hsin; Liang, Chi-Lung; Chen, Shin-Cheh; Lee, Jia-Lin

2015-08-15

Wnt signaling contributes to the reprogramming and maintenance of cancer stem cell (CSC) states that are activated by epithelial-mesenchymal transition (EMT). However, the mechanistic relationship between EMT and the Wnt pathway in CSC is not entirely clear. Chromatin immunoprecipitation with high-throughput sequencing (ChIP-seq) indicated that EMT induces a switch from the β-catenin/E-cadherin/Sox15 complex to the β-catenin/Twist1/TCF4 complex, the latter of which then binds to CSC-related gene promoters. Tandem coimmunoprecipitation and re-ChIP experiments with epithelial-type cells further revealed that Sox15 associates with the β-catenin/E-cadherin complex, which then binds to the proximal promoter region of CASP3. Through this mechanism, Twist1 cleavage is triggered to regulate a β-catenin-elicited promotion of the CSC phenotype. During EMT, we documented that Twist1 binding to β-catenin enhanced the transcriptional activity of the β-catenin/TCF4 complex, including by binding to the proximal promoter region of ABCG2, a CSC marker. In terms of clinical application, our definition of a five-gene CSC signature (nuclear β-catenin(High)/nuclear Twist1(High)/E-cadherin(Low)/Sox15(Low)/CD133(High)) may provide a useful prognostic marker for human lung cancer. ©2015 American Association for Cancer Research.

Silencing of Prrx2 Inhibits the Invasion and Metastasis of Breast Cancer both In Vitro and In Vivo by Reversing Epithelial-Mesenchymal Transition.

PubMed

Lv, Zhi-Dong; Wang, Hai-Bo; Liu, Xiang-Ping; Jin, Li-Ying; Shen, Ruo-Wu; Wang, Xin-Gang; Kong, Bin; Qu, Hui-Li; Li, Fu-Nian; Yang, Qi-Feng

2017-01-01

Epithelial-mesenchymal transition (EMT) is recognized as a crucial mechanism in breast cancer progression and metastasis. Paired-related homeobox 2 (Prrx2) has been identified as a new EMT inducer in cancer, but the underlying mechanisms are still poorly understood. The expression of Prrx2 was assessed by immunohistochemistry in breast cancer tissues to evaluate the clinicopathological significance of Prrx2, as well as the correlation between Prrx2 and EMT. Short hairpin RNA knockdown of Prrx2 was used to examine cellular effects of Prrx2, detecte the expression of Wnt/β-catenin signaling and EMT-associated proteins, and observe cell proliferation, invasion and migration abilities in vitro and in vivo. Clinical association studies showed that Prrx2 expression was related to tumor size, lymph node metastasis, tumor node metastasis stages, EMT and poor survival. Results also showed that knockdown of Prrx2 could alter cell morphology, suppressed the abilities of cell proliferation, invasion and migration in breast cancer. Moreover, silencing of Prrx2 induced the mesenchymal-epithelial transition and prevented nuclear translocation of β-catenin, inhibited wnt/β-catenin signaling pathway. Our study indicated that Prrx2 may be an important activator of EMT in human breast cancer and it can serve as a molecular target of therapeutic interventions for breast cancer. © 2017 The Author(s). Published by S. Karger AG, Basel.

Rapamycin inhibits epithelial-to-mesenchymal transition of peritoneal mesothelium cells through regulation of Rho GTPases.

PubMed

Xiang, Shilong; Li, Meng; Xie, Xishao; Xie, Zhoutao; Zhou, Qin; Tian, Yuanshi; Lin, Weiqiang; Zhang, Xiaohui; Jiang, Hong; Shou, Zhangfei; Chen, Jianghua

2016-06-01

Epithelial-mesenchymal transition (EMT) of peritoneal mesothelial cells (PMCs) is a key process of peritoneal fibrosis. Rapamycin has been previously shown to inhibit EMT of PMCs and prevent peritoneal fibrosis. In this study, we investigated the undefined molecular mechanisms by which rapamycin inhibits EMT of PMCs. To define the protective effect of rapamycin, we initially used a rat PD model which was daily infused with 20 mL of 4.25% high glucose (HG) dialysis solution for 6 weeks to induce fibrosis. The HG rats showed decreased ultrafiltration volume and obvious fibroproliferative response, with markedly increased peritoneal thickness and higher expression of α-smooth muscle actin (α-SMA) and transforming growth factor-β1. Rapamycin significantly ameliorated those pathological changes. Next, we treated rat PMCs with HG to induce EMT and/or rapamycin for indicated time. Rapamycin significantly inhibited HG-induced EMT, which manifests as increased expression of α-SMA, fibronectin, and collagen I, decreased expression of E-cadherin, and increased mobility. HG increased the phosphorylation of PI3K, Akt, and mTOR. Importantly, rapamycin inhibits the RhoA, Rac1, and Cdc42 activated by HG. Moreover, rapamycin repaired the pattern of F-actin distribution induced by HG, reducing the formation of stress fiber, focal adhesion, lamellipodia, and filopodia. Thus, rapamycin shows an obvious protective effect on HG-induced EMT, by inhibiting the activation of Rho GTPases (RhoA, Rac1, and Cdc42). © 2016 Federation of European Biochemical Societies.

Co-culture of 3D tumor spheroids with fibroblasts as a model for epithelial–mesenchymal transition in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Sun-Ah, E-mail: j.sarah.k@gmail.com; Lee, Eun Kyung, E-mail: leeek@catholic.ac.kr; Cancer Evolution Research Center, College of Medicine, The Catholic University of Korea, Seoul 137-701

Epithelial–mesenchymal transition (EMT) acts as a facilitator of metastatic dissemination in the invasive margin of malignant tumors where active tumor–stromal crosstalks take place. Co-cultures of cancer cells with cancer-associated fibroblasts (CAFs) are often used as in vitro models of EMT. We established a tumor–fibroblast proximity co-culture using HT-29 tumor spheroids (TSs) with CCD-18co fibroblasts. When co-cultured with TSs, CCD-18co appeared activated, and proliferative activity as well as cell migration increased. Expression of fibronectin increased whereas laminin and type I collagen decreased in TSs co-cultured with fibroblasts compared to TSs alone, closely resembling the margin of in vivo xenograft tissue. Activemore » TGFβ1 in culture media significantly increased in TS co-cultures but not in 2D co-cultures of cancer cells–fibroblasts, indicating that 3D context-associated factors from TSs may be crucial to crosstalks between cancer cells and fibroblasts. We also observed in TSs co-cultured with fibroblasts increased expression of α-SMA, EGFR and CTGF; reduced expression of membranous β-catenin and E-cadherin, together suggesting an EMT-like changes similar to a marginal region of xenograft tissue in vivo. Overall, our in vitro TS–fibroblast proximity co-culture mimics the EMT-state of the invasive margin of in vivo tumors in early metastasis. - Highlights: • An adjacent co-culture of tumor spheroids and fibroblasts is presented as EMT model. • Activation of fibroblasts and increased cell migration were shown in co-culture. • Expression of EMT-related factors in co-culture was similar to that in tumor tissue. • Crosstalk between spheroids and fibroblasts was demonstrated by secretome analysis.« less

Targeting IκB kinase β/NF-κB signaling in human prostate cancer by a novel IκB kinase β inhibitor CmpdA

PubMed Central

Zhang, Yanting; Lapidus, Rena G.; Liu, Peiyan; Choi, Eun Yong; Adediran, Samusi; Hussain, Arif; Wang, Xinghuan; Liu, Xuefeng; Dan, Han C.

2016-01-01

NF-κB plays an important role in many types of cancer, including prostate cancer (PCa), but the role of the upstream kinase of NF-κB, IKKβ, in PCa has not been fully documented, nor are there any effective IKKβ inhibitors used in clinical settings. Here, we have shown that IKKβ activity is mediated by multiple kinases including IKKα in human PCa cell lines that express activated IKKβ. Immunohistochemical analysis (IHC) of human PCa tissue microarrays (TMA) demonstrates that phosphorylation of IKKα/β within its activation loop gradually increases in low to higher stage tumors as compared to normal tissue. The expression of cell proliferation and survival markers (Ki67, Survivin), epithelial-to-mesenchymal transition (EMT) markers (Slug, Snail), as well as cancer stem cell (CSC) related transcription factors (Nanog, Sox2, Oct-4), also increase in parallel among the respective TMA samples analyzed. IKKβ, but not NF-κB, is found to regulate Nanog, which, in turn, modulates the levels of Oct4, Sox2, Snail and Slug, indicating an essential role of IKKβ in regulating cancer stem cells and EMT. The novel IKKβ inhibitor CmpdA inhibits constitutively activated IKKβ/NF-κB signaling, leading to induction of apoptosis and inhibition of proliferation, migration and stemness in these cells. CmpdA also significantly inhibits tumor growth in xenografts without causing apparent in vivo toxicity. Furthermore, CmpdA and docetaxel act synergistically to inhibit proliferation of PCa cells. These results indicate that IKKβ plays a pivotal role in PCa, and targeting IKKβ, including in combination with docetaxel, may be a potentially useful strategy for treating advanced PCa. PMID:27196761

Stabilization of gene expression and cell morphology after explant recycling during fin explant culture in goldfish.

PubMed

Chenais, Nathalie; Lareyre, Jean-Jacques; Le Bail, Pierre-Yves; Labbe, Catherine

2015-07-01

The development of fin primary cell cultures for in vitro cellular and physiological studies is hampered by slow cell outgrowth, low proliferation rate, poor viability, and sparse cell characterization. Here, we investigated whether the recycling of fresh explants after a first conventional culture could improve physiological stability and sustainability of the culture. The recycled explants were able to give a supplementary cell culture showing faster outgrowth, cleaner cell layers and higher net cell production. The cells exhibited a highly stabilized profile for marker gene expression including a low cytokeratin 49 (epithelial marker) and a high collagen 1a1 (mesenchymal marker) expression. Added to the cell spindle-shaped morphology, motility behavior, and actin organization, this suggests that the cells bore stable mesenchymal characteristics. This contrast with the time-evolving expression pattern observed in the control fresh explants during the first 2 weeks of culture: a sharp decrease in cytokeratin 49 expression was concomitant with a gradual increase in col1a1. We surmise that such loss of epithelial features for the benefit of mesenchymal ones was triggered by an epithelial to mesenchymal transition (EMT) process or by way of a progressive population replacement process. Overall, our findings provide a comprehensive characterization of this new primary culture model bearing mesenchymal features and whose stability over culture time makes those cells good candidates for cell reprogramming prior to nuclear transfer, in a context of fish genome preservation. Copyright © 2015 Elsevier Inc. All rights reserved.

miR-598 inhibits metastasis in colorectal cancer by suppressing JAG1/Notch2 pathway stimulating EMT.

PubMed

Chen, Jia; Zhang, Haichen; Chen, Ying; Qiao, Guanglei; Jiang, Weihua; Ni, Peihua; Liu, Xiangfan; Ma, Lijun

2017-03-01

MicroRNAs (miRNAs) are a class of endogenous, evolutionarily conserved small non-coding RNA molecules that mediate the posttranscriptional process of target gene, leading to translational repression or degradation of target mRNAs. A series of studies have indicated that miRNAs play an important role in tumor initiation, development and progression. In this study, we found that down regulation of miR-598 was a frequent event in CRC tissues compared to the paracarcinoma tissues. And the study demonstrated that miR-598 was implicated in CRC metastasis. Transwell migration assay revealed that elevated miR-598 expression reduces CRC cell migration. Moreover, our study showed that suppression of miR-598 expression induces CRC cell epithelialmesenchymal transition(EMT) and overexpression of miR-598 inhibits CRC cell EMT. In addition, bioinformatics target prediction identified JAG1 as a putative target of miR-598. Knockdown of miR-598 was shown to upregulate JAG1 expression. Furthermore, overexpression of miR-598 suppressed the expression of JAG1. Consistent results were also obtained when the regulation of JAG1 expression by miR-598 was further specified in CRC tissues. Moreover, overexpression of JAG1 induces epithelialmesenchymal transition(EMT) and promotes the metastasis of CRC cells. Decreased Notch2 expression suppresses CRC cells metastasis and EMT. Together, these results indicate that miR-598 is a novel regulator of colorectal cancer metastasis. Our data suggest miR-598 is implicated in regulating Epithelial-mesenchymal transitions by directly suppressing its downstream target gene JAG1 to inactivate Notch signaling pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

Hypoxia-induced PLOD2 promotes proliferation, migration and invasion via PI3K/Akt signaling in glioma.

PubMed

Song, Ye; Zheng, Shihao; Wang, Jizhou; Long, Hao; Fang, Luxiong; Wang, Gang; Li, Zhiyong; Que, Tianshi; Liu, Yi; Li, Yilei; Zhang, Xi'an; Fang, Weiyi; Qi, Songtao

2017-06-27

Gliomas are the most common form of malignant primary brain tumors with poor 5-year survival rate. Dysregulation of procollagen-lysine, 2-oxoglutarate 5-dioxygenase 2 (PLOD2) was observed in gliomas, but the specific role and molecular mechanism of PLOD2 in glioma have not been reported yet. In this study, PLOD2 was found to be frequently up-regulated in glioma and could serve as an independent prognostic marker to identify patients with poor clinical outcome. Knockdown of PLOD2 inhibited proliferation, migration and invasion of glioma cells in vitro and in vivo. Mechanistically, inhibition of PLOD2 inactivated PI3K/AKT signaling pathway and thus regulated the expression of its downstream epithelial-mesenchymal transition (EMT)-associated regulators, including E-cadherin, vimentin, N-cadherin, β-catenin, snail and slug in glioma cells. Moreover, PLOD2 could be induced by hypoxia-inducible factor-1α (HIF-1α) via hypoxia, thereby promoting hypoxia-induced EMT in glioma cells. Our data suggests that PLOD2 may be a potential therapeutic target for patients with glioma.

Transcriptome meta-analysis reveals common differential and global gene expression profiles in cystic fibrosis and other respiratory disorders and identifies CFTR regulators.

PubMed

Clarke, Luka A; Botelho, Hugo M; Sousa, Lisete; Falcao, Andre O; Amaral, Margarida D

2015-11-01

A meta-analysis of 13 independent microarray data sets was performed and gene expression profiles from cystic fibrosis (CF), similar disorders (COPD: chronic obstructive pulmonary disease, IPF: idiopathic pulmonary fibrosis, asthma), environmental conditions (smoking, epithelial injury), related cellular processes (epithelial differentiation/regeneration), and non-respiratory "control" conditions (schizophrenia, dieting), were compared. Similarity among differentially expressed (DE) gene lists was assessed using a permutation test, and a clustergram was constructed, identifying common gene markers. Global gene expression values were standardized using a novel approach, revealing that similarities between independent data sets run deeper than shared DE genes. Correlation of gene expression values identified putative gene regulators of the CF transmembrane conductance regulator (CFTR) gene, of potential therapeutic significance. Our study provides a novel perspective on CF epithelial gene expression in the context of other lung disorders and conditions, and highlights the contribution of differentiation/EMT and injury to gene signatures of respiratory disease. Copyright © 2015 Elsevier Inc. All rights reserved.

Pericytes/vessel-associated mural cells (VAMCs) are the major source of key epithelial-mesenchymal transition (EMT) factors SLUG and TWIST in human glioma.

PubMed

Mäder, Lisa; Blank, Anna E; Capper, David; Jansong, Janina; Baumgarten, Peter; Wirsik, Naita M; Zachskorn, Cornelia; Ehlers, Jakob; Seifert, Michael; Klink, Barbara; Liebner, Stefan; Niclou, Simone; Naumann, Ulrike; Harter, Patrick N; Mittelbronn, Michel

2018-05-08

Epithelial-to-mesenchymal transition (EMT) is supposed to be responsible for increased invasion and metastases in epithelial cancer cells. The activation of EMT genes has further been proposed to be important in the process of malignant transformation of primary CNS tumors. Since the cellular source and clinical impact of EMT factors in primary CNS tumors still remain unclear, we aimed at deciphering their distribution in vivo and clinico-pathological relevance in human gliomas. We investigated 350 glioma patients for the expression of the key EMT factors SLUG and TWIST by immunohistochemistry and immunofluorescence related to morpho-genetic alterations such as EGFR -amplification, IDH-1 (R132H) mutation and 1p/19q LOH. Furthermore, transcriptional cluster and survival analyses were performed. Our data illustrate that SLUG and TWIST are overexpressed in gliomas showing vascular proliferation such as pilocytic astrocytomas and glioblastomas. EMT factors are exclusively expressed by non-neoplastic pericytes/vessel-associated mural cells (VAMCs). They are not associated with patient survival but correlate with pericytic/VAMC genes in glioblastoma cluster analysis. In summary, the upregulation of EMT genes in pilocytic astrocytomas and glioblastomas reflects the level of activation of pericytes/VAMCs in newly formed blood vessels. Our results underscore that the negative prognostic potential of the EMT signature in the group of diffuse gliomas of WHO grade II-IV does most likely not derive from glioma cells but rather reflects the degree of proliferating mural cells thereby constituting a potential target for future alternative treatment approaches.

Lenticular cytoprotection, part 2: link between glycogen synthase kinase-3β, epithelial to mesenchymal transition, and mitochondrial depolarization.

PubMed

Neelam, Sudha; Brooks, Morgan M; Cammarata, Patrick R

2014-01-01

The inhibition of GSK-3β blocks mitochondrial membrane permeability transition (mMPT) for HLE-B3 cells in atmospheric oxygen. GSK-3β, as part of a multifactorial complex, also regulates nuclear levels of β-catenin, a known coordinator of cell survival and adhesion. The purpose of these studies was to demonstrate a novel, but likely disadvantageous, link between β-catenin's influence on the expression of the pro-survival protein, vascular endothelial growth factor (VEGF), resulting in enhanced lens epithelial cell mitochondrial protection against depolarization and nuclear β-catenin as an inducer of epithelial to mesenchymal transition (EMT). Virally transformed human lens epithelial cells (HLE-B3) were treated with SB216763, a specific inhibitor of GSK-3β catalytic activity and XAV939, a specific β-catenin inhibitor that bars the translocation of β-catenin from cytoplasm to the nucleus. Western blot analysis was employed to detect the levels of cytoplasmic and nuclear β-catenin and phospho-β-catenin, pBcl-2 and the EMT proteins, α-smooth muscle actin (α-SMA), and fibronectin. ELISA was used to measure the levels of VEGF in cell culture supernatants. JC-1 analysis was performed to analyze the influence of either SB216763 or XAV939 on mitochondrial depolarization. Cultured lens epithelial cells maintained in hypoxia (1% oxygen) and subsequently reintroduced into atmospheric oxygen and treated with the GSK-3β inhibitor SB216763 illustrated a marked inhibition of phosphorylation of glycogen synthase (downstream substrate of GSK-3β) and significant increase in nuclear translocation of β-catenin. The augmented nuclear β-catenin levels positively correlated with increased expression of α-SMA and fibronectin, both marker proteins indicative of EMT. The enhanced nuclear β-catenin activity also elicited increased VEGF and pBcl-2 expression, resulting in increased resistance to mitochondrial depolarization. Treatment of the cells with the β-catenin inhibitor XAV939 resulted in decreased expression of nuclear β-catenin, VEGF levels, pBcl-2, and EMT proteins, as well as increased mitochondrial depolarization. The data support a model whereby the onset of epithelial to mesenchymal transition may circuitously benefit from the enhanced synthesis of VEGF by setting up a potentially harmful situation whereby the resulting mesenchymal cell population may be more resistant to mitochondrial depolarization than the lens epithelial cell population from which it originated. These findings support the potential therapeutic relevance of developing strategies to undermine the progression of normal cells to mesenchymal transition without subverting cell viability.

The Nuclear Receptor, RORγ, Regulates Pathways Necessary for Breast Cancer Metastasis.

PubMed

Oh, Tae Gyu; Wang, Shu-Ching M; Acharya, Bipul R; Goode, Joel M; Graham, J Dinny; Clarke, Christine L; Yap, Alpha S; Muscat, George E O

2016-04-01

We have previously reported that RORγ expression was decreased in ER-ve breast cancer, and increased expression improves clinical outcomes. However, the underlying RORγ dependent mechanisms that repress breast carcinogenesis have not been elucidated. Here we report that RORγ negatively regulates the oncogenic TGF-β/EMT and mammary stem cell (MaSC) pathways, whereas RORγ positively regulates DNA-repair. We demonstrate that RORγ expression is: (i) decreased in basal-like subtype cancers, and (ii) inversely correlated with histological grade and drivers of carcinogenesis in breast cancer cohorts. Furthermore, integration of RNA-seq and ChIP-chip data reveals that RORγ regulates the expression of many genes involved in TGF-β/EMT-signaling, DNA-repair and MaSC pathways (including the non-coding RNA, LINC00511). In accordance, pharmacological studies demonstrate that an RORγ agonist suppresses breast cancer cell viability, migration, the EMT transition (microsphere outgrowth) and mammosphere-growth. In contrast, RNA-seq demonstrates an RORγ inverse agonist induces TGF-β/EMT-signaling. These findings suggest pharmacological modulation of RORγ activity may have utility in breast cancer. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

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

NASA Astrophysics Data System (ADS)

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.

Electric field-induced suppression of PTEN drives epithelial-to-mesenchymal transition via mTORC1 activation.

PubMed

Yan, Tiantian; Jiang, Xupin; Guo, Xiaowei; Chen, Wen; Tang, Di; Zhang, Junhui; Zhang, Xingyue; Zhang, Dongxia; Zhang, Qiong; Jia, Jiezhi; Huang, Yuesheng

2017-02-01

Naturally occurring electric fields (EFs) are an intrinsic property of wounds. Endogenous EFs in skin wounds play critical roles in the dynamic and well-ordered biological process of wound healing. The epithelial-to-mesenchymal transition (EMT) allows keratinocytes to transition from sedentary cells to motile cells, facilitating wound healing. However, EMT-related studies have been performed without considering endogenous EFs. Thus, the relationship between electrical signals and the EMT remain elusive. Phosphatase and tension homolog (PTEN) and mammalian target of rapamycin complex 1 (mTORC1) are key molecules in sensing electrical cues, and they play significant roles in cellular responses to EFs. In addition, these molecules are closely related to the occurrence of the EMT in other cells. We used primary human keratinocytes to investigate the influence of EFs on the EMT as well as the roles of PTEN and mTORC1 in this process. The effects of EFs on the EMT were investigated by analyzing the levels of specific proteins and transcription factors. The roles of mTORC1 and PTEN and their relationship with each other were studied via pharmacological inhibition or genetic knockdown. A Zeiss imaging system and scratch assays were used to study single-cell motility and monolayer cell migration. EFs induced a range of both biochemical changes (e.g., increased Snail, Slug, vimentin, and N-cadherin expression, decreased E-cadherin expression) and functional changes (e.g., enhanced migratory capacity) that are characteristic of the EMT. EF-stimulated cells exhibited suppressed PTEN expression, and further PTEN downregulation led to the acquisition of more mesenchymal features and the loss of epithelial characteristics, which was accompanied by increased migratory capacity. PTEN overexpression reversed the EF-induced EMT and inhibited the migratory capacity of keratinocytes. EF-induced mTORC1 activation was a required component of the causal relationship between PTEN suppression and the EMT, as mTORC1 inhibition reversed the EMT induced by PTEN downregulation. Our data demonstrate that the EF-induced suppression of PTEN drives the EMT via mTORC1 activation, thereby revealing a new and promising role of EFs in facilitating wound reepithelialization. These results provide a novel perspective regarding the significance of EFs in wound healing; therefore, electrical stimulation offers a new avenue of wound management for improved and accelerated wound healing. Copyright © 2016 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.

MiR-424-5p reversed epithelial-mesenchymal transition of anchorage-independent HCC cells by directly targeting ICAT and suppressed HCC progression

PubMed Central

Zhang, Ying; Li, Tao; Guo, Pengbo; Kang, Jia; Wei, Qing; Jia, Xiaoqing; Zhao, Wei; Huai, Wanwan; Qiu, Yumin; Sun, Lei; Han, Lihui

2014-01-01

Resistance to anoikis and Epithelial-mesenchymal transition (EMT) are two processes critically involved in cancer metastasis. In this study, we demonstrated that after anchorage deprival, hepatocellular carcinoma (HCC) cells not only resisted anoikis, but also exhibited EMT process. Microarray expression profiling revealed that expression of miR-424-5p was significantly decreased in anoikis-resistant HCC cells. Ectopic overexpression of miR-424-5p was sufficient to reverse resistance to anoikis, block EMT process and inhibit malignant behaviors of HCC cells. Target analysis showed that a potent β-catenin inhibitor, ICAT/CTNNBIP1 was a direct target of miR-424-5p. Further study demonstrated that miR-424-5p reversed resistance to anoikis and EMT of HCCs by directly targeting ICAT and further maintaining the E-cadherin/β-catanin complex on the cellular membrance. In vivo study further demonstrated that miR-424-5p significantly inhibited the tumorigenicity of HCC cells in nude mice. Clinical investigation demonstrated that miR-424-5p was significantly downregulated in HCC tissues compared with that of the non-cancerous liver tissues, and this decreased expression of miR-424-5p was significantly correlated with higher pathological grades and more advanced TNM stages. Therefore, aberrant expression of miR-424-5p is critically involved in resistance to anoikis and EMT during the metastatic process of HCC, and its downregulation significantly contributes to liver cancer progression. PMID:25175916

Triptolide suppresses paraquat induced idiopathic pulmonary fibrosis by inhibiting TGFB1-dependent epithelial mesenchymal transition.

PubMed

Chen, Hong; Chen, Qun; Jiang, Chun-Ming; Shi, Guang-Yue; Sui, Bo-Wen; Zhang, Wei; Yang, Li-Zhen; Li, Zhu-Ying; Liu, Li; Su, Yu-Ming; Zhao, Wen-Cheng; Sun, Hong-Qiang; Li, Zhen-Zi; Fu, Zhou

2018-03-01

Idiopathic pulmonary fibrosis (IPF) and tumor are highly similar to abnormal cell proliferation that damages the body. This malignant cell evolution in a stressful environment closely resembles that of epithelial-mesenchymal transition (EMT). As a popular EMT-inducing factor, TGFβ plays an important role in the progression of multiple diseases. However, the drugs that target TGFB1 are limited. In this study, we found that triptolide (TPL), a Chinese medicine extract, exerts an anti-lung fibrosis effect by inhibiting the EMT of lung epithelial cells. In addition, triptolide directly binds to TGFβ and subsequently increase E-cadherin expression and decrease vimentin expression. In in vivo studies, TPL improves the survival state and inhibits lung fibrosis in mice. In summary, this study revealed the potential therapeutic effect of paraquat induced TPL in lung fibrosis by regulating TGFβ-dependent EMT progression. Copyright © 2017 Elsevier B.V. All rights reserved.

The Prevalence of CD146 Expression in Breast Cancer Subtypes and Its Relation to Outcome.

PubMed

de Kruijff, Ingeborg E; Timmermans, Anna M; den Bakker, Michael A; Trapman-Jansen, Anita M A C; Foekens, Renée; Meijer-Van Gelder, Marion E; Oomen-de Hoop, Esther; Smid, Marcel; Hollestelle, Antoinette; van Deurzen, Carolien H M; Foekens, John A; Martens, John W M; Sleijfer, Stefan

2018-05-05

CD146, involved in epithelial-to-mesenchymal transition (EMT), might affect cancer aggressiveness. We here investigated the prevalence of CD146 expression in breast cancer subtypes, its relation to prognosis, the relation between CD146 and EMT and the outcome to tamoxifen. Primary breast cancer tissues from 1342 patients were available for this retrospective study and immunohistochemically stained for CD146. For survival analyses, pure prognosis was studied by only including lymph-node negative patients who did not receive (neo)adjuvant systemic treatment ( n = 551). 11% of the tumors showed CD146 expression. CD146 expression was most prevalent in triple-negative cases (64%, p < 0.001). In univariable analysis, CD146 expression was a prognostic factor for both metastasis-free survival (MFS) ( p = 0.020) and overall survival (OS) ( p = 0.037), but not in multivariable analysis (including age, tumor size, grade, estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2) and Ki-67). No correlation between CD146 and EMT nor difference in outcome to first-line tamoxifen was seen. In this large series, our data showed that CD146 is present in primary breast cancer and is a pure prognostic factor for MFS and OS in breast cancer patients. We did not see an association between CD146 expression and EMT nor on outcome to tamoxifen.

Notch3 negatively regulates chemoresistance in breast cancers.

PubMed

Gu, Xiaoting; Lu, Chunxiao; He, Dongxu; Lu, Yangfan; Jin, Jian; Liu, Dequan; Ma, Xin

2016-10-14

To define the role of the NOTCH signaling pathway in the development of chemoresistance and the associated epithelial-mesenchymal transition (EMT), we investigated the effect of Notch3 on adriamycin (ADM)-resistant human breast cancer cells (MCF-7/ADM cells). We found that Notch3 was downregulated and involved in the chemoresistance of MCF-7/ADM cells, while forced expression of Notch3 reversed the chemoresistance. Furthermore, fos-related antigen 1 (Fra1) was negatively regulated by Notch3 and was highly expressed in MCF-7/ADM cells. Increased Fra1 activated the EMT process. Finally, Notch3 expression was confirmed in clinically chemoresistant samples of breast cancers from patients receiving anthracycline-based chemotherapy. Low expression of Notch3 was an unfavorable predictor of distant relapse-free survival in ER positive breast cancers. Taken together, our findings demonstrate that the Notch3-Fra1 signaling pathway mediates chemoresistance via the EMT.

Histone Deacetylase Inhibitor Induces the Expression of Select Epithelial Genes in Mouse Utricle Sensory Epithelia-Derived Progenitor Cells

PubMed Central

Wang, Jue

2014-01-01

Abstract Mouse utricle sensory epithelial cell–derived progenitor cells (MUCs), which have hair cell progenitor and mesenchymal features via epithelial-to-mesenchymal transition (EMT) as previously described, provide a potential approach for hair cell regeneration via cell transplantation. In this study, we treated MUCs with trichostatin A (TSA) to determine whether histone deacetylase inhibitor is able to stimulate the expression of epithelial genes in MUCs, an essential step for guiding mesenchymal-like MUCs to become sensory epithelial cells. After 72 h of TSA treatment, MUCs acquired epithelial-like features, which were indicated by increased expression of epithelial markers such as Cdh1, Krt18, and Dsp. Additionally, TSA decreased the expression of mesenchymal markers, including Zeb1, Zeb2, Snai1, and Snai2, and prosensory genes Lfng, Six1, and Dlx5. Moreover, the expression of the hair cell genes Atoh1 and Myo6 was increased in TSA-treated MUCs. We also observed significantly decreased expression of Hdac2 and Hdac3 in TSA-treated MUCs. However, no remarkable change was detected in protein expression using immunofluorescence, indicating that TSA-induced HDAC inhibition may contribute to the initial stage of the mesenchymal-to-epithelial phenotypic change. In the future, more work is needed to induce hair cell regeneration using inner ear tissue–derived progenitors to achieve an entire mesenchymal-to-epithelial transition. PMID:24945595

The histone variant H2A.X is a regulator of the epithelial-mesenchymal transition.

PubMed

Weyemi, Urbain; Redon, Christophe E; Choudhuri, Rohini; Aziz, Towqir; Maeda, Daisuke; Boufraqech, Myriem; Parekh, Palak R; Sethi, Taresh K; Kasoji, Manjula; Abrams, Natalie; Merchant, Anand; Rajapakse, Vinodh N; Bonner, William M

2016-02-15

The epithelial-mesenchymal transition (EMT), considered essential for metastatic cancer, has been a focus of much research, but important questions remain. Here, we show that silencing or removing H2A.X, a histone H2A variant involved in cellular DNA repair and robust growth, induces mesenchymal-like characteristics including activation of EMT transcription factors, Slug and ZEB1, in HCT116 human colon cancer cells. Ectopic H2A.X re-expression partially reverses these changes, as does silencing Slug and ZEB1. In an experimental metastasis model, the HCT116 parental and H2A.X-null cells exhibit a similar metastatic behaviour, but the cells with re-expressed H2A.X are substantially more metastatic. We surmise that H2A.X re-expression leads to partial EMT reversal and increases robustness in the HCT116 cells, permitting them to both form tumours and to metastasize. In a human adenocarcinoma panel, H2A.X levels correlate inversely with Slug and ZEB1 levels. Together, these results point to H2A.X as a regulator of EMT.

miR-644a Inhibits Cellular Proliferation and Invasion via Suppression of CtBP1 in Gastric Cancer Cells.

PubMed

Li, Yingchao; Yan, Xiaoni; Ren, Li; Li, Yang

2018-01-19

Epithelial-mesenchymal transition (EMT) is one of the most important mechanisms in the metastasis of various cancers, including gastric cancer (GC). In this study, we explored the putative significance of miR-644a and its role in EMT-mediated metastasis of GC. We first detected the expression of miR-644a in a cohort of 107 GC tissues using quantitative RT-PCR. The expression of miR-644a was suppressed in GC tissues and was associated with a later clinical stage and tumor metastasis. Restoring the expression of miR-644a could significantly suppress the migration and invasion of HGC-27 and SGC-7901 cells, which might be correlated to its suppressive effect on the EMT process. We also found that carboxyl-terminal-binding protein 1 (CtBP1) was a putative target gene of miR-644a in GC and might be involved in the suppressive effect. Collectively, through targeting CtBP1-mediated suppression of the EMT process, miR-644a might suppress the tumor metastasis of GC cells.

Motility and stem cell properties induced by the epithelial-mesenchymal transition require destabilization of lipid rafts

PubMed Central

Prijic, Sara; Chen, Xiaoling; Levental, Ilya; Chang, Jeffrey T.

2016-01-01

The Epithelial-Mesenchymal Transition (EMT) is a developmental program that provides cancer cells with the characteristics necessary for metastasis, including increased motility and stem cell properties. The cellular and molecular mechanisms underlying this process are not yet fully understood, hampering efforts to develop therapeutics. In recent years, it has become apparent that EMT is accompanied by wholesale changes in diverse signaling pathways that are initiated by proteins at the plasma membrane (PM). The PM contains thousands of lipid and protein species that are dynamically and spatially organized into lateral membrane domains, an example of which are lipid rafts. Since one of the major functions of rafts is modulation of signaling originating at the PM, we hypothesized that the signaling changes occurring during an EMT are associated with alterations in PM organization. To test this hypothesis, we used Giant Plasma Membrane Vesicles (GPMVs) to study the organization of intact plasma membranes isolated from live cells. We observed that induction of EMT significantly destabilized lipid raft domains. Further, this reduction in stability was crucial for the maintenance of the stem cell phenotype and EMT-induced remodeling of PM-orchestrated pathways. Exogenously increasing raft stability by feeding cells with ω-3 polyunsaturated fatty acid docosahexaenoic acid (DHA) repressed these phenotypes without altering EMT markers, and inhibited the metastatic capacity of breast cancer cells. Hence, modulating raft properties regulates cell phenotype, suggesting a novel approach for targeting the impact of EMT in cancer. PMID:27303921

Relevance of MicroRNA200 Family and MicroRNA205 for Epithelial to Mesenchymal Transition and Clinical Outcome in Biliary Tract Cancer Patients

PubMed Central

Urbas, Romana; Mayr, Christian; Klieser, Eckhard; Fuereder, Julia; Bach, Doris; Stättner, Stefan; Primavesi, Florian; Jaeger, Tarkan; Stanzer, Stefanie; Ress, Anna Lena; Löffelberger, Magdalena; Wagner, Andrej; Berr, Frieder; Ritter, Markus; Pichler, Martin; Neureiter, Daniel; Kiesslich, Tobias

2016-01-01

Extensive stromal interaction is one reason for the dismal outcome of biliary tract cancer (BTC) patients. Epithelial to mesenchymal transition (EMT) is involved in tumor invasion and metastasis and is partly regulated by microRNAs (miRs). This study explores the expression of anti-EMT miR200 family (miR141, −200a/b/c, −429) and miR205 as well as the EMT-related proteins E-cadherin and vimentin in a panel of BTC cell lines and clinical specimens by quantitative real-time polymerase chain reaction, Western blot and immunohistochemistry, respectively. MicroRNA expression was correlated to (i) the expression patterns of E-cadherin and vimentin; (ii) clinicopathological characteristics; and (iii) survival data. MicroRNA-200 family and miR205 were expressed in all BTC cells and clinical specimens. E-cadherin and vimentin showed a mutually exclusive expression pattern in both, in vitro and in vivo. Expression of miR200 family members positively correlated with E-cadherin and negatively with vimentin expression in BTC cells and specimens. High expression of miR200 family members (but not miR205) and E-cadherin was associated with longer survival, while low miR200 family and high vimentin expression was a predictor of unfavorable survival. Overall, the current study demonstrates the relevance of the miR200 family in EMT of BTC tumors and suggests these miRs as predictors for positive outcome. PMID:27941621

Wnt5a / planar cell polarity signaling pathway in urothelial carcinoma, a potential prognostic biomarker

PubMed Central

Saling, Mark; Duckett, Jordan K; Ackers, Ian; Coschigano, Karen; Jenkinson, Scott; Malgor, Ramiro

2017-01-01

Bladder cancer is the fourth most common cancer in men and the most common malignancy of the urinary tract. Bladder cancers detected at an early stage have a very high five-year survival rate, but when detected after local metastasis the rate is only about 50%. Our group recently reported a positive correlation between the expression of Wnt5a, a member of the Wnt proteins family, and histopathological grade and stage of urothelial carcinoma (UC). The objective of this study was to analyze UC cases reported in Athens, Ohio and investigate the major components of Wnt5a / planar cell polarity (PCP) signaling pathway in UC human tissue samples and UC cell lines. Formalin fixed and paraffin embedded transurethral resection tissues were immunostained for Wnt5a, Ror-2, CTHRC1 and E-cadherin. In addition, in vitro studies using UC cell lines were investigated for Wnt5a/PCP signaling and epithelial mesenchymal transition (EMT) gene expression. The IHC results showed a correlation between the expression of Wnt5a, Ror2 and CTHRC1 with high histological grade of the tumor, while E-cadherin showed an opposite trend of expression. Real time RT-PCR results showed that RNA expression of the Wnt5a/ PCP pathway genes vary in low and high grade UC cell lines and that the high grade cell lines exhibited signs of EMT. These findings support that Wnt5a-Ror2 signaling plays a role in UC, support the potential use of Wnt5a as a prognostic marker and provide evidence that Wnt5a signaling may be used as an effective molecular target for novel therapeutic tools. PMID:28427201

Smad3 linker phosphorylation attenuates Smad3 transcriptional activity and TGF-β1/Smad3-induced epithelial-mesenchymal transition in renal epithelial cells.

PubMed

Bae, Eunjin; Kim, Seong-Jin; Hong, Suntaek; Liu, Fang; Ooshima, Akira

2012-10-26

Transforming growth factor-β1 (TGF-β1) has a distinct role in renal fibrosis associated with epithelial-mesenchymal transition (EMT) of the renal tubules and synthesis of extracellular matrix. Smad3 plays an essential role in fibrosis initiated by EMT. Phosphorylation of Smad3 in the C-terminal SSXS motif by type I TGF-β receptor kinase is essential for mediating TGF-β response. Smad3 activity is also regulated by phosphorylation in the linker region. However, the functional role of Smad3 linker phosphorylation is not well characterized. We now show that Smad3 EPSM mutant, which mutated the four phosphorylation sites in the linker region, markedly enhanced TGF-β1-induced EMT of Smad3-deficient primary renal tubular epithelial cells, whereas Smad3 3S-A mutant, which mutated the C-terminal phosphorylation sites, was unable to induce EMT in response to TGF-β1. Furthermore, immunoblotting and RT-PCR analysis showed a marked induction of fibrogenic gene expression with a significant reduction in E-cadherin in HK2 human renal epithelial cells expressing Smad3 EPSM. TGF-β1 could not induce the expression of α-SMA, vimentin, fibronectin and PAI-1 or reduce the expression of E-cadherin in HK2 cells expressing Smad3 3S-A in response to TGF-β1. Our results suggest that Smad3 linker phosphorylation has a negative regulatory role on Smad3 transcriptional activity and TGF-β1/Smad3-induced renal EMT. Elucidation of mechanism regulating the Smad3 linker phosphorylation can provide a new strategy to control renal fibrosis. Copyright © 2012 Elsevier Inc. All rights reserved.

MicroRNA-200c-141 and ∆Np63 are required for breast epithelial differentiation and branching morphogenesis.

PubMed

Hilmarsdóttir, Bylgja; Briem, Eirikur; Sigurdsson, Valgardur; Franzdóttir, Sigrídur Rut; Ringnér, Markus; Arason, Ari Jon; Bergthorsson, Jon Thor; Magnusson, Magnus Karl; Gudjonsson, Thorarinn

2015-07-15

The epithelial compartment of the breast contains two lineages, the luminal- and the myoepithelial cells. D492 is a breast epithelial cell line with stem cell properties that forms branching epithelial structures in 3D culture with both luminal- and myoepithelial differentiation. We have recently shown that D492 undergo epithelial to mesenchymal transition (EMT) when co-cultured with endothelial cells. This 3D co-culture model allows critical analysis of breast epithelial lineage development and EMT. In this study, we compared the microRNA (miR) expression profiles for D492 and its mesenchymal-derivative D492M. Suppression of the miR-200 family in D492M was among the most profound changes observed. Exogenous expression of miR-200c-141 in D492M reversed the EMT phenotype resulting in gain of luminal but not myoepithelial differentiation. In contrast, forced expression of ∆Np63 in D492M restored the myoepithelial phenotype only. Co-expression of miR-200c-141 and ∆Np63 in D492M restored the branching morphogenesis in 3D culture underlining the requirement for both luminal and myoepithelial elements for obtaining full branching morphogenesis in breast epithelium. Introduction of a miR-200c-141 construct in both D492 and D492M resulted in resistance to endothelial induced EMT. In conclusion, our data suggests that expression of miR-200c-141 and ∆Np63 in D492M can reverse EMT resulting in luminal- and myoepithelial differentiation, respectively, demonstrating the importance of these molecules in epithelial integrity in the human breast. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

Arsenite evokes IL-6 secretion, autocrine regulation of STAT3 signaling, and miR-21 expression, processes involved in the EMT and malignant transformation of human bronchial epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Luo, Fei; Xu, Yuan; The Key Laboratory of Modern Toxicology, Ministry of Education, School of Public Health, Nanjing Medical University

2013-11-15

Arsenite is an established human carcinogen, and arsenite-induced inflammation contributes to malignant transformation of cells, but the molecular mechanisms by which cancers are produced remain to be established. The present results showed that, evoked by arsenite, secretion of interleukin-6 (IL-6), a pro-inflammatory cytokine, led to the activation of STAT3, a transcription activator, and to increased levels of a microRNA, miR-21. Blocking IL-6 with anti-IL-6 antibody and inhibiting STAT3 activation reduced miR-21 expression. For human bronchial epithelial cells, cultured in the presence of anti-IL-6 antibody for 3 days, the arsenite-induced EMT and malignant transformation were reversed. Thus, IL-6, acting on STAT3more » signaling, which up-regulates miR-21in an autocrine manner, contributes to the EMT induced by arsenite. These data define a link from inflammation to EMT in the arsenite-induced malignant transformation of HBE cells. This link, mediated through miRNAs, establishes a mechanism for arsenite-induced lung carcinogenesis. - Highlights: • Arsenite evokes IL-6 secretion. • IL-6 autocrine mediates STAT3 signaling and up-regulates miR-21expression. • Inflammation is involved in arsenite-induced EMT.« less

Loss of BMI-1 dampens migration and EMT of colorectal cancer in inflammatory microenvironment through TLR4/MD-2/MyD88-mediated NF-κB signaling.

PubMed

Ye, Kai; Chen, Qi-Wei; Sun, Ya-Feng; Lin, Jian-An; Xu, Jian-Hua

2018-02-01

Increasing evidence from various clinical and experimental studies has demonstrated that the inflammatory microenvironment created by immune cells facilitates tumor migration. Epithelial-mesenchymal transition (EMT) is involved in the progression of cancer invasion and metastasis in an inflammatory microenvironment. B-lymphoma Moloney murine leukemia virus insertion region 1 (BMI-1) acts as an oncogene in various tumors. Ectopic expression of Bmi-1 have an effect on EMT and invasiveness. The purpose of this study was to investigate the efficacy of BMI-1 on inflammation-induced tumor migration and EMT and the underlying mechanism. We observed that the expression of BMI-1, TNF-α, and IL-1β was significantly increased in HT29 and HCT116 cells after THP-1 Conditioned-Medium (THP-1-CM) stimulation. Additionally, inhibition of BMI-1 impeded cell invasion induced by THP-1-CM-stimulation in both HT29 and HCT116 cells. BMI-1 knockdown remarkably repressed THP-1-CM-induced EMT by regulating the expression of EMT biomarkers with an increase in E-cadherin accompanied by decrease in N-cadherin and vimentin. Furthermore, downregulation of BMI-1 dramatically impeded THP-1-CM-triggered Toll-like receptor 4(TLR4)/myeloid differentiation protein 2(MD-2)/myeloid differentiation factor 88(MyD88) activity by repressing the expression of the TLR4/MD-2 complex and MyD88. Further data demonstrated that knockout of BMI-1 also dampened NF-κB THP-1-CM-triggered activity. Taken all data together, our findings established that BMI-1 modulated TLR4/MD-2/MyD88 complex-mediated NF-κB signaling involved in inflammation-induced cancer cells invasion and EMT, and therefore, could be a potential chemopreventive agent against inflammation-associated colorectal cancer. Establishment of an inflammatory microenvironment. Suppression of BMI-1 reverses THP-1-CM-induced inflammatory cytokine production in CRC. Loss of BMI-1 suppressed TLR4/MD-2/MyD88 complex-mediated NF-κB signaling. © 2017 Wiley Periodicals, Inc.

Resveratrol Inhibits Proliferation, Invasion, and Epithelial-Mesenchymal Transition by Increasing miR-200c Expression in HCT-116 Colorectal Cancer Cells.

PubMed

Karimi Dermani, Fatemeh; Saidijam, Massoud; Amini, Razieh; Mahdavinezhad, Ali; Heydari, Korosh; Najafi, Rezvan

2017-06-01

colorectal cancer (CRC) is one of the most common malignancies, associated with high rates of relapse. A notable challenge in treatment is low response rate to current therapies for advanced CRC. The miR-200c plays an essential role in tumor suppression by inhibiting epithelial-mesenchymal transition (EMT). Resveratrol, a natural compound found in red wine, reveals anti-cancer properties in several types of cancers such as CRC. The aim of current study was to evaluate the effects of resveratrol on proliferation, apoptosis, and invasion of HCT-116 cells and also expression of EMT-related genes in presences or absence of miR-200c. the effect of resveratrol on viability was examined by MTT assay. LNA-anti-miR-200c transfection of HCT-116 cells was carried out in a time dependent manner. Then, the expression of miR-200c and EMT-related genes were quantified by qRT-PCR. Further, expression of EMT-related proteins, apoptosis, and invasion were analyzed by Western blot, Annexin V/PI staining and scratch test, respectively. resveratrol could significantly inhibit viability of HCT-116 cells. LNA-anti-miR-200c suppressed the endogenous miR-200c in transfected cells compared with the control. qRT-PCR and Western blot analysis of LNA-anti-miR-200c transfected cells revealed a considerable increase in vimentin and ZEB-1 expression, with a concomitant reduction in E-cadherin expression level. Migration of HCT-116 cells increased, and apoptosis significantly reduced in transfected cells. While, resveratrol could entirely reverse these changes by modulation of miR-200c expression. our findings revealed a major role of resveratrol in apoptosis, invasion, and switching of EMT to MET phenotype through upregulation of miR-200c in CRC. J. Cell. Biochem. 118: 1547-1555, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Analyses of publicly available genomics resources define FGF-2-expressing bladder carcinomas as EMT-prone, proliferative tumors with low mutation rates and high expression of CTLA-4, PD-1 and PD-L1.

PubMed

McNiel, Elizabeth A; Tsichlis, Philip N

2017-01-01

FGF-2 is overexpressed in a subset of invasive bladder carcinomas and its overexpression correlates with poor prognosis. Analyses of publicly available databases addressing the molecular mechanisms that may be responsible for the poor prognosis of these tumors, revealed that FGF-2 expression correlates positively with the expression of EMT-promoting transcription factors and with changes in gene expression that are characteristic of EMT. The same analyses also revealed that FGF-2 correlates negatively with the expression, mutation and copy number variations of FGFR-3, all of which are associated with non-invasive bladder carcinomas. Finally, they showed that FGF-2 expression correlates with the expression of FGFR-1, the expression of the IIIc variant of FGFR-2 and with the expression of Akt3. The latter observation is significant because our earlier studies had shown that Akt3 regulates FGFR-2 alternative splicing, shifting the balance toward the IIIc relative to the IIIb FGFR-2 splice variant. Since the IIIc variant is recognized by FGF-2, while the IIIb variant is not, we conclude that Akt3 may facilitate the FGF-2 response. FGF-2 is known to promote the expression of KDM2B, which functions in concert with EZH2 to repress the EZH2-targeting microRNA miR-101, activating a switch, which stably upregulates EZH2. TCGA data showing a correlation between KDM2B and EZH2 expression and Oncomine data, showing a correlation between KDM2B and tumor progression, strongly support the role of the FGF-2/KDM2B/miR-101/EZH2 pathway in bladder cancer. These observations combined, suggest a model according to which FGF-2 induces EMT, cell proliferation and cancer stem cell self-renewal by coupling the Akt3 and KDM2B-controlled pathways outlined above, in bladder carcinomas. Further analyses of publicly-available databases, revealed that FGF-2-expressing bladder carcinomas carry fewer genetic alterations and they tend to express high levels of CTLA-4, PD-1 and PD-L1, which suggests immune blockade by checkpoint activation. EMT, enhanced proliferation and immune checkpoint activation combined, may be responsible for the poor prognosis of FGF-2-expressing bladder carcinomas.

NFATC3-PLA2G15 Fusion Transcript Identified by RNA Sequencing Promotes Tumor Invasion and Proliferation in Colorectal Cancer Cell Lines.

PubMed

Jang, Jee-Eun; Kim, Hwang-Phill; Han, Sae-Won; Jang, Hoon; Lee, Si-Hyun; Song, Sang-Hyun; Bang, Duhee; Kim, Tae-You

2018-06-14

This study was designed to identify novel fusion transcripts (FTs) and their functional significance in colorectal cancer lines. We performed paired-end RNA sequencing of 28 colorectal cancer (CRC) cell lines. FT candidates were identified using TopHat-fusion, ChimeraScan, and FusionMap tools and further experimental validation was conducted through reverse transcription-polymerase chain reaction and Sanger sequencing. FT was depleted in human CRC line and the effects on cell proliferation, cell migration, and cell invasion were analyzed. 1,380 FT candidates were detected through bioinformatics filtering. We selected 6 candidate FTs, including 4 inter-chromosomal and 2 intra-chromosomal FTs and each FT was found in at least 1 of the 28 cell lines. Moreover, when we tested 19 pairs of CRC tumor and adjacent normal tissue samples, NFATC3-PLA2G15 FT was found in 2. Knockdown of NFATC3-PLA2G15 using siRNA reduced mRNA expression of epithelial-mesenchymal transition (EMT) markers such as vimentin, twist, and fibronectin and increased mesenchymal-epithelial transition markers of E-cadherin, claudin-1, and FOXC2 in colo-320 cell line harboring NFATC3-PLA2G15 FT. The NFATC3-PLA2G15 knockdown also inhibited invasion, colony formation capacity, and cell proliferation. These results suggest that that NFATC3-PLA2G15 FTs may contribute to tumor progression by enhancing invasion by EMT and proliferation.

Knockdown of POLDIP2 suppresses tumor growth and invasion capacity and is linked to unfavorable transformation ability and metastatic feature in non-small cell lung cancer.

PubMed

Chen, Ying-Chieh; Kuo, Chih-Chi; Chian, Chih-Feng; Tzao, Ching; Chang, Shan-Yueh; Shih, Yu-Lueng; Lin, Ya-Wen; Yu, Mu-Hsien; Su, Her-Young

2018-07-01

The main problem in the treatment of non-small cell lung cancer (NSCLC) is metastasis. Epithelial-mesenchymal transition (EMT) is known as the critical signaling in tumor progression, metastasis, and also the drug resistance. In this study, we reported a novel gene Polymerase delta-interacting protein 2 (POLDIP2) was downregulated in NSCLC tissues and first demonstrated that overexpression of POLDIP2 increased the anchorage-independent growth (AIG) and invasiveness of H1299 cells. In addition, we examined that knockdown of POLDIP2 in H1299 and A549 cells reduced tumorigenicity and metastatic capacity in vitro and also in vivo. Moreover, downregulation of the cell proliferation marker cyclin D1 and EMT markers CDH2, Slug, and Twist was showed in H1299 cells by POLDIP2 knockdown, suggesting that the inhibition of malignancy was affected by modulating key genes for tumor growth and invasiveness. Taken together, our study is the first study that demonstrated that POLDIP2 gene was function as an oncogene in NSCLC and implied the oncogenic ability might be through promoting cell proliferation or EMT. Copyright © 2018 Elsevier Inc. All rights reserved.

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.

PubMed

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 < 5 μm) are able to induce 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 translocation to nucleus - likely reinforces signalling for EMT promotion. In vivo results supported the occurrence of pulmonary fibrosis following MWCNT exposure. We demonstrate a new molecular mechanism of MWCNT-mediated EMT, which is Smad-independent and involves TGF-β and its intracellular effectors Akt/GSK-3β that activate the SNAIL-1 signalling pathway. This finding suggests potential novel targets in the development of therapeutic and preventive approaches.

Thioredoxin 1 mediates TGF-β-induced epithelial-mesenchymal transition in salivary adenoid cystic carcinoma.

PubMed

Jiang, Yang; Feng, Xin; Zheng, Lei; Li, Sheng-Lin; Ge, Xi-Yuan; Zhang, Jian-Guo

2015-09-22

Epithelial-mesenchymal transition (EMT) plays an important role in the invasion and metastasis of salivary adenoid cystic carcinoma (SACC) which is characterized by wide local infiltration, perineural spread, a propensity to local recurrence and late distant metastasis. Our recent studies have disclosed that TGF-β is a crucial factor for EMT in metastatic SACC. In this study, we further uncovered small redox protein thioredoxin 1 (TXN) as a critical mediator of TGF-β induced EMT. Immunohistochemistry analysis revealed significantly higher expressions of TXN, thioredoxin reductase 1 (TXNRD1) and N-cadherin, and lower expression of E-cadherin in human metastatic SACC compared to non-metastatic SACC tissues. Consistently, cultured SACC cells with stable TXN overexpression had decreased E-cadherin and increased N-cadherin as well as Snail and Slug expressions. The enhanced migration and invasion potential of these cells was abrogated by Akt or TXNRD1 inhibitors. Expression of N-cadherin and Akt p-Akt decreased, whereas E-cadherin expression increased in a BBSKE (TXNRD1 inhibitor)-dose-dependent manner. In a xenograft mouse model, TXN overexpression facilitated the metastatic potential of SACC-83 cells to the lung. Our results indicate that TXN plays a key role in SACC invasion and metastasis through the modulation of TGF-β-Akt/GSK-3β on EMT. TXN could be a potential therapeutic target for SACC.

Thymoquinone inhibits cancer metastasis by downregulating TWIST1 expression to reduce epithelial to mesenchymal transition.

PubMed

Khan, Md Asaduzzaman; Tania, Mousumi; Wei, Chunli; Mei, Zhiqiang; Fu, Shelly; Cheng, Jingliang; Xu, Jianming; Fu, Junjiang

2015-08-14

Proteins that promote epithelial to mesenchymal transition (EMT) are associated with cancer metastasis. Inhibition of EMT regulators may be a promising approach in cancer therapy. In this study, Thymoquinone (TQ) was used to treat cancer cell lines to investigate its effects on EMT-regulatory proteins and cancer metastasis. We show that TQ inhibited cancer cell growth, migration and invasion in a dose-dependent manner. At the molecular level, TQ treatment decreased the transcriptional activity of the TWIST1 promoter and the mRNA expression of TWIST1, an EMT-promoting transcription factor. Accordingly, TQ treatment also decreased the expression of TWIST1-upregulated genes such as N-Cadherin and increased the expression of TWIST1-repressed genes such as E-Cadherin, resulting in a reduction of cell migration and invasion. TQ treatment also inhibited the growth and metastasis of cancer cell-derived xenograft tumors in mice but partially attenuated the migration and invasion in TWIST1-overexpressed cell lines. Furthermore, we found that TQ treatment enhanced the promoter DNA methylation of the TWIST1 gene in BT 549 cells. Together, these results demonstrate that TQ treatment inhibits TWIST1 promoter activity and decreases its expression, leading to the inhibition of cancer cell migration, invasion and metastasis. These findings suggest TQ as a potential small molecular inhibitor of cancer growth and metastasis.

Thymoquinone inhibits cancer metastasis by downregulating TWIST1 expression to reduce epithelial to mesenchymal transition

PubMed Central

Khan, Md. Asaduzzaman; Tania, Mousumi; Wei, Chunli; Mei, Zhiqiang; Fu, Shelly; Cheng, Jingliang; Xu, Jianming; Fu, Junjiang

2015-01-01

Proteins that promote epithelial to mesenchymal transition (EMT) are associated with cancer metastasis. Inhibition of EMT regulators may be a promising approach in cancer therapy. In this study, Thymoquinone (TQ) was used to treat cancer cell lines to investigate its effects on EMT-regulatory proteins and cancer metastasis. We show that TQ inhibited cancer cell growth, migration and invasion in a dose-dependent manner. At the molecular level, TQ treatment decreased the transcriptional activity of the TWIST1 promoter and the mRNA expression of TWIST1, an EMT-promoting transcription factor. Accordingly, TQ treatment also decreased the expression of TWIST1-upregulated genes such as N-Cadherin and increased the expression of TWIST1-repressed genes such as E-Cadherin, resulting in a reduction of cell migration and invasion. TQ treatment also inhibited the growth and metastasis of cancer cell-derived xenograft tumors in mice but partially attenuated the migration and invasion in TWIST1-overexpressed cell lines. Furthermore, we found that TQ treatment enhanced the promoter DNA methylation of the TWIST1 gene in BT 549 cells. Together, these results demonstrate that TQ treatment inhibits TWIST1 promoter activity and decreases its expression, leading to the inhibition of cancer cell migration, invasion and metastasis. These findings suggest TQ as a potential small molecular inhibitor of cancer growth and metastasis. PMID:26023736

Dbl oncogene expression in MCF-10 A epithelial cells disrupts mammary acinar architecture, induces EMT and angiogenic factor secretion

PubMed Central

Vanni, Cristina; Ognibene, Marzia; Finetti, Federica; Mancini, Patrizia; Cabodi, Sara; Segalerba, Daniela; Torrisi, Maria Rosaria; Donnini, Sandra; Bosco, Maria Carla; Varesio, Luigi; Eva, Alessandra

2015-01-01

The proteins of the Dbl family are guanine nucleotide exchange factors (GEFs) of Rho GTPases and are known to be involved in cell growth regulation. Alterations of the normal function of these proteins lead to pathological processes such as developmental disorders, neoplastic transformation, and tumor metastasis. We have previously demonstrated that expression of Dbl oncogene in lens epithelial cells modulates genes encoding proteins involved in epithelial-mesenchymal-transition (EMT) and induces angiogenesis in the lens. Our present study was undertaken to investigate the role of Dbl oncogene in epithelial cells transformation, providing new insights into carcinoma progression.To assess how Dbl oncogene can modulate EMT, cell migration, morphogenesis, and expression of pro-apoptotic and angiogenic factors we utilized bi- and 3-dimensional cultures of MCF-10 A cells. We show that upon Dbl expression MCF-10 A cells undergo EMT. In addition, we found that Dbl overexpression sustains Cdc42 and Rac activation inducing morphological alterations, characterized by the presence of lamellipodia and conferring a high migratory capacity to the cells. Moreover, Dbl expressing MCF-10 A cells form altered 3D structures and can induce angiogenesis by producing proangiogenic factors such as CCL2. These results support a role for Dbl oncogene in epithelial cell differentiation and transformation and suggest the relevance of GEF deregulation in tumor onset and progression. PMID:25723869

Cytotoxic and mutagenic properties of shale oil byproducts. II. Comparison of mutagenic effects at five genetic markers induced by retort process water plus near ultraviolet light in Chinese hamster ovary cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, D.J.C.; Strniste, G.F.

1982-01-01

A Chinese hamster ovary (CHO) cell line heterozygous at the adenine phosphoribosyl transferase (APRT) locus was used for selection of induced mutants resistant to 8-azaadenine (8AA), 6-thioguanine (6TG), ouabain (OUA), emetine (EMT) and diphtheria toxin (DIP). The expression times necessary for optimizing the number of mutants recovered at the different loci have been determined using the known direct acting mutagen, far ultraviolet light (FUV), and a complex aqueous organic mixture (shale oil process water) activated with near ultraviolet light (NUV). The results indicate that optimal expression times following treatment with either mutagen was between 2 and 8 days. For CHOmore » cells treated with shale oil process water and subsequently exposed to NUV a linear dose response for mutant induction was observed for all five genetic loci. At 10% surviving fraction of cells, between 35- and 130-fold increases above backgound mutation frequencies were observed for the various markers examined.« less

Mitochondrial ribosomal protein S18-2 is highly expressed in endometrial cancers along with free E2F1

PubMed Central

Iurchenko, Natalia; Kovalevska, Larysa; Stip, Maria C; Budnikova, Daria; Andersson, Sonia; Polischuk, Ludmila; Buchynska, Lubov; Kashuba, Elena

2016-01-01

Endometrial cancer (EC) is one of the most frequent causes of cancer death among women in developed countries. Histopathological diagnosis and imaging techniques for EC are limited, thus new prognostic markers are needed to offer patients the best treatment and follow-up. In the present paper we showed that the level of mitochondrial ribosomal protein MRPS18-2 (S18-2) increased in EC compared with the normal endometrium and hyperplasia, based on a study of 42 patient biopsies. Importantly, high expression of free E2F1 in EC correlates well with high S18-2 expression. The EC cell line HEC-1-A, which overexpresses S18-2 constitutively, showed an increased proliferation capacity in vitro and in vivo (in SCID mice). Moreover, pan-keratin, beta-catenin and E-cadherin signals are diminished in these cells, compared to the parental HEC-1-A line, in contrast to vimentin signal that is increased. This may be associated with epithelial-mesenchymal cell transition (EMT). We conclude that high expression of S18-2 and free E2F1, and low pan-keratin, beta-catenin, and E-cadherin signals might be a good set of prognostic markers for EC. PMID:26959119

Inhibition of Bevacizumab-induced Epithelial-Mesenchymal Transition by BATF2 Overexpression Involves the Suppression of Wnt/β-Catenin Signaling in Glioblastoma Cells.

PubMed

Huang, Wenqiu; Zhang, Chenguang; Cui, Mengtian; Niu, Jing; Ding, Wei

2017-08-01

Bevacizumab (BV) has been used for the treatment of recurrent glioblastoma. However, it also induces epithelial-mesenchymal transition (EMT) in glioblastoma cells, which compromises its efficacy. BATF2 (basic leucine zipper ATF-like transcription factor 2), a multi-target transcriptional repressor, has been found to suppress cancer development partly through inhibition of Wnt/β-catenin singling. The roles of BATF2 and Wnt/β-catenin signaling in BV-induced EMT in glioblastoma cells were investigated in this study. BV was used to treat U87MG cells, and TOP/FOP FLASH luciferase reporters were employed to determine the activity of Wnt/β-catenin signaling. EMT markers were detected with quantitative reverse transcription-PCR and western blotting. Immunofluorescence (IF) was used to determine the compartmentation of β-catenin. Wound-healing, TransWell and ECIS assays were used to analyze cell adhesion, invasion and migration. BV induced EMT phenotype in U87MG cells, and BATF2 overexpression significantly inhibited BV-induced EMT with suppression of Wnt/β-catenin signaling. Our findings expanded the understanding of the role of BATF2 in tumors, and also suggested a potential of using BATF2 as a therapeutic target to hinder bevacizumab induced EMT in glioblastoma. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

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

PubMed

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.

Interplay between Trx-1 and S100P promotes colorectal cancer cell epithelial-mesenchymal transition by up-regulating S100A4 through AKT activation.

PubMed

Zuo, Zhigui; Zhang, Peili; Lin, Feiyan; Shang, Wenjing; Bi, Ruichun; Lu, Fengying; Wu, Jianbo; Jiang, Lei

2018-04-01

We previously reported a novel positive feedback loop between thioredoxin-1 (Trx-1) and S100P, which promotes the invasion and metastasis of colorectal cancer (CRC). However, the underlying molecular mechanisms remain poorly understood. In this study, we examined the roles of Trx-1 and S100P in CRC epithelial-to-mesenchymal transition (EMT) and their underlying mechanisms. We observed that knockdown of Trx-1 or S100P in SW620 cells inhibited EMT, whereas overexpression of Trx-1 or S100P in SW480 cells promoted EMT. Importantly, S100A4 and the phosphorylation of AKT were identified as potential downstream targets of Trx-1 and S100P in CRC cells. Silencing S100A4 or inhibition of AKT phosphorylation eliminated S100P- or Trx-1-mediated CRC cell EMT, migration and invasion. Moreover, inhibition of AKT activity reversed S100P- or Trx-1-induced S100A4 expression. The expression of S100A4 was higher in human CRC tissues compared with their normal counterpart tissues and was significantly correlated with lymph node metastasis and poor survival. The overexpression of S100A4 protein was also positively correlated with S100P or Trx-1 protein overexpression in our cohort of CRC tissues. In addition, overexpression of S100P reversed the Trx-1 knockdown-induced inhibition of S100A4 expression, EMT and migration and invasion in SW620 cells. The data suggest that interplay between Trx-1 and S100P promoted CRC EMT as well as migration and invasion by up-regulating S100A4 through AKT activation, thus providing further potential therapeutic targets for suppressing the EMT in metastatic CRC. © 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Recruitment and retention of emergency medical technicians: a qualitative study.

PubMed

Patterson, P Daniel; Probst, Janice C; Leith, Katherine H; Corwin, Sara J; Powell, M Paige

2005-01-01

Emergency medical technicians (EMTs) are critical to out-of-hospital care, but maintaining staff can be difficult. The study objective was to identify factors that contribute to recruitment and retention of EMTs and paramedics. Information was drawn from three focus groups of EMT-Basic, EMT-Intermediate, and EMT-Paramedic personnel recruited from participants at an annual conference. Thoughts and feelings of EMTs and paramedics were investigated using eight questions designed to explore entry into emergency medical services, what it is like to be an EMT or paramedic, and the EMT educational process. Data were analyzed at the group level for common themes using NVivo. For a majority of respondents, emergency medical services was not a primary career path. Most respondents entered the industry as an alternate or replacement for a nursing career or as a second career following military medic service. The majority of respondents believed the job was stressful yet rewarding, and although it negatively affected their personal lives, the occupation gave them a sense of accomplishment and belonging. Respondents expressed a preference for EMT education resulting in college credit or licensure versus professional certification. Job-related stress produced by numerous factors appears to be a likely contributor to low employee retention. Recruitment and retention efforts should address study findings, incorporating key findings into educational, evaluation, and job enhancement programs.

Migration speed and directionality switch of normal epithelial cells after TGF-β1-induced EMT (tEMT) on micro-structured polydimethylsiloxane (PDMS) substrates with variations in stiffness and topographic patterning.

PubMed

Wu, Tsung-Hsien; Li, Chia-Hui; Tang, Ming-Jer; Liang, Jen-I; Chen, Chia-Hsin; Yeh, Ming-Long

2013-10-01

The epithelial to mesenchymal transition (EMT) involves several physiological and pathological phenomena and endows cells with invasive and migratory properties. However, the effects of substrate stiffness and topography on the migration of cells before or after transforming growth factor-β1 (TGF-β1)-induced EMT (tEMT) are unknown. Herein, we seed control or tEMT NMuMG cells on the 2D patterns consisted of 1 μm or 5 μm line-widths and groove or cone patterns on either 2 MPa (1.96 ± 0.48 MPa) or 4 MPa (3.70 ± 0.74 MPa) polydimethylsiloxane (PDMS) substrates. After tEMT, the increased expression of α-SMA with vinculin in focal adhesion (FA) sites led to an acceleration of tEMT cell motility. On the 2 MPa substrate, the most influenced substrate was the 1 μm, cone-patterned substrate, where the tEMT cells' motility decelerated by 0.13 μm/min (36% slower than the cells on groove pattern). However, on the 5 μm, groove-patterned substrate, where the tEMT cells demonstrated the most rapid motility relative to the control cells, with an increment of 0.18 μm/min (100%). Among the different physical cues from substrate, the cone pattern could impede the migration speed of tEMT cells. Furthermore, we recommend the groove-patterned with a 5 μm line-width substrate as a useful tool to differentiate control and tEMT cells by migration speed.

Tangeretin enhances radiosensitivity and inhibits the radiation-induced epithelial-mesenchymal transition of gastric cancer cells.

PubMed

Zhang, Xukui; Zheng, Luming; Sun, Yinggang; Wang, Tianxiao; Wang, Baocheng

2015-07-01

Irradiation has been reported to increase radioresistance and epithelial-mesenchymal transition (EMT) in gastric cancer (GC) cells. The Notch pathway is critically implicated in cancer EMT and radioresistance. In the present study, we investigated the use of a Notch-1 inhibiting compound as a novel therapeutic candidate to regulate radiation-induced EMT in GC cells. According to previous screening, tangeretin, a polymethoxylated flavonoid from citrus fruits was selected as a Notch-1 inhibitor. Tangeretin enhanced the radiosensitivity of GC cells as demonstrated by MTT and colony formation assays. Tangeretin also attenuated radiation-induced EMT, invasion and migration in GC cells, accompanied by a decrease in Notch-1, Jagged1/2, Hey-1 and Hes-1 expressions. Tangeretin triggered the upregulation of miR-410, a tumor-suppressive microRNA. Furthermore, re-expression of miR-410 prevented radiation-induced EMT and cell invasion. An in vivo tumor xenograft model confirmed the antimetastasis effect of tangeretin as we observed in vitro. In nude mice, tumor size was considerably diminished by radiation plus tangeretin co-treatment. Tangeretin almost completely inhibited lung metastasis induced by irradiation. Tangeretin may be a novel antimetastatic agent for radiotherapy.

14-3-3ζ and aPKC-ι synergistically facilitate epithelial-mesenchymal transition of cholangiocarcinoma via GSK-3β/Snail signaling pathway.

PubMed

Yang, Yan; Liu, Yan; He, Jun-Chuang; Wang, Jian-Ming; Schemmer, Peter; Ma, Chao-Qun; Qian, Ya-Wei; Yao, Wei; Zhang, Jian; Qi, Wei-Peng; Fu, Yang; Feng, Wei; Yang, Tao

2016-08-23

Cholangiocarcinoma (CCA) invasion and metastasis are the primary causes of poor survival rates in patients. The epithelial-mesenchymal transition (EMT) is a crucial step in cancer invasion and metastasis. However, it is still unclear of the molecular mechanism. In this study, the expression of 14-3-3ζ and atypical protein kinase C-ι (aPKC-ι) was further detected in CCA tissues and cell lines. Meanwhile, we established the EMT model of CCA cells and investigated 14-3-3ζ and aPKC-ι co-regulatory effect on the EMT in vitro and in vivo. Further, we identified the downstream molecular glycogen synthase kinase 3 beta (GSK-3β)/Snail signalling pathway that contribute to regulating the EMT. Our data showed that the expression of 14-3-3ζ and aPKC-ι was synergistically increased in CCA tissues compared with adjacent noncancerous tissues and was intimately associated with differentiation and the tumour-node-metastasis (TNM) stage. Multivariate Cox regression analysis indicated that high 14-3-3ζ and aPKC-ι expression separately predicted a poor prognosis and were independent prognostic indicators in patients with CCA. The CO-IP experiment confirmed that the mutual binding relationship between 14-3-3ζ and aPKC-ι. Small interfering RNAs and siRNA rescue experiment demonstrated that 14-3-3ζ and aPKC-ι regulated each other. In addition, 14-3-3ζ and aPKC-ι pretreatment by si-RNA inhibit the phosphorylated GSK-3β and Snail expression during EMT. Meanwhile, silence of 14-3-3ζ or aPKC-ι suppressed CCA cells migration, metastasis and proliferation in vitro and in vivo. Our study demonstrates that 14-3-3ζ and aPKC-ι synergistically facilitate EMT of CCA via GSK-3β/Snail signalling pathway, and may be potential therapeutic target for CCA.

14-3-3ζ and aPKC-ι synergistically facilitate epithelial-mesenchymal transition of cholangiocarcinoma via GSK-3β/snail signaling pathway

PubMed Central

He, Jun-chuang; Wang, Jian-ming; Schemmer, Peter; Ma, Chao-qun; Qian, Ya-wei; Yao, Wei; Zhang, Jian; Qi, Wei-peng; Fu, Yang; Feng, Wei; Yang, Tao

2016-01-01

Cholangiocarcinoma (CCA) invasion and metastasis are the primary causes of poor survival rates in patients. The epithelial-mesenchymal transition (EMT) is a crucial step in cancer invasion and metastasis. However, it is still unclear of the molecular mechanism. In this study, the expression of 14-3-3ζ and atypical protein kinase C-ι (aPKC-ι) was further detected in CCA tissues and cell lines. Meanwhile, we established the EMT model of CCA cells and investigated 14-3-3ζ and aPKC-ι co-regulatory effect on the EMT in vitro and in vivo. Further, we identified the downstream molecular glycogen synthase kinase 3 beta (GSK-3β)/Snail signalling pathway that contribute to regulating the EMT. Our data showed that the expression of 14-3-3ζ and aPKC-ι was synergistically increased in CCA tissues compared with adjacent noncancerous tissues and was intimately associated with differentiation and the tumour-node-metastasis (TNM) stage. Multivariate Cox regression analysis indicated that high 14-3-3ζ and aPKC-ι expression separately predicted a poor prognosis and were independent prognostic indicators in patients with CCA. The CO-IP experiment confirmed that the mutual binding relationship between 14-3-3ζ and aPKC-ι. Small interfering RNAs and siRNA rescue experiment demonstrated that 14-3-3ζ and aPKC-ι regulated each other. In addition, 14-3-3ζ and aPKC-ι pretreatment by si-RNA inhibit the phosphorylated GSK-3β and Snail expression during EMT. Meanwhile, silence of 14-3-3ζ or aPKC-ι suppressed CCA cells migration, metastasis and proliferation in vitro and in vivo. Our study demonstrates that 14-3-3ζ and aPKC-ι synergistically facilitate EMT of CCA via GSK-3β/Snail signalling pathway, and may be potential therapeutic target for CCA. PMID:27409422

Decreased eIF3e/Int6 expression causes epithelial-to-mesenchymal transition in breast epithelial cells.

PubMed

Gillis, L D; Lewis, S M

2013-08-01

eIF3e/Int6 is a component of the multi-subunit eIF3 complex, which binds directly to the 40S ribosome to facilitate ribosome recruitment to mRNA and hence protein synthesis. Reduced expression of eIF3e/Int6 has been found in up to 37% of human breast cancers, and expression of a truncated mutant version of the mouse eIF3e/Int6 protein leads to malignant transformation of normal mammary cells. These findings suggest that eIF3e/Int6 is a tumor suppressor; however, a recent study has reported that a reduction of eIF3e/Int6 expression in breast cancer cells leads to reduced translation of oncogenes, suggesting that eIF3e/Int6 may in fact have an oncogenic role in breast cancer. To gain a better understanding of the role of eIF3e/Int6 in breast cancer, we have examined the effects of decreased eIF3e/Int6 expression in an immortalized breast epithelial cell line, MCF-10A. Surprisingly, we find that decreased expression of eIF3e/Int6 causes breast epithelial cells to undergo epithelial-to-mesenchymal transition (EMT). We show that EMT induced by a decrease in eIF3e/Int6 expression imparts invasive and migratory properties to breast epithelial cells, suggesting that regulation of EMT by eIF3e/Int6 may have an important role in breast cancer metastasis. Furthermore, we show that reduced eIF3e/Int6 expression in breast epithelial cells causes a specific increase in the expression of the key EMT regulators Snail1 and Zeb2, which occurs at both the transcriptional and post-transcriptional levels. Together, our data indicate a novel role of eIF3e/Int6 in the regulation of EMT in breast epithelial cells and support a tumor suppressor role of eIF3e/Int6.

Epithelial–mesenchymal transition-associated secretory phenotype predicts survival in lung cancer patients

PubMed Central

Reka, Ajaya Kumar; Chen, Guoan; Keshamouni, Venkateshwar G.

2014-01-01

In cancer cells, the process of epithelial–mesenchymal transition (EMT) confers migratory and invasive capacity, resistance to apoptosis, drug resistance, evasion of host immune surveillance and tumor stem cell traits. Cells undergoing EMT may represent tumor cells with metastatic potential. Characterizing the EMT secretome may identify biomarkers to monitor EMT in tumor progression and provide a prognostic signature to predict patient survival. Utilizing a transforming growth factor-β-induced cell culture model of EMT, we quantitatively profiled differentially secreted proteins, by GeLC-tandem mass spectrometry. Integrating with the corresponding transcriptome, we derived an EMT-associated secretory phenotype (EASP) comprising of proteins that were differentially upregulated both at protein and mRNA levels. Four independent primary tumor-derived gene expression data sets of lung cancers were used for survival analysis by the random survival forests (RSF) method. Analysis of 97-gene EASP expression in human lung adenocarcinoma tumors revealed strong positive correlations with lymph node metastasis, advanced tumor stage and histological grade. RSF analysis built on a training set (n = 442), including age, sex and stage as variables, stratified three independent lung cancer data sets into low-, medium- and high-risk groups with significant differences in overall survival. We further refined EASP to a 20 gene signature (rEASP) based on variable importance scores from RSF analysis. Similar to EASP, rEASP predicted survival of both adenocarcinoma and squamous carcinoma patients. More importantly, it predicted survival in the early-stage cancers. These results demonstrate that integrative analysis of the critical biological process of EMT provides mechanism-based and clinically relevant biomarkers with significant prognostic value. PMID:24510113

The role of hERG1 ion channels in epithelial-mesenchymal transition and the capacity of riluzole to reduce cisplatin resistance in colorectal cancer cells.

PubMed

Fortunato, Angelo

2017-08-01

The transition of cells from the epithelial to the mesenchymal state (EMT) plays an important role in tumor progression. EMT allows cells to acquire mobility, stem-like behavior and resistance to apoptosis and drug treatment. These features turn EMT into a central process in tumor biology. Ion channels are attractive targets for the treatment of cancer since they play critical roles in controlling a wide range of physiological processes that are frequently deregulated in cancer. Here, we investigated the role of ether-a-go-go-related 1 (hERG1) ion channels in the EMT of colorectal cancer cells. We studied the epithelial-mesenchymal profile of different colorectal cancer-derived cell lines and the expression of hERG1 potassium channels in these cell lines using real-time PCR. Next, we knocked down hERG1 expression in HCT116 cells using lentivirus mediated RNA interference and characterized the hERG1 silenced cells in vitro and in vivo. Finally, we investigated the capacity of riluzole, an ion channel-modulating drug used in humans to treat amyotrophic lateral sclerosis, to reduce the resistance of the respective colorectal cancer cells to the chemotherapeutic drug cisplatin. We found that of the colorectal cancer-derived cell lines tested, HCT116 showed the highest mesenchymal profile and a high hERG1 expression. Subsequent hERG1 expression knockdown induced a change in cell morphology, which was accompanied by a reduction in the proliferative and tumorigenic capacities of the cells. Notably, we found that hERG1expression knockdown elicited a reversion of the EMT profile in HCT116 cells with a reacquisition of the epithelial-like profile. We also found that riluzole increased the sensitivity of HCT116 cisplatin-resistant cells to cisplatin. Our data indicate that hERG1 plays a role in the EMT of colorectal cancer cells and that its knockdown reduces the proliferative and tumorigenic capacities of these cells. In addition, we conclude that riluzole may be used in combination with cisplatin to reduce chemo-resistance in colorectal cancer cells.

Promotion of Cancer Stem-Like Cell Properties in Hepatitis C Virus-Infected Hepatocytes

PubMed Central

Kwon, Young-Chan; Bose, Sandip K.; Steele, Robert; Meyer, Keith; Di Bisceglie, Adrian M.; Ray, Ratna B.

2015-01-01

ABSTRACT We have previously reported that hepatitis C virus (HCV) infection of primary human hepatocytes (PHH) induces the epithelial mesenchymal transition (EMT) state and extends hepatocyte life span (S. K. Bose, K. Meyer, A. M. Di Bisceglie, R. B. Ray, and R. Ray, J Virol 86:13621–13628, 2012, http://dx.doi.org/10.1128/JVI.02016-12). These hepatocytes displayed sphere formation on ultralow binding plates and survived for more than 12 weeks. The sphere-forming hepatocytes expressed a number of cancer stem-like cell (CSC) markers, including high levels of the stem cell factor receptor c-Kit. The c-Kit receptor is regarded as one of the CSC markers in hepatocellular carcinoma (HCC). Analysis of c-Kit mRNA displayed a significant increase in the liver biopsy specimens of chronically HCV-infected patients. We also found c-Kit is highly expressed in transformed human hepatocytes (THH) infected in vitro with cell culture-grown HCV genotype 2a. Further studies suggested that HCV core protein significantly upregulates c-Kit expression at the transcriptional level. HCV infection of THH led to a significant increase in the number of spheres displayed on ultralow binding plates and in enhanced EMT and CSC markers and tumor growth in immunodeficient mice. The use of imatinib or dasatinib as a c-Kit inhibitor reduced the level of sphere-forming cells in culture. The sphere-forming cells were sensitive to treatment with sorafenib, a multikinase inhibitor, that is used for HCC treatment. Further, stattic, an inhibitor of the Stat3 molecule, induced sphere-forming cell death. A combination of sorafenib and stattic had a significantly stronger effect, leading to cell death. These results suggested that HCV infection potentiates CSC generation, and selected drugs can be targeted to efficiently inhibit cell growth. IMPORTANCE HCV infection may develop into HCC as an end-stage liver disease. We focused on understanding the mechanism for the risk of HCC from chronic HCV infection and identified targets for treatment. HCV-infected primary and transformed human hepatocytes (PHH or THH) generated CSC. HCV-induced spheres were highly sensitive to cell death from sorafenib and stattic treatment. Thus, our study is highly significant for HCV-associated HCC, with the potential for developing a target-specific strategy for improved therapies. PMID:26355082

CDKL2 promotes epithelial-mesenchymal transition and breast cancer progression

PubMed Central

Li, Linna; Liu, Chunping; Amato, Robert J.; Chang, Jeffrey T.; Du, Guangwei; Li, Wenliang

2014-01-01

The epithelial–mesenchymal transition (EMT) confers mesenchymal properties on epithelial cells and has been closely associated with the acquisition of aggressive traits by epithelial cancer cells. To identify novel regulators of EMT, we carried out cDNA screens that covered 500 human kinases. Subsequent characterization of candidate kinases led us to uncover cyclin-dependent kinase-like 2 (CDKL2) as a novel potent promoter for EMT and breast cancer progression. CDKL2-expressing human mammary gland epithelial cells displayed enhanced mesenchymal traits and stem cell-like phenotypes, which was acquired through activating a ZEB1/E-cadherin/β-catenin positive feedback loop and regulating CD44 mRNA alternative splicing to promote conversion of CD24high cells to CD44high cells. Furthermore, CDKL2 enhanced primary tumor formation and metastasis in a breast cancer xenograft model. Notably, CDKL2 is expressed significantly higher in mesenchymal human breast cancer cell lines than in epithelial lines, and its over-expression/amplification in human breast cancers is associated with shorter disease-free survival. Taken together, our study uncovered a major role for CDKL2 in promoting EMT and breast cancer progression. PMID:25333262

BAG3 regulates epithelial-mesenchymal transition and angiogenesis in human hepatocellular carcinoma.

PubMed

Xiao, Heng; Cheng, Shaobing; Tong, Rongliang; Lv, Zheng; Ding, Chaofeng; Du, Chengli; Xie, Haiyang; Zhou, Lin; Wu, Jian; Zheng, Shusen

2014-03-01

Bcl2-associated athanogene 3 (BAG3) protein is a co-chaperone of heat-shock protein (Hsp) 70 and may regulate major physiological and pathophysiological processes. However, few reports have examined the role of BAG3 in human hepatocellular carcinoma (HCC). In this study, we show that BAG3 regulates epithelial-mesenchymal transition (EMT) and angiogenesis in HCC. BAG3 was overexpressed in HCC tissues and cell lines. BAG3 knockdown resulted in reduction in migration and invasion of HCC cells, which was linked to reversion of EMT by increasing E-cadherin expression and decreasing N-cadherin, vimentin and slug expression, as well as suppressing matrix metalloproteinase 2 (MMP-2) expression. In a xenograft tumorigenicity model, BAG3 knockdown effectively inhibited tumor growth and metastasis through reduction in CD34 and VEGF expression and reversal of the EMT pathway. In conclusion, BAG3 is associated with the invasiveness and angiogenesis in HCC, and the BAG3 gene may be a novel therapeutic approach against HCC.

The Z-cad dual fluorescent sensor detects dynamic changes between the epithelial and mesenchymal cellular states.

PubMed

Toneff, M J; Sreekumar, A; Tinnirello, A; Hollander, P Den; Habib, S; Li, S; Ellis, M J; Xin, L; Mani, S A; Rosen, J M

2016-06-17

The epithelial to mesenchymal transition (EMT) has been implicated in metastasis and therapy resistance of carcinomas and can endow cancer cells with cancer stem cell (CSC) properties. The ability to detect cancer cells that are undergoing or have completed EMT has typically relied on the expression of cell surface antigens that correlate with an EMT/CSC phenotype. Alternatively these cells may be permanently marked through Cre-mediated recombination or through immunostaining of fixed cells. The EMT process is dynamic, and these existing methods cannot reveal such changes within live cells. The development of fluorescent sensors that mirror the dynamic EMT state by following the expression of bona fide EMT regulators in live cells would provide a valuable new tool for characterizing EMT. In addition, these sensors will allow direct observation of cellular plasticity with respect to the epithelial/mesenchymal state to enable more effective studies of EMT in cancer and development. We generated a lentiviral-based, dual fluorescent reporter system, designated as the Z-cad dual sensor, comprising destabilized green fluorescent protein containing the ZEB1 3' UTR and red fluorescent protein driven by the E-cadherin (CDH1) promoter. Using this sensor, we robustly detected EMT and mesenchymal to epithelial transition (MET) in breast cancer cells by flow cytometry and fluorescence microscopy. Importantly, we observed dynamic changes in cellular populations undergoing MET. Additionally, we used the Z-cad sensor to identify and isolate minor subpopulations of cells displaying mesenchymal properties within a population comprising predominately epithelial-like cells. The Z-cad dual sensor identified cells with CSC-like properties more effectively than either the ZEB1 3' UTR or E-cadherin sensor alone. The Z-cad dual sensor effectively reports the activities of two factors critical in determining the epithelial/mesenchymal state of carcinoma cells. The ability of this stably integrating dual sensor system to detect dynamic fluctuations between these two states through live cell imaging offers a significant improvement over existing methods and helps facilitate the study of EMT/MET plasticity in response to different stimuli and in cancer pathogenesis. Finally, the versatile Z-cad sensor can be adapted to a variety of in vitro or in vivo systems to elucidate whether EMT/MET contributes to normal and disease phenotypes.

LINE-1 couples EMT programming with acquisition of oncogenic phenotypes in human bronchial epithelial cells.

PubMed

Reyes-Reyes, Elsa M; Aispuro, Ivan; Tavera-Garcia, Marco A; Field, Matthew; Moore, Sara; Ramos, Irma; Ramos, Kenneth S

2017-11-28

Although several lines of evidence have established the central role of epithelial-to-mesenchymal-transition (EMT) in malignant progression of non-small cell lung cancers (NSCLCs), the molecular events connecting EMT to malignancy remain poorly understood. This study presents evidence that Long Interspersed Nuclear Element-1 (LINE-1) retrotransposon couples EMT programming with malignancy in human bronchial epithelial cells (BEAS-2B). This conclusion is supported by studies showing that: 1) activation of EMT programming by TGF-β1 increases LINE-1 mRNAs and protein; 2) the lung carcinogen benzo(a)pyrene coregulates TGF-β1 and LINE-1 mRNAs, with LINE-1 positioned downstream of TGF-β1 signaling; and, 3) forced expression of LINE-1 in BEAS-2B cells recapitulates EMT programming and induces malignant phenotypes and tumorigenesis in vivo . These findings identify a TGFβ1-LINE-1 axis as a critical effector pathway that can be targeted for the development of precision therapies during malignant progression of intractable NSCLCs.

Complement 3 activates the renal renin-angiotensin system by induction of epithelial-to-mesenchymal transition of the nephrotubulus in mice.

PubMed

Zhou, Xueli; Fukuda, Noboru; Matsuda, Hiroyuki; Endo, Morito; Wang, Xiaofei; Saito, Kosuke; Ueno, Takahiro; Matsumoto, Taro; Matsumoto, Koichi; Soma, Masayoshi; Kobayashi, Naohiko; Nishiyama, Akira

2013-10-01

We have demonstrated that mesenchymal cells from spontaneously hypertensive rats genetically express complement 3 (C3). Mature tubular epithelial cells can undergo epithelial-to-mesenchymal transition (EMT) that is linked to the pathogenesis of renal fibrosis and injury. In this study, we investigated the contribution of C3 in EMT and in the renal renin-angiotensin (RA) systems associated with hypertension. C3a induced EMT in mouse TCMK-1 epithelial cells, which displayed increased expression of renin and Krüppel-like factor 5 (KLF5) and nuclear localization of liver X receptor α (LXRα). C3 and renin were strongly stained in the degenerated nephrotubulus and colocalized with LXRα and prorenin receptor in unilateral ureteral obstruction (UUO) kidneys from wild-type mice. In C3-deficient mice, hydronephrus and EMT were suppressed, with no expression of renin and C3. After UUO, systolic blood pressure was increased in wild-type but not C3-deficient mice. In wild-type mice, intrarenal angiotensin II (ANG II) levels were markedly higher in UUO kidneys than normal kidneys and decreased with aliskiren. There were no increases in intrarenal ANG II levels after UUO in C3-deficient mice. Thus C3 induces EMT and dedifferentiation of epithelial cells, which produce renin through induction of LXRα. These data indicate for the first time that C3 may be a primary factor to activate the renal RA systems to induce hypertension.

Amyloid precursor protein regulates migration and metalloproteinase gene expression in prostate cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miyazaki, Toshiaki; Ikeda, Kazuhiro; Horie-Inoue, Kuniko

Highlights: • APP knockdown reduced proliferation and migration of prostate cancer cells. • APP knockdown reduced expression of metalloproteinase and EMT-related genes. • APP overexpression promoted LNCaP cell migration. • APP overexpression increased expression of metalloproteinase and EMT-related genes. - Abstract: Amyloid precursor protein (APP) is a type I transmembrane protein, and one of its processed forms, β-amyloid, is considered to play a central role in the development of Alzheimer’s disease. We previously showed that APP is a primary androgen-responsive gene in prostate cancer and that its increased expression is correlated with poor prognosis for patients with prostate cancer. APPmore » has also been implicated in several human malignancies. Nevertheless, the mechanism underlying the pro-proliferative effects of APP on cancers is still not well-understood. In the present study, we explored a pathophysiological role for APP in prostate cancer cells using siRNA targeting APP (siAPP). The proliferation and migration of LNCaP and DU145 prostate cancer cells were significantly suppressed by siAPP. Differentially expressed genes in siAPP-treated cells compared to control siRNA-treated cells were identified by microarray analysis. Notably, several metalloproteinase genes, such as ADAM10 and ADAM17, and epithelial–mesenchymal transition (EMT)-related genes, such as VIM, and SNAI2, were downregulated in siAPP-treated cells as compared to control cells. The expression of these genes was upregulated in LNCaP cells stably expressing APP when compared with control cells. APP-overexpressing LNCaP cells exhibited enhanced migration in comparison to control cells. These results suggest that APP may contribute to the proliferation and migration of prostate cancer cells by modulating the expression of metalloproteinase and EMT-related genes.« less

A1E reduces stemness and self-renewal in HPV 16-positive cervical cancer stem cells.

PubMed

Kwon, Taeho; Bak, Yesol; Ham, Sun-Young; Yu, Dae-Yeul; Yoon, Do-Young

2016-02-02

Cervical cancer is the second most common cancer in females. Recent reports have revealed the critical role of cervical cancer stem cells (CSCs) in tumorigenicity and metastasis. Previously we demonstrated that A1E exerts an anti-proliferative action, which inhibits the growth of cervical cancer cells. A1E is composed of 11 oriental medicinal herbs. Cervical cancer cell culture, wund healing and invasion assay, flow cytometry, sheroid formation assay, and wstern blot assays were performed in HPV 16-positive SiHa cell and HPV 16-negative C33A cells. A1E targets the E6 and E7 oncogenes; thus, A1E significantly inhibited proliferation of human papilloma virus (HPV) 16-positive SiHa cells, it did not inhibit the proliferation of HPV-negative C33A cells. Accordingly, we investigated whether A1E can regulate epithelial-to-mesenchymal transition (EMT), CSC self-renewal, and stemness-related gene expression in cervical cancer cells. Down rgulation of cell migration, cell invasion, and EMT was observed in A1E-treated SiHa cells. Specifically, A1E-treated SiHa cells showed significant decreases in OCT-3/4 and Sox2 expression levels and in sphere formation. Moreover, CSCs makers ALDH+ and ALDH, CD133 double positive cell were significantly decreased in A1E-treated SiHa cells. However, A1E treatment did not down regulate ALDH+ expression and the number of ALDH/CD133 double positive cells in C33A cells. Taken together, A1E can inhibit CSCs and reduce the expression of stemness markers. Treating CSCs with A1E may be a potential therapy for cervical cancer.

Fatty Acid Synthase Mediates the Epithelial-Mesenchymal Transition of Breast Cancer Cells

PubMed Central

Li, Junqin; Dong, Lihua; Wei, Dapeng; Wang, Xiaodong; Zhang, Shuo; Li, Hua

2014-01-01

This study aimed to investigate the role of fatty acid synthase (FASN) in the epithelial-mesenchymal transition (EMT) of breast cancer cells. MCF-7 cells and MCF-7 cells overexpressing mitogen-activated protein kinase 5 (MCF-7-MEK5) were used in this study. MCF-7-MEK5 cells showed stable EMT characterized by increased vimentin and decreased E-cadherin expression. An In vivo animal model was established using the orthotopic injection of MCF-7 or MCF-7-MEK5 cells. Real-time quantitative PCR and western blotting were used to detect the expression levels of FASN and its downstream proteins liver fatty acid-binding protein (L-FABP) and VEGF/VEGFR-2 in both in vitro and in vivo models (nude mouse tumor tissues). In MCF-7-MEK5 cells, significantly increased expression of FASN was associated with increased levels of L-FABP and VEGF/VEGFR-2. Cerulenin inhibited MCF-7-MEK5 cell migration and EMT, and reduced FASN expression and down-stream proteins L-FABP, VEGF, and VEGFR-2. MCF-7-MEK5 cells showed higher sensitivity to Cerulenin than MCF-7 cells. Immunofluorescence revealed an increase of co-localization of FASN with VEGF on the cell membrane and with L-FABP within MCF-7-MEK5 cells. Immunohistochemistry further showed that increased percentage of FASN-positive cells in the tumor tissue was associated with increased percentages of L-FABP- and VEGF-positive cells and the Cerulenin treatment could reverse the effect. Altogether, our results suggest that FASN is essential to EMT possibly through regulating L-FABP, VEGF and VEGFR-2. This study provides a theoretical basis and potential strategy for effective suppression of malignant cells with EMT. PMID:24520215

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takahashi, Atsushi; Graduate School of Science and Engineering, Saitama University, Saitama 338-8570; Green Tea Laboratory, Saitama Prefectural Agriculture and Forestry Research Center, Saitama 358-0042

Highlights: •EGCG reduced cell motility of highly metastatic human lung cancer cells. •EGCG increased cell stiffness of the cells, indicating the inhibition of phenotypes of EMT. •EGCG inhibited expression of vimentin and Slug in the cells at the leading edge of scratch. •Treatment of MβCD increased cell stiffness, and inhibited cell motility and vimentin expression. •Inhibition of EMT phenotypes with EGCG is a mechanism-based inhibition of cancer metastasis. -- Abstract: Cell motility and cell stiffness are closely related to metastatic activity of cancer cells. (−)-Epigallocatechin gallate (EGCG) has been shown to inhibit spontaneous metastasis of melanoma cell line into themore » lungs of mice, so we studied the effects of EGCG on cell motility, cell stiffness, and expression of vimentin and Slug, which are molecular phenotypes of epithelial–mesenchymal transition (EMT). Treatments of human non-small cell lung cancer cell lines H1299 and Lu99 with 50 and 100 μM EGCG reduced cell motility to 67.5% and 43.7% in H1299, and 71.7% and 31.5% in Lu99, respectively in in vitro wound healing assay. Studies on cell stiffness using atomic force microscope (AFM) revealed that treatment with 50 μM EGCG increased Young’s modulus of H1299 from 1.24 to 2.25 kPa and that of Lu99 from 1.29 to 2.28 kPa, showing a 2-fold increase in cell stiffness, i.e. rigid elasticity of cell membrane. Furthermore, treatment with 50 μM EGCG inhibited high expression of vimentin and Slug in the cells at a leading edge of scratch. Methyl-β-cyclodextrin, a reagent to deplete cholesterol in plasma membrane, showed inhibition of EMT phenotypes similar that by EGCG, suggesting that EGCG induces inhibition of EMT phenotypes by alteration of membrane organization.« less

The actin filament cross-linker L-plastin confers resistance to TNF-α in MCF-7 breast cancer cells in a phosphorylation-dependent manner

PubMed Central

Janji, Bassam; Vallar, Laurent; Tanoury, Ziad Al; Bernardin, François; Vetter, Guillaume; Schaffner-Reckinger, Elisabeth; Berchem, Guy; Friederich, Evelyne; Chouaib, Salem

2010-01-01

Abstract We used a tumour necrosis factor (TNF)-α resistant breast adenocarcinoma MCF-7 cell line to investigate the involvement of the actin cytoskeleton in the mechanism of cell resistance to this cytokine. We found that TNF resistance correlates with the loss of cell epithelial properties and the gain of a mesenchymal phenotype, reminiscent of an epithelial-to-mesenchymal transition (EMT). Morphological changes were associated with a profound reorganization of the actin cytoskeleton and with a change in the repertoire of expressed actin cytoskeleton genes and EMT markers, as revealed by DNA microarray-based expression profiling. L-plastin, an F-actin cross-linking and stabilizing protein, was identified as one of the most significantly up-regulated genes in TNF-resistant cells. Knockdown of L-plastin in these cells revealed its crucial role in conferring TNF resistance. Importantly, overexpression of wild-type L-plastin in TNF-sensitive MCF-7 cells was sufficient to protect them against TNF-mediated cell death. Furthermore, we found that this effect is dependent on serine-5 phosphorylation of L-plastin and that non-conventional protein kinase C isoforms and the ceramide pathway may regulate its phosphorylation state. The protective role of L-plastin was not restricted to TNF-α resistant MCF-7 cells because a correlation between the expression of L-plastin and the resistance to TNF-α was observed in other breast cancer cell lines. Together, our study discloses a novel unexpected role of the actin bundling protein L-plastin as a cell protective protein against TNF-cytotoxicity. PMID:19799649

Hypoxia-induced Bmi1 promotes renal tubular epithelial cell–mesenchymal transition and renal fibrosis via PI3K/Akt signal

PubMed Central

Du, Rui; Xia, Lin; Ning, Xiaoxuan; Liu, Limin; Sun, Wenjuan; Huang, Chen; Wang, Hanmin; Sun, Shiren

2014-01-01

Hypoxia is an important microenvironmental factor in the development of renal fibrosis; however, the underlying mechanisms are not well elucidated. Here we show that hypoxia induces Bmi1 mRNA and protein expression in human tubular epithelial cells. We further demonstrate that Bmi1 expression might be directly regulated by hypoxia-inducible factor-1a (HIF-1a) under low oxygen. Moreover, chromatin immunoprecipitation and reporter gene assay studies reveal cooperative transactivation of Bmi1 by HIF-1α and Twist. Enforced Bmi1 expression induces epithelial–mesenchymal transition (EMT), whereas silencing endogenous Bmi-1 expression reverses hypoxia-induced EMT. Up-regulation of Bmi1 leads to stabilization of Snail via modulation of PI3K/Akt signaling, whereas ablation of PI3K/Akt signaling partially rescues the phenotype of Bmi1-overexpressing cells, indicating that PI3K/Akt signaling might be a major mediator of Bmi1-induced EMT. In a rat model of obstructive nephropathy, Bmi1 expression increases in a time-dependent manner. Furthermore, we demonstrate that increased levels of Bmi1, correlated with HIF-1α and Twist, are associated with patients with chronic kidney disease. We provide in vitro and in vivo evidence that activation of HIF-1a/Twist-Bmi1 signaling in renal epithelial cells is associated with the development of chronic renal disease and may promote fibrogenesis via modulation of PI3K/Akt/Snail signaling by facilitating EMT. PMID:25009285

Loss of 4E-BP1 function induces EMT and promotes cancer cell migration and invasion via cap-dependent translational activation of snail

PubMed Central

She, Qing-Bai

2014-01-01

The cap-dependent translation is frequently deregulated in a variety of cancers associated with tumor progression. However, the molecular basis of the translation activation for metastatic progression of cancer remains largely elusive. Here, we demonstrate that activation of cap-dependent translation by silencing the translational repressor 4E-BP1 causes cancer epithelial cells to undergo epithelial-mesenchymal transition (EMT), which is associated with selective upregulation of the EMT inducer Snail followed by repression of E-cadherin expression and promotion of cell migratory and invasive capabilities as well as metastasis. Conversely, inhibition of cap-dependent translation by a dominant active mutant 4E-BP1 effectively downregulates Snail expression and suppresses cell migration and invasion. Furthermore, dephosphorylation of 4E-BP1 by mTORC1 inhibition or directly targeting the translation initiation also profoundly attenuates Snail expression and cell motility, whereas knockdown of 4E-BP1 or overexpression of Snail significantly rescues the inhibitory effects. Importantly, 4E-BP1-regulated Snail expression is not associated with its changes in the level of transcription or protein stability. Together, these findings indicate a novel role of 4E-BP1 in the regulation of EMT and cell motility through translational control of Snail expression and activity, and suggest that targeting cap-dependent translation may provide a promising approach for blocking Snail-mediated metastatic potential of cancer. PMID:24970798

Epsilon-aminocaproic acid prevents high glucose and insulin induced-invasiveness in MDA-MB-231 breast cancer cells, modulating the plasminogen activator system.

PubMed

Viedma-Rodríguez, Rubí; Martínez-Hernández, María Guadalupe; Flores-López, Luis Antonio; Baiza-Gutman, Luis Arturo

2018-01-01

Obesity and type II diabetes mellitus have contributed to the increase of breast cancer incidence worldwide. High glucose concentration promotes the proliferation of metastatic cells, favoring the activation of the plasminogen/plasmin system, thus contributing to tumor progression. The efficient formation of plasmin is dependent on the binding of plasminogen to the cell surface. We studied the effect of ε-aminocaproic acid (EACA), an inhibitor of the binding of plasminogen to cell surface, on proliferation, migration, invasion, epithelial-mesenchymal transition (EMT), and plasminogen activation system, in metastatic MDA-MB-231 breast cancer cells grown in a high glucose microenvironment and treated with insulin. MDA-MB-231 cells were treated with EACA 12.5 mmol/L under high glucose 30 mmol/L (HG) and high glucose and insulin 80 nmol/L (HG-I) conditions, evaluating: cell population growth, % of viability, migratory, and invasive abilities, as well as the expression of uPA, its receptor (uPAR), and its inhibitor (PAI-1), by real-time reverse transcription-polymerase chain reaction (RT-PCR) and Western blot, MMP-2 and MMP-9 mRNAs were evaluated by RT-PCR. Markers of EMT were evaluated by Western blot. Additionally, the presence of active uPA was studied by gel zymography, using casein-plasminogen as substrates. EACA prevented the increase in cell population, migration and invasion induced by HG and insulin, which was associated with the inhibition of EMT and the attenuation of HG- and insulin-dependent expression of uPA, uPAR, PAI-1, MMP-2, MMP-9, α-enolase (ENO A), and HCAM. The interaction of plasminogen to the cell surface and plasmin formation are mediators of the prometastasic action of hyperglycemia and insulin, potentially, EACA can be employed in the prevention and as adjuvant treatment of breast tumorigenesis promoted by hyperglycemia and insulin.

Hypothalamic beta-endorphin neurons suppress preneoplastic and neoplastic lesions development in 1,2-dimethylhydrazine induced rat colon cancer model.

PubMed

Murugan, Sengottuvelan; Dave, Yatee; Rakhit, Ankush; Sarkar, Dipak K

2017-01-01

In recent years, experimental studies demonstrated negative impacts of impaired body stress response on colonic pathologies. In this study, we tested if reducing body stress response by the use of β-endorphin (BEP) neuronal transplants in the hypothalamus suppresses pre-neoplastic and neoplastic lesions. Colon cancer was induced by injecting 1,2-dimethylhydrazine (DMH) for sixteen weeks in Sprague Dawley rats with BEP neuron transplants or control neuron transplants, and their colonic histopathologies, colon tissue levels of pro-inflammatory cytokines and epithelial-mesenchymal transition (EMT) proteins and splenic levels of cytotoxic proteins were measured. Our results revealed that DMH induced tumors in colon at 100% incidence in control rats but failed to induce colonic tumors in 70% of animal with BEP neuronal transplants. The mean volume of tumor at the colon was smaller in BEP neurons transplanted rats than those in controls. Histopathologies of colon tissues revealed that BEP neurons transplanted animals had lesser tissue lesions such as aberrant crypt foci (ACF) and adenocarcinoma development in the colon than those in control groups. Immunohistochemical and western blot analyses identified reduced expression of Ki-67, TNF-α and NF-κB nuclear translocation in colonic tissues of BEP neurons transplanted rats than those in controls. BEP neurons transplanted rats also showed reduced expressions of transcription factors linked to EMT like Snail, Twist, and N-cadherin, but increased the levels of an epithelial cell marker E-cadherin in colon tissue. Furthermore, splenic NK cells cytolytic proteins such as perforin, granzyme B and IFN-γ levels in BEP neurons transplanted rats were higher than those in control rats. These data suggest that BEP neuron transplants suppress the growth and progression of colonic tumors possibly by decreasing inflammatory mileu and EMT via activation of innate immune responses.

Doxycycline reverses epithelial-to-mesenchymal transition and suppresses the proliferation and metastasis of lung cancer cells.

PubMed

Qin, Yuan; Zhang, Qiang; Lee, Shan; Zhong, Wei-Long; Liu, Yan-Rong; Liu, Hui-Juan; Zhao, Dong; Chen, Shuang; Xiao, Ting; Meng, Jing; Jing, Xue-Shuang; Wang, Jing; Sun, Bo; Dai, Ting-Ting; Yang, Cheng; Sun, Tao; Zhou, Hong-Gang

2015-12-01

The gelatinase inhibitor doxycycline is the prototypical antitumor antibiotic. We investigated the effects of doxycycline on the migration, invasion, and metastasis of human lung cancer cell lines and in a mouse model. We also measured the effect of doxycycline on the transcription of epithelial-mesenchymal transition (EMT) markers, and used immunohistochemistry to determine whether EMT reversal was associated with doxycycline inhibition. Doxycycline dose-dependently inhibited proliferation, migration, and invasion of NCI-H446 human small cell lung cancer cells. It also suppressed tumor growth from NCI-H446 and A549 lung cancer cell xenografts without altering body weight, inhibited Lewis lung carcinoma cell migration, and prolonged survival. The activities of the transcription factors Twist1/2, SNAI1/2, AP1, NF-κB, and Stat3 were suppressed by doxycycline, which reversed EMT and inhibited signal transduction, thereby suppressing tumor growth and metastasis. Our data demonstrate functional targeting of transcription factors by doxycycline to reverse EMT and suppress tumor proliferation and metastasis. Thus, doxycycline selectively targets malignant tumors and reduces its metastatic potential with less cytotoxicity in lung cancer patients.

Silibinin suppresses EMT-driven erlotinib resistance by reversing the high miR-21/low miR-200c signature in vivo

PubMed Central

Cufí, Sílvia; Bonavia, Rosa; Vazquez-Martin, Alejandro; Oliveras-Ferraros, Cristina; Corominas-Faja, Bruna; Cuyàs, Elisabet; Martin-Castillo, Begoña; Barrajón-Catalán, Enrique; Visa, Joana; Segura-Carretero, Antonio; Joven, Jorge; Bosch-Barrera, Joaquim; Micol, Vicente; Menendez, Javier A.

2013-01-01

The flavolignan silibinin was studied for its ability to restore drug sensitivity to EGFR-mutant NSCLC xenografts with epithelial-to-mesenchymal transition (EMT)-driven resistance to erlotinib. As a single agent, silibinin significantly decreased the tumor volumes of erlotinib-refractory NSCLC xenografts by approximately 50%. Furthermore, the complete abrogation of tumor growth was observed with the co-treatment of erlotinib and silibinin. Silibinin fully reversed the EMT-related high miR-21/low miR-200c microRNA signature and repressed the mesenchymal markers SNAIL, ZEB, and N-cadherin observed in erlotinib-refractory tumors. Silibinin was sufficient to fully activate a reciprocal mesenchymal-to-epithelial transition (MET) in erlotinib-refractory cells and prevent the highly migratogenic phenotype of erlotinib-resistant NSCLC cells. Given that the various mechanisms of resistance to erlotinib result from EMT, regardless of the EGFR mutation status, a water-soluble, silibinin-rich milk thistle extract might be a suitable candidate therapy for upcoming clinical trials aimed at preventing or reversing NSCLC progression following erlotinib treatment. PMID:23963283

DOE Office of Scientific and Technical Information (OSTI.GOV)

Castillo, Gaelle del; Murillo, Miguel M.; IDIBELL-Institut de Recerca Oncologica, Gran Via s/n, Km 2.7, 08907 L'Hospitalet, Barcelona

Transforming growth factor-beta (TGF-{beta}) induces apoptosis in fetal rat hepatocytes. However, a subpopulation of these cells survives, concomitant with changes in phenotype, reminiscent of an epithelial-mesenchymal transition (EMT). We have previously suggested that EMT might confer cell resistance to apoptosis (Valdes et al., Mol. Cancer Res., 1: 68-78, 2002). However, the molecular mechanisms responsible for this resistance are not explored yet. In this work, we have isolated and subcultured the population of hepatocytes that suffered the EMT process and are resistant to apoptosis (TGF-{beta}-treated fetal hepatocytes: T{beta}T-FH). We prove that they secrete mitogenic and survival factors, as analyzed by themore » proliferative and survival capacity of conditioned medium. Inhibition of the epidermal growth factor receptor (EGFR) sensitizes T{beta}T-FH to die after serum withdrawal. T{beta}T-FH expresses high levels of transforming growth factor-alpha (TGF-{alpha}) and heparin-binding EGF-like growth factor (HB-EGF) and shows constitutive activation of the EGFR pathway. A blocking anti-TGF-{alpha} antibody restores the capacity of cells to die. TGF-{beta}, which is expressed by T{beta}T-FH, mediates up-regulation of TGF-{alpha} and HB-EGF expression in those cells. In summary, results suggest that an autocrine loop of TGF-{beta} confers resistance to apoptosis after an EMT process in hepatocytes, through the increase in the expression of EGFR ligands.« less

Escin Ia suppresses the metastasis of triple-negative breast cancer by inhibiting epithelial-mesenchymal transition via down-regulating LOXL2 expression.

PubMed

Wang, Yuhui; Xu, Xiaotian; Zhao, Peng; Tong, Bei; Wei, Zhifeng; Dai, Yue

2016-04-26

The saponin fraction of Aesculus chinensis Bunge fruits (SFAC) could inhibit the invasion and migration of MDA-MB-231 cells. Among which, escin Ia showed more potent inhibition of the invasion than other five main saponin constituents. It selectively reduced the expression of LOXL2 mRNA and promoted the expression of E-cadherin mRNA, and prevented the EMT process of MDA-MB-231 cells and TNF-α/TGF-β-stimulated MCF-7 cells. Moreover, it reduced the LOXL2 level in MDA-MB-231 cells but not in MCF-7 cells. When MCF-7 cells were stimulated with TNF-α/TGF-β, transfected with LOXL2 or treated with hypoxia, escin Ia down-regulated the level of LOXL2 in MCF-7 cells. Meanwhile, escin Ia suppressed the EMT process in LOXL2-transfected or hypoxia-treated MCF-7 cells. Of interest, escin Ia did not alter the level of HIF-1α in hypoxia-induced MCF-7 cells. In TNBC xenograft mice, the metastasis and EMT of MDA-MB-231 cells were suppressed by escin Ia. In conclusion, escin Ia was the main active ingredient of SFAC for the anti-TNBC metastasis activity, and its action mechanisms involved inhibition of EMT process by down-regulating LOXL2 expression.

Escin Ia suppresses the metastasis of triple-negative breast cancer by inhibiting epithelial-mesenchymal transition via down-regulating LOXL2 expression

PubMed Central

Zhao, Peng; Tong, Bei; Wei, Zhifeng; Dai, Yue

2016-01-01

The saponin fraction of Aesculus chinensis Bunge fruits (SFAC) could inhibit the invasion and migration of MDA-MB-231 cells. Among which, escin Ia showed more potent inhibition of the invasion than other five main saponin constituents. It selectively reduced the expression of LOXL2 mRNA and promoted the expression of E-cadherin mRNA, and prevented the EMT process of MDA-MB-231 cells and TNF-α/TGF-β-stimulated MCF-7 cells. Moreover, it reduced the LOXL2 level in MDA-MB-231 cells but not in MCF-7 cells. When MCF-7 cells were stimulated with TNF-α/TGF-β, transfected with LOXL2 or treated with hypoxia, escin Ia down-regulated the level of LOXL2 in MCF-7 cells. Meanwhile, escin Ia suppressed the EMT process in LOXL2-transfected or hypoxia-treated MCF-7 cells. Of interest, escin Ia did not alter the level of HIF-1α in hypoxia-induced MCF-7 cells. In TNBC xenograft mice, the metastasis and EMT of MDA-MB-231 cells were suppressed by escin Ia. In conclusion, escin Ia was the main active ingredient of SFAC for the anti-TNBC metastasis activity, and its action mechanisms involved inhibition of EMT process by down-regulating LOXL2 expression. PMID:27008697

Post-transcription mediated Snail stabilization is involved in radiation exposure induced invasion and migration of hepatocarcinoma cells.

PubMed

Dong, Liyang; Zhang, Xuebang; Xiang, Wei; Ni, Junwei; Zhou, Weizhong; Li, Haiyan

2018-04-20

Increasing evidences suggested that radiotherapy can paradoxically promote tumor invasion and metastatic processes, while its detailed mechanism is not well illustrated. Our present study found that radiation can promote the migration and invasion of hepatocellular carcinoma (HCC) cells via induction of epithelial mesenchymal transition (EMT), which was evidenced by the results that radiation induced up regulation of vimentin while down regulation of E-Cadherin. As to the EMT-related transcription factors, radiation increased the expression of Snail, while not Slug, ZEB1 or TWIST. This was confirmed by the results that radiation increased the nuclear translocation of Snail in HCC cells. However, radiation had no effect on the expression or half-life of Snail mRNA. In HCC cells treated by cycloheximide (CHX, the translation inhibitor), radiation significantly increased the half-life of Snail protein, which suggested that radiation increased the expression of Snail via up regulation of its protein stability. Radiation increased the expression of COP9 signalosome 2 (CSN2), which has been reported to block the ubiquitination and degradation of Snail. Silence of CSN2/Snail can attenuate radiation induced cell migration and EMT of HCC cells. Collectively, our data suggested that radiation can promote HCC cell invasion and EMT by stabilization of Snail via CSN2 signals. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

ΔNp63α promotes breast cancer cell motility through the selective activation of components of the epithelial-to-mesenchymal transition program

PubMed Central

Dang, Tuyen T.; Esparza, Matthew A.; Maine, Erin A.; Westcott, Jill M.; Pearson, Gray W.

2015-01-01

Cell identity signals influence the invasive capability of tumor cells, as demonstrated by the selection for programs of epithelial-to-mesenchymal transition (EMT) during malignant progression. Breast cancer cells retain canonical epithelial traits and invade collectively as cohesive groups of cells, but the signaling pathways critical to their invasive capabilities are still incompletely understood. Here we report that the transcription factor ΔNp63α drives the migration of basal-like breast cancer (BLBC) cells by inducing a hybrid mesenchymal/epithelial state. Through a combination of expression analysis and functional testing across multiple BLBC cell populations, we determined that ΔNp63α induces migration by elevating the expression of the EMT program components Slug and Axl. Interestingly, ΔNp63α also increased the expression of miR205, which can silence ZEB1/2 to prevent the loss of epithelial character caused by EMT induction. In clinical specimens, co-expression of various elements of the ΔNp63α pathway confirmed its implication in motility signaling in BLBC. We observed that activation of the ΔNp63α pathway occurred during the transition from noninvasive ductal carcinoma in situ to invasive breast cancer. Notably, in an orthotopic tumor model, Slug expression was sufficient to induce collective invasion of E-cadherin expressing BLBC cells. Together, our results illustrate how ΔNp63α can drive breast cancer cell invasion by selectively engaging pro-migratory components of the EMT program while, in parallel, still promoting the retention of epithelial character. PMID:26292362

Thrombospondin 1 promotes an aggressive phenotype through epithelial-to-mesenchymal transition in human melanoma.

PubMed

Jayachandran, Aparna; Anaka, Matthew; Prithviraj, Prashanth; Hudson, Christopher; McKeown, Sonja J; Lo, Pu-Han; Vella, Laura J; Goding, Colin R; Cebon, Jonathan; Behren, Andreas

2014-07-30

Epithelial-to-mesenchymal transition (EMT), in which epithelial cells loose their polarity and become motile mesenchymal cells, is a determinant of melanoma metastasis. We compared gene expression signatures of mesenchymal-like melanoma cells with those of epithelial-like melanoma cells, and identified Thrombospondin 1 (THBS1) as highly up-regulated in the mesenchymal phenotype. This study investigated whether THBS1, a major physiological activator of transforming growth factor (TGF)-beta, is involved in melanoma EMT-like process. We sought to examine expression patterns in distinct melanoma phenotypes including invasive, de-differentiated, label-retaining and drug resistant populations that are putatively associated with an EMT-like process. Here we show that THBS1 expression and secretion was elevated in melanoma cells exhibiting invasive, drug resistant, label retaining and mesenchymal phenotypes and correlated with reduced expression of genes involved in pigmentation. Elevated THBS1 levels were detected in Vemurafenib resistant melanoma cells and inhibition of THBS1 led to significantly reduced chemoresistance in melanoma cells. Notably, siRNA-mediated silencing of THBS1 and neutralizing antibody to THBS1 reduced invasion in mesenchymal-like melanoma cells, while ectopic THBS1 expression in epithelial-like melanoma cells enhanced invasion. Furthermore, the loss of THBS1 inhibited in vivo motility of melanoma cells within the embryonic chicken neural tube. In addition, we found aberrant THBS1 protein expression in metastatic melanoma tumor biopsies. These results implicate a role for THBS1 in EMT, and hence THBS1 may serve as a novel target for strategies aimed at the treatment of melanoma invasion and drug resistance.

Thrombospondin 1 promotes an aggressive phenotype through epithelial-to-mesenchymal transition in human melanoma

PubMed Central

Jayachandran, Aparna; Anaka, Matthew; Prithviraj, Prashanth; Hudson, Christopher; McKeown, Sonja J; Lo, Pu-Han; Vella, Laura J; Goding, Colin R; Cebon, Jonathan; Behren, Andreas

2014-01-01

Epithelial-to-mesenchymal transition (EMT), in which epithelial cells loose their polarity and become motile mesenchymal cells, is a determinant of melanoma metastasis. We compared gene expression signatures of mesenchymal-like melanoma cells with those of epithelial-like melanoma cells, and identified Thrombospondin 1 (THBS1) as highly up-regulated in the mesenchymal phenotype. This study investigated whether THBS1, a major physiological activator of transforming growth factor (TGF)-beta, is involved in melanoma EMT-like process. We sought to examine expression patterns in distinct melanoma phenotypes including invasive, de-differentiated, label-retaining and drug resistant populations that are putatively associated with an EMT-like process. Here we show that THBS1 expression and secretion was elevated in melanoma cells exhibiting invasive, drug resistant, label retaining and mesenchymal phenotypes and correlated with reduced expression of genes involved in pigmentation. Elevated THBS1 levels were detected in Vemurafenib resistant melanoma cells and inhibition of THBS1 led to significantly reduced chemoresistance in melanoma cells. Notably, siRNA-mediated silencing of THBS1 and neutralizing antibody to THBS1 reduced invasion in mesenchymal-like melanoma cells, while ectopic THBS1 expression in epithelial-like melanoma cells enhanced invasion. Furthermore, the loss of THBS1 inhibited in vivo motility of melanoma cells within the embryonic chicken neural tube. In addition, we found aberrant THBS1 protein expression in metastatic melanoma tumor biopsies. These results implicate a role for THBS1 in EMT, and hence THBS1 may serve as a novel target for strategies aimed at the treatment of melanoma invasion and drug resistance. PMID:25051363

Irradiated fibroblasts promote epithelial–mesenchymal transition and HDGF expression of esophageal squamous cell carcinoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bao, Ci-Hang; Wang, Xin-Tong; Ma, Wei

2015-03-06

Recent evidence suggested that nonirradiated cancer-associated fibroblasts (CAFs) promoted aggressive phenotypes of cancer cells through epithelial–mesenchymal transition (EMT). Hepatoma-derived growth factor (HDGF) is a radiosensitive gene of esophageal squamous cell carcinoma (ESCC). This study aimed to investigate the effect of irradiated fibroblasts on EMT and HDGF expression of ESCC. Our study demonstrated that coculture with nonirradiated fibroblasts significantly increased the invasive ability of ESCC cells and the increased invasiveness was further accelerated when they were cocultured with irradiated fibroblasts. Scattering of ESCC cells was also accelerated by the supernatant from irradiated fibroblasts. Exposure of ESCC cells to supernatant from irradiatedmore » fibroblasts resulted in decreased E-cadherin, increased vimentin in vitro and β-catenin was demonstrated to localize to the nucleus in tumor cells with irradiated fibroblasts in vivo models. The expression of HDGF and β-catenin were increased in both fibroblasts and ESCC cells of irradiated group in vitro and in vivo models. Interestingly, the tumor cells adjoining the stromal fibroblasts displayed strong nuclear HDGF immunoreactivity, which suggested the occurrence of a paracrine effect of fibroblasts on HDGF expression. These data suggested that irradiated fibroblasts promoted invasion, growth, EMT and HDGF expression of ESCC. - Highlights: • Irradiated CAFs accelerated invasiveness and scattering of ESCC cell lines. • Irradiated CAFs promoted EMT of ESCC cells. • Irradiated fibroblasts induced nuclear β-catenin relocalization in ESCC cells. • Irradiated fibroblasts increased HDGF expression in vitro and in vivo.« less

Epithelial-mesenchymal transition abolishes the susceptibility of polarized epithelial cell lines to measles virus.

PubMed

Shirogane, Yuta; Takeda, Makoto; Tahara, Maino; Ikegame, Satoshi; Nakamura, Takanori; Yanagi, Yusuke

2010-07-02

Measles virus (MV), an enveloped negative-strand RNA virus, remains a major cause of morbidity and mortality in developing countries. MV predominantly infects immune cells by using signaling lymphocyte activation molecule (SLAM; also called CD150) as a receptor, but it also infects polarized epithelial cells, forming tight junctions in a SLAM-independent manner. Although the ability of MV to infect polarized epithelial cells is thought to be important for its transmission, the epithelial cell receptor for MV has not been identified. A transcriptional repressor, Snail, induces epithelial-mesenchymal transition (EMT), in which epithelial cells lose epithelial cell phenotypes, such as adherens and tight junctions. In this study, EMT was induced by expressing Snail in a lung adenocarcinoma cell line, II-18, which is highly susceptible to wild-type MV. Snail-expressing II-18 cells lost adherens and tight junctions. Microarray analysis confirmed the induction of EMT in II-18 cells and suggested a novel function of Snail in protein degradation and distribution. Importantly, wild-type MV no longer entered EMT-induced II-18 cells, suggesting that the epithelial cell receptor is down-regulated by the induction of EMT. Other polarized cell lines, NCI-H358 and HT-29, also lost susceptibility to wild-type MV when EMT was induced. However, the complete formation of tight junctions rather reduced MV entry into HT-29 cells. Taken together, these data suggest that the unidentified epithelial cell receptor for MV is involved in the formation of epithelial intercellular junctions.

Analyses of publicly available genomics resources define FGF-2-expressing bladder carcinomas as EMT-prone, proliferative tumors with low mutation rates and high expression of CTLA-4, PD-1 and PD-L1

PubMed Central

McNiel, Elizabeth A; Tsichlis, Philip N

2017-01-01

Fibroblast growth factor 2 (FGF-2) is overexpressed in a subset of invasive bladder carcinomas and its overexpression correlates with poor prognosis. Analyses of publicly available databases addressing the molecular mechanisms that may be responsible for the poor prognosis of these tumors, revealed that FGF-2 expression correlates positively with the expression of epithelial to mesenchymal transition (EMT)-promoting transcription factors and with changes in gene expression that are characteristic of EMT. The same analyses also revealed that FGF-2 correlates negatively with the expression, mutation and copy number variations of FGFR-3, all of which are associated with noninvasive bladder carcinomas. Finally, they showed that FGF-2 expression correlates with the expression of FGFR-1, the expression of the IIIc variant of FGFR-2 and with the expression of Akt3. The latter observation is significant because our earlier studies had shown that Akt3 regulates FGFR-2 alternative splicing, shifting the balance toward the IIIc relative to the IIIb FGFR-2 splice variant. As the IIIc variant is recognized by FGF-2, while the IIIb variant is not, we conclude that Akt3 may facilitate the FGF-2 response. FGF-2 is known to promote the expression of KDM2B, which functions in concert with EZH2 to repress the EZH2-targeting microRNA miR-101, activating a switch, which stably upregulates EZH2. The cancer genome atlas (TCGA) data showing a correlation between KDM2B and EZH2 expression and Oncomine data, showing a correlation between KDM2B and tumor progression, strongly support the role of the FGF-2/KDM2B/miR-101/EZH2 pathway in bladder cancer. These observations combined, suggest a model according to which FGF-2 induces EMT, cell proliferation and cancer stem cell self-renewal by coupling the Akt3 and KDM2B-controlled pathways outlined above, in bladder carcinomas. Further analyses of publicly available databases, revealed that FGF-2-expressing bladder carcinomas carry fewer genetic alterations and they tend to express high levels of CTLA-4, PD-1 and PD-L1, which suggests immune blockade by checkpoint activation. EMT, enhanced proliferation and immune checkpoint activation combined, may be responsible for the poor prognosis of FGF-2-expressing bladder carcinomas. PMID:28515962

Nanotechnology for the detection and kill of circulating tumor cells

NASA Astrophysics Data System (ADS)

Gao, Yang; Yuan, Zhou

2014-09-01

Circulating tumor cells (CTCs) represent a surrogate biomarker of hematogenous metastases and thus could be considered as a `liquid biopsy' which reveals metastasis in action. But it is absolutely a challenge to detect CTCs due to their extreme rarity. At present, the most common principle is to take advantage of the epithelial surface markers of CTCs which attach to a specific antibody. Antibody-magnetic nanobeads combine with the epithelial surface markers, and then the compound is processed by washing, separation, and detection. However, a proportion of CTC antigen expressions are down-regulated or lost in the process of epithelial-mesenchymal transition (EMT), and thus, this part of CTCs cannot be detected by classical detection methods such as CellSearch. To resolve this problem, some multiple-marker CTC detections have been developed rapidly. Additionally, nanotechnology is a promising approach to kill CTCs with high efficiency. Implantable nanotubes coated with apoptosis-promoting molecules improve the disease-free survival and overall survival. The review introduces some novel CTC detection techniques and therapeutic methods by virtue of nanotechnology to provide a better knowledge of the progress about CTC study.

Upregulation of MicroRNA-4262 Targets Kaiso (ZBTB33) to Inhibit the Proliferation and EMT of Cervical Cancer Cells.

PubMed

Feng, Jing

2017-08-11

More and more studies have reported that dysregulation of microRNAs (miRNAs) lead to the proliferation and EMT of multiple cancers. Recently, several reports have demonstrated that dysregulation of miR-4262 is in numerous cancers. However, its role and precise mechanism in human cervical cancer (CC) have not been well clarified. Hence, my study was aim to explore the biological roles and precise mechanisms of miR-4262 in CC cell lines. In my study, I found that the level of miR-4262 is significantly decreased in CC tissues and cell lines. Moreover, decreased expression of miR-4262 was closely related to increased expression of Kaiso (ZBTB33) that belongs to the BTB/POZ family in CC tissues and cell lines. The proliferation and EMT of CC cells were inhibited by miR-4262 mimic. However, down-regulation of miR-4262 enhanced the proliferation and EMT of CC cells. Next, bioinformatics analysis predicted that miR-4262 might directly target the Kaiso gene. Besides, luciferase reporter assay had confirmed this result. Moreover, introduction of Kaiso in CC cells partially blocked the effects of miR-4262 mimic. In conclusion, miR-4262 suppressed the proliferation and EMT of CC cells by directly down-regulation of Kaiso.

Effect of midkine on gemcitabine resistance in biliary tract cancer

PubMed Central

Guo, Huihui; Qiu, Li; Sun, Xinrong; Wang, Xiang; Shi, Qian

2018-01-01

Gemcitabine-based chemotherapy is one of the most effective and commonly used chemotherapeutic regimens for biliary tract cancer (BTC). However, development of resistance to this drug limits its efficacy. The present study aimed to explore the effects of midkine (MDK) on the resistance of BTC cells to gemcitabine. Cell viability and proliferation were measured by a Cell Counting Kit-8 assay and 5-ethynyl-2′-deoxyuridine staining, respectively. Western blot analysis was used to detect the expression of E-cadherin and vimentin. The results indicated that BTC cell lines were more resistant to gemcitabine plus MDK compared with gemcitabine alone. In terms of the underlying mechanism, MDK promoted the epithelial to mesenchymal transition (EMT) of BTC cells and the enhancing effect of MDK on gemcitabine resistance was abrogated when the EMT was blocked with small interfering (si)RNA targeting Twist. In addition, MDK promoted the expression of Notch-1, while knockdown of Notch-1 by siRNA blocked the EMT process in the BTC cell lines. Taken together, these results indicated that MDK promoted gemcitabine resistance of BTC through inducing EMT via upregulating Notch-1. It was suggested that inhibition of the EMT is a promising strategy to overcome MDK-induced drug resistance. PMID:29344648

Differential roles of kallikrein-related peptidase 6 in malignant transformation and ΔNp63β-mediated epithelial-mesenchymal transition of oral squamous cell carcinoma.

PubMed

Kaneko, Naoki; Kawano, Shintaro; Yasuda, Kaori; Hashiguchi, Yuma; Sakamoto, Taiki; Matsubara, Ryota; Goto, Yuichi; Jinno, Teppei; Maruse, Yasuyuki; Morioka, Masahiko; Hattori, Taichi; Tanaka, Shoichi; Tanaka, Hideaki; Kiyoshima, Tamotsu; Nakamura, Seiji

2017-12-01

We previously reported that epithelial-to-mesenchymal transition (EMT) was mediated by ΔNp63β in oral squamous cell carcinoma (OSCC). In this study, DNA microarray analyses were performed using ΔNp63β-overexpressing OSCC cells to identify genes associated with ΔNp63β-mediated EMT. Thereby, we focused on kallikrein-related peptidase (KLK) 6, most up-regulated following ΔNp63β-overexpression, that activates protease-activated receptors (PARs). In RT-PCR analyses, ΔNp63 was positively associated with KLK6 and PAR2 and negatively with PAR1 in OSCC cells. By ΔNp63 knockdown, KLK6 and PAR2 expression was decreased and PAR1 was increased. Furthermore, KLK6 knockdown led to enhancing migration and invasion, and inhibiting proliferation, suggesting EMT-phenotypes. Although, in the KLK6 or PAR2 knockdown cells, phosphorylation of ERK was reduced, it was restored in the KLK6 knockdown OSCC cells treated with recombinant KLK6 proteins. Immunohistochemistry showed ΔNp63, KLK6, and PAR2 were more strongly expressed in the epithelial dysplasia and central region of OSCC than normal oral epithelium, whereas PAR1 expression was undetectable. Interestingly, at the invasive front of OSCC, ΔNp63, KLK6, and PAR2 were reduced, but PAR1 was elevated. In addition, the OSCC patients with decreasing KLK6 expression at the invasive front had more unfavourable prognosis. These results suggested differential roles of KLK6 in malignant transformation and EMT; high ΔNp63β expression up-regulates KLK6-PAR2 and down-regulates PAR1, inducing malignant transformation in oral epithelium with stimulating proliferation through ERK signal activation. Moreover, KLK6-PAR2 expression is down-regulated and PAR1 is up-regulated when ΔNp63β expression is decreased, leading to EMT with enhancing migration and invasion through ERK signal reduction at the invasive front. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Engagement of I-Branching β-1, 6-N-Acetylglucosaminyltransferase 2 in Breast Cancer Metastasis and TGF-β Signaling

PubMed Central

Zhang, Haijun; Meng, Fanyan; Wu, Sherwin; Kreike, Bas; Sethi, Seema; Chen, Wei; Miller, Fred R.; Wu, Guojun

2014-01-01

In this study, we have showed that GCNT2, a gene-encoding glucosaminyl (N-acetyl) transferase 2, I-branching enzyme, is overexpressed in highly metastatic breast cancer cell lines of human and mouse origin and basal-like breast tumor samples. GCNT2 expression is also significantly correlated to the metastatic phenotype in breast tumor samples. Functional studies showed that ectopic expression of GCNT2 enhances cell detachment, adhesion to endothelial cells, cell migration and invasion in vitro, and lung metastasis of breast cancer cells in vivo. Knockdown of GCNT2 expression decreases cell migration and invasion in vitro and lung metastasis in vivo. We have further shown the involvement of GCNT2 in the epithelial-to-mesenchymal transition (EMT). Specifically, the expression of E-cadherin is significantly changed upon GCNT2 expression at the protein level but not at the RNA level. Moreover, we have shown that GCNT2 is a direct target of the TGF-β–smad pathway and that change in GCNT2 expression modulates EMT induced by TGF-β1 treatment. Finally, we have shown that diminution of the glycosyltransferase activity of I-branching β-1, 6-N-acetylglucosaminyl transferase 2 (GCNT2) abrogates its cell migration and invasion-promoting function and synergistic effect with TGF-β to induce EMT. Our study for the first time showed that GCNT2 is a novel gene contributing to breast cancer metastasis with preferential expression in basal-like breast cancer. Moreover, we discovered that involvement of GCNT2 in EMT and TGF-β signaling, and further glycosylation modification of E-cadherin by GCNT2, are the underlying integrative mechanisms for breast cancer metastasis, implying that blocking TGF-β/GCNT2 signaling is a promising approach for targeting metastatic breast cancer. PMID:21750175

MicroRNA-144-3p suppresses gastric cancer progression by inhibiting epithelial-to-mesenchymal transition through targeting PBX3

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Butian; Zhang, Shengping; Shen, Hao

MicroRNAs are aberrantly expressed in a wide variety of human cancers. The present study aims to elucidate the effects and molecular mechanisms of miR-144-3p that underlie gastric cancer (GC) development. It was observed that miR-144-3p expression was significantly decreased in GC tissues compared to that in paired non-tumor tissues; moreover, its expression was lower in tissues of advanced stage and larger tumor size, as well as in lymph node metastasis tissues compared to that in control groups. miR-144-3p expression was associated with depth of invasion (P = 0.030), tumor size (P = 0.047), lymph node metastasis (P = 0.047), and TNM stage (P = 0.048). Additionally, miR-144-3p significantlymore » inhibited proliferation, migration, and invasion in GC cells. It also reduced F-actin expression and suppressed epithelial-to-mesenchymal transition (EMT) in GC cells. Furthermore, pre-leukemia transcription factor 3 (PBX3) was a direct target gene of miR-144-3p. PBX3 was overexpressed in GC tissues and promoted EMT in GC cells. The effects of miR-144-3p mimics or inhibitors on cell migration, invasion, and proliferation were reversed by PBX3 overexpression or downregulation respectively. These results suggest that miR-144-3p suppresses GC progression by inhibiting EMT through targeting PBX3. - Highlights: • miR-144-3p is downregulated in gastric cancer tissues and associated with malignant clinical factors. • miR-144-3p inhibits proliferation, migration, and invasion in gastric cancer cells. • PBX3 is a direct target of miR-144-3p and promotes EMT in gastric cancer. • miR-144-3p suppresses EMT in gastric cancer by regulating PBX3.« less

Role of DNA methylation in miR-200c/141 cluster silencing in invasive breast cancer cells.

PubMed

Neves, Rui; Scheel, Christina; Weinhold, Sandra; Honisch, Ellen; Iwaniuk, Katharina M; Trompeter, Hans-Ingo; Niederacher, Dieter; Wernet, Peter; Santourlidis, Simeon; Uhrberg, Markus

2010-08-03

The miR-200c/141 cluster has recently been implicated in the epithelial to mesenchymal transition (EMT) process. The expression of these two miRNAs is inversely correlated with tumorigenicity and invasiveness in several human cancers. The role of these miRNAs in cancer progression is based in part on their capacity to target the EMT activators ZEB1 and ZEB2, two transcription factors, which in turn repress expression of E-cadherin. Little is known about the regulation of the mir200c/141 cluster, whose targeting has been proposed as a promising new therapy for the most aggressive tumors. We show that the miR-200c/141 cluster is repressed by DNA methylation of a CpG island located in the promoter region of these miRNAs. Whereas in vitro methylation of the miR-200c/141 promoter led to shutdown of promoter activity, treatment with a demethylating agent caused transcriptional reactivation in breast cancer cells formerly lacking expression of miR-200c and miR-141. More importantly, we observed that DNA methylation of the identified miR-200c/141 promoter was tightly correlated with phenotype and the invasive capacity in a panel of 8 human breast cancer cell lines. In line with this, in vitro induction of EMT by ectopic expression of the EMT transcription factor Twist in human immortalized mammary epithelial cells (HMLE) was accompanied by increased DNA methylation and concomitant repression of the miR-200c/141 locus. The present study demonstrates that expression of the miR-200c/141 cluster is regulated by DNA methylation, suggesting epigenetic regulation of this miRNA locus in aggressive breast cancer cell lines as well as untransformed mammary epithelial cells. This epigenetic silencing mechanism might represent a novel component of the regulatory circuit for the maintenance of EMT programs in cancer and normal cells.

The inhibition of lung cancer cell migration by AhR-regulated autophagy.

PubMed

Tsai, Chi-Hao; Li, Ching-Hao; Cheng, Yu-Wen; Lee, Chen-Chen; Liao, Po-Lin; Lin, Cheng-Hui; Huang, Shih-Hsuan; Kang, Jaw-Jou

2017-02-14

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is highly expressed in multiple organs and tissues. Whereas AhR mediates the metabolism of xenobiotic and endogenous compounds, its novel function in cancer epithelial-mesenchymal transition (EMT) remains controversial. Autophagy also participates in tumour progression through its functions in cell homeostasis and facilitates adaptation to EMT progression. In the present study, we found that AhR-regulated autophagy positively modulates EMT in non-small cell lung cancer cells. The motility of A549, H1299, and CL1-5 cells were correlated with different AhR expression levels. Invasive potential and cell morphology also changed when AhR protein expression was altered. Moreover, AhR levels exerted a contrasting effect on autophagy potential. Autophagy was higher in CL1-5 and H1299 cells with lower AhR levels than in A549 cells. Both AhR overexpression and autophagy inhibition decreased CL1-5 metastasis in vivo. Furthermore, AhR promoted BNIP3 ubiquitination for proteasomal degradation. AhR silencing in A549 cells also reduced BNIP3 ubiquitination. Taken together, these results provide a novel insight into the cross-linking between AhR and autophagy, we addressed the mechanistic BNIP3 modulation by endogenous AhR, which affect cancer cell EMT progression.

Histochemical and Immunohistochemical Study of α-SMA, Collagen, and PCNA in Epithelial Ovarian Neoplasm

PubMed

Anggorowati, Nungki; Ratna Kurniasari, Chatarina; Damayanti, Karina; Cahyanti, Titik; Widodo, Irianiwati; Ghozali, Ahmad; Romi, Muhammad Mansyur; Sari, Dwi Cahyani Ratna; Arfian, Nur

2017-03-01

Background: Alpha-smooth muscle actin (α-SMA) is an isoform of actin, positive in myofibroblasts and is an epithelial to mesenchymal transition (EMT) marker. EMT is a process by which tumor cells develop to be more hostile and able to metastasize. Progression of tumor cells is always followed by cell composition and extracellular matrix component alteration. Increased α-SMA expression and collagen alteration may predict the progressivity of ovarian neoplasms. Objective: The aim of this research was to analyse the characteristic of α-SMA and collagen in tumor cells and stroma of ovarian neoplasms. In this study, PCNA (proliferating cell nuclear antigen) expression was also investigated. Methods: Thirty samples were collected including serous, mucinous, endometrioid, and clear cell subtypes. The expression of α-SMA and PCNA were calculated in cells and stroma of ovarian tumors. Collagen was detected using Sirius Red staining and presented as area fraction. Results: The overexpressions of α-SMA in tumor cells were only detected in serous and clear cell ovarian carcinoma. The histoscore of α-SMA was higher in malignant than in benign or borderline ovarian epithelial neoplasms (105.3±129.9 vs. 17.3±17.1, P=0.011; mean±SD). Oppositely, stromal α-SMA and collagen area fractions were higher in benign than in malignant tumors (27.2±6.6 vs 20.5±8.4, P=0.028; 31.0±5.6 vs. 23.7±6.4, P=0.04). The percentages of epithelial and stromal PCNA expressions were not significantly different between benign and malignant tumors. Conclusion: Tumor cells of serous and clear cell ovarian carcinoma exhibit mesenchymal characteristic as shown by α-SMA positive expression. This expression might indicate that these subtypes were more aggressive. This research showed that collagen and α-SMA area fractions in stroma were higher in benign than in malignant neoplasms. 10.22034/APJCP.2017.18.3.667

CLCA2 epigenetic regulation by CTBP1, HDACs, ZEB1, EP300 and miR-196b-5p impacts prostate cancer cell adhesion and EMT in metabolic syndrome disease.

PubMed

Porretti, Juliana; Dalton, Guillermo N; Massillo, Cintia; Scalise, Georgina D; Farré, Paula L; Elble, Randolph; Gerez, Esther N; Accialini, Paula; Cabanillas, Ana M; Gardner, Kevin; De Luca, Paola; De Siervi, Adriana

2018-03-14

Prostate cancer (PCa) is the most common cancer among men. Metabolic syndrome (MeS) is associated with increased PCa aggressiveness and recurrence. Previously, we proposed C-terminal binding protein 1 (CTBP1), a transcriptional co-repressor, as a molecular link between these two conditions. Notably, CTBP1 depletion decreased PCa growth in MeS mice. The aim of this study was to investigate the molecular mechanisms that explain the link between MeS and PCa mediated by CTBP1. We found that CTBP1 repressed chloride channel accessory 2 (CLCA2) expression in prostate xenografts developed in MeS animals. CTBP1 bound to CLCA2 promoter and repressed its transcription and promoter activity in PCa cell lines. Furthermore, we found that CTBP1 formed a repressor complex with ZEB1, EP300 and HDACs that modulates the CLCA2 promoter activity. CLCA2 promoted PCa cell adhesion inhibiting epithelial-mesenchymal transition (EMT) and activating CTNNB1 together with epithelial marker (CDH1) induction, and mesenchymal markers (SNAI2 and TWIST1) repression. Moreover, CLCA2 depletion in PCa cells injected subcutaneously in MeS mice increased the circulating tumor cells foci compared to control. A microRNA (miRNA) expression microarray from PCa xenografts developed in MeS mice, showed 21 miRNAs modulated by CTBP1 involved in angiogenesis, extracellular matrix organization, focal adhesion and adherents junctions, among others. We found that miR-196b-5p directly targets CLCA2 by cloning CLCA2 3'UTR and performing reporter assays. Altogether, we identified a new molecular mechanism to explain PCa and MeS link based on CLCA2 repression by CTBP1 and miR-196b-5p molecules that might act as key factors in the progression onset of this disease. © 2018 UICC.

Influence of Bicarbonate/Low-GDP Peritoneal Dialysis Fluid (Bicavera) on In Vitro and Ex Vivo Epithelial-to-Mesenchymal Transition of Mesothelial Cells

PubMed Central

Fernández–Perpén, Antonio; Pérez–Lozano, María Luisa; Bajo, María–Auxiliadora; Albar–Vizcaino, Patricia; Correa, Pilar Sandoval; del Peso, Gloria; Castro, María–José; Aguilera, Abelardo; Ossorio, Marta; Peter, Mirjam E.; Passlick–Deetjen, Jutta; Aroeira, Luiz S.; Selgas, Rafael; López–Cabrera, Manuel; Sánchez–Tomero, J. Antonio

2012-01-01

♦ Background: Peritoneal membrane damage induced by peritoneal dialysis (PD) is largely associated with epithelial-to-mesenchymal transition (EMT) of mesothelial cells (MCs), which is believed to be a result mainly of the glucose degradation products (GDPs) present in PD solutions. ♦ Objectives: This study investigated the impact of bicarbonate-buffered, low-GDP PD solution (BicaVera: Fresenius Medical Care, Bad Homburg, Germany) on EMT of MCs in vitro and ex vivo. ♦ Methods: In vitro studies: Omentum-derived MCs were incubated with lactate-buffered standard PD fluid or BicaVera fluid diluted 1:1 with culture medium. Ex vivo studies: From 31 patients randomly distributed to either standard or BicaVera solution and followed for 24 months, effluents were collected every 6 months for determination of EMT markers in effluent MCs. ♦ Results: Culturing of MCs with standard fluid in vitro resulted in morphology change to a non-epithelioid shape, with downregulation of E-cadherin (indicative of EMT) and strong induction of vascular endothelial growth factor (VEGF) expression. By contrast, in vitro exposure of MCs to bicarbonate/low-GDP solution had less impact on both EMT parameters. Ex vivo studies partially confirmed the foregoing results. The BicaVera group, with a higher prevalence of the non-epithelioid MC phenotype at baseline (for unknown reasons), showed a clear and significant trend to gain and maintain an epithelioid phenotype at medium- and longer-term and to show fewer fibrogenic characteristics. By contrast, the standard solution group demonstrated a progressive and significantly higher presence of the non-epithelioid phenotype. Compared with effluent MCs having an epithelioid phenotype, MCs with non-epithelioid morphology showed significantly lower levels of E-cadherin and greater levels of fibronectin and VEGF. In comparing the BicaVera and standard solution groups, MCs from the standard solution group showed significantly higher secretion of interleukin 8 and lower secretion of collagen I, but no differences in the levels of other EMT-associated molecules, including fibronectin, VEGF, E-cadherin, and transforming growth factor β1. Peritonitis incidence was similar in both groups. Functionally, the use of BicaVera fluid was associated with higher transport of small molecules and lower ultrafiltration capacity. ♦ Conclusions: Effluent MCs grown ex vivo from patients treated with bicarbonate/low-GDP BicaVera fluid showed a trend to acquire an epithelial phenotype, with lower production of proinflammatory cytokines and chemokines (such as interleukin 8) than was seen with MCs from patients treated with a lactate-buffered standard PD solution. PMID:22215656

EML4-ALK induces epithelial-mesenchymal transition consistent with cancer stem cell properties in H1299 non-small cell lung cancer cells.

PubMed

Guo, Fuchun; Liu, Xiaoke; Qing, Qin; Sang, Yaxiong; Feng, Chengjun; Li, Xiaoyu; Jiang, Li; Su, Pei; Wang, Yongsheng

2015-04-10

The echinoderm microtubule-associated protein-like 4(EML4)--anaplastic lymphoma kinase (ALK) fusion gene has been identified as a driver mutation in non-small-cell lung cancer (NSCLC). However, the role of EML4-ALK in malignant transformation is not entirely clear. Here, for the first time, we showed that H1299 NSCLC cells stably expressing EML4-ALK acquire EMT phenotype, associated with enhanced invasive migration and increased expression of EMT-inducing transcription factors. H1299-EML4-ALK cells also displayed cancer stem cell-like properties with a concomitant up-regulation of CD133 and enhanced ability of mammospheres formation. Moreover, we found that inhibition of ERK1/2 reversed EMT induced by EML4-ALK in H1299 cells. Taken together, these results suggested that EML4-ALK induced ERK activation is mechanistically associated with EMT phenotype. Thus, inhibition of ERK signaling pathway could be a potential strategy in treatment of NSCLC patients with EML4-ALK translocation. Copyright © 2015 Elsevier Inc. All rights reserved.

Translational regulation of inhibin βA by TGFβ via the RNA-binding protein hnRNP E1 enhances the invasiveness of epithelial-to-mesenchymal transitioned cells.

PubMed

Howley, B V; Hussey, G S; Link, L A; Howe, P H

2016-03-31

The epithelial-to-mesenchymal transition (EMT) is a cellular process that functions during embryonic development and tissue regeneration, thought to be aberrantly activated in epithelial-derived cancer and has an important role in the process of metastasis. The transforming growth factor (TGF)-β signaling pathway is a key inducer of EMT and we have elucidated a posttranscriptional mechanism by which TGFβ modulates expression of select transcripts via the RNA-binding protein hnRNP E1 during EMT. One such transcript inhibin βA is a member of the TGFβ superfamily. Here, we show by polysome profiling that inhibin βA is translationally regulated by TGFβ via hnRNP E1. TGFβ treatment or knockdown of hnRNP E1 relieves silencing of the inhibin βA transcript, resulting in increased protein expression and secreted levels of the inhibin βA homodimer, activin A. Our data indicate that the translational upregulation of inhibin βA enhances the migration and invasion of cells that have undergone an EMT and promotes cancer progression in vivo.

Stabilization of gene expression and cell morphology after explant recycling during fin explant culture in goldfish

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chenais, Nathalie; Lareyre, Jean-Jacques; Le Bail, Pierre-Yves

The development of fin primary cell cultures for in vitro cellular and physiological studies is hampered by slow cell outgrowth, low proliferation rate, poor viability, and sparse cell characterization. Here, we investigated whether the recycling of fresh explants after a first conventional culture could improve physiological stability and sustainability of the culture. The recycled explants were able to give a supplementary cell culture showing faster outgrowth, cleaner cell layers and higher net cell production. The cells exhibited a highly stabilized profile for marker gene expression including a low cytokeratin 49 (epithelial marker) and a high collagen 1a1 (mesenchymal marker) expression.more » Added to the cell spindle-shaped morphology, motility behavior, and actin organization, this suggests that the cells bore stable mesenchymal characteristics. This contrast with the time-evolving expression pattern observed in the control fresh explants during the first 2 weeks of culture: a sharp decrease in cytokeratin 49 expression was concomitant with a gradual increase in col1a1. We surmise that such loss of epithelial features for the benefit of mesenchymal ones was triggered by an epithelial to mesenchymal transition (EMT) process or by way of a progressive population replacement process. Overall, our findings provide a comprehensive characterization of this new primary culture model bearing mesenchymal features and whose stability over culture time makes those cells good candidates for cell reprogramming prior to nuclear transfer, in a context of fish genome preservation. - Highlights: • Recycled fin explants outgrow cells bearing stable mesenchymal traits. • Cell production and quality is enhanced in the recycled explant culture system. • Fresh fin primary culture is highly variable and loose epithelial traits over time.« less

Human RON receptor tyrosine kinase induces complete epithelial-to-mesenchymal transition but causes cellular senescence

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cote, Marceline; Miller, A. Dusty; Liu, Shan-Lu

2007-08-17

The RON receptor tyrosine kinase is a member of the MET proto-oncogene family and is important for cell proliferation, differentiation, and cancer development. Here, we created a series of Madin-Darby canine kidney (MDCK) epithelial cell clones that express different levels of RON, and have investigated their biological properties. While low levels of RON correlated with little morphological change in MDCK cells, high levels of RON expression constitutively led to morphological scattering or complete and stabilized epithelial-to-mesenchymal transition (EMT). Unexpectedly, MDCK clones expressing higher levels of RON exhibited retarded proliferation and senescence, despite increased motility and invasiveness. RON was constitutively tyrosine-phosphorylatedmore » in MDCK cells expressing high levels of RON and undergoing EMT, and the MAPK signaling pathway was activated. This study reveals for the first time that RON alone is sufficient to induce complete and stabilized EMT in MDCK cells, and overexpression of RON does not cause cell transformation but rather induces cell cycle arrest and senescence, leading to impaired cell proliferation.« less

Growth hormone-releasing hormone antagonists reduce prostatic enlargement and inflammation in carrageenan-induced chronic prostatitis.

PubMed

Popovics, Petra; Cai, Renzhi; Sha, Wei; Rick, Ferenc G; Schally, Andrew V

2018-05-21

Inflammation plays a key role in the etiology of benign prostatic hyperplasia (BPH) through multiple pathways involving the stimulation of proliferation by cytokines and growth factors as well as the induction of the focal occurrence of epithelial-to-mesenchymal transition (EMT). We have previously reported that GHRH acts as a prostatic growth factor in experimental BPH and in autoimmune prostatitis models and its blockade with GHRH antagonists offer therapeutic approaches for these conditions. Our current study was aimed at the investigation of the beneficial effects of GHRH antagonists in λ-carrageenan-induced chronic prostatitis and at probing the downstream molecular pathways that are implicated in GHRH signaling. To demonstrate the complications triggered by recurrent/chronic prostatic inflammation in Sprague-Dawley rats, 50 μL 3% carrageenan was injected into both ventral prostate lobes two times, 3 weeks apart. GHRH antagonist, MIA-690, was administered 5 days after the second intraprostatic injection at 20 μg daily dose for 4 weeks. GHRH-induced signaling events were identified in BPH-1 and in primary prostate epithelial (PrEp) cells at 5, 15, 30, and 60 min with Western blot. Inflammation induced prostatic enlargement and increased the area of the stromal compartment whereas treatment with the GHRH antagonist significantly reduced these effects. This beneficial activity was consistent with a decrease in prostatic GHRH, inflammatory marker COX-2, growth factor IGF-1 and inflammatory and EMT marker TGF-β1 protein levels and the expression of multiple genes related to EMT. In vitro, GHRH stimulated multiple pathways involved in inflammation and growth in both BPH-1 and PrEp cells including NFκB p65, AKT, ERK1/2, EGFR, STAT3 and increased the levels of TGF-β1 and Snail/Slug. Most interestingly, GHRH also stimulated the transactivation of the IGF receptor. The study demonstrates that GHRH antagonists could be beneficial for the treatment of prostatic inflammation and BPH in part by inhibiting the growth-promoting and inflammatory effects of locally produced GHRH. © 2018 Wiley Periodicals, Inc.

Mechanism of c-Met and EGFR tyrosine kinase inhibitor resistance through epithelial mesenchymal transition in non-small cell lung cancer

DOE Office of Scientific and Technical Information (OSTI.GOV)

Rastogi, Ichwaku; Rajanna, Supriya; Webb, Andrew

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) inmore » 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.« less

1,25(OH)2D3 attenuates TGF-β1/β2-induced increased migration and invasion via inhibiting epithelial–mesenchymal transition in colon cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Shanwen; Zhu, Jing; Zuo, Shuai

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-actinmore » 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. - Highlights: • TGF-β1/β2-induced model of EMT was used in this study to test the effect of 1,25(OH)2D3 on EMT in colon cancer cells. • 1,25(OH)2D3 inhibited TGF-β1/β2-induced increased migration and invasion. • 1,25(OH)2D3 inhibited TGF-β1/β2-induced increased level of EMT-related transcription factors. • 1,25(OH)2D3 inhibited TGF-β1/β2-induced increased expression of F-actin in SW-480 cells.« less

Snail1 transcription factor controls telomere transcription and integrity

PubMed Central

Mazzolini, Rocco; Gonzàlez, Núria; Garcia-Garijo, Andrea; Millanes-Romero, Alba; Peiró, Sandra; Smith, Susan

2018-01-01

Abstract Besides controlling epithelial-to-mesenchymal transition (EMT) and cell invasion, the Snail1 transcriptional factor also provides cells with cancer stem cell features. Since telomere maintenance is essential for stemness, we have examined the control of telomere integrity by Snail1. Fluorescence in situ hybridization (FISH) analysis indicates that Snail1-depleted mouse mesenchymal stem cells (MSC) have both a dramatic increase of telomere alterations and shorter telomeres. Remarkably, Snail1-deficient MSC present higher levels of both telomerase activity and the long non-coding RNA called telomeric repeat-containing RNA (TERRA), an RNA that controls telomere integrity. Accordingly, Snail1 expression downregulates expression of the telomerase gene (TERT) as well as of TERRA 2q, 11q and 18q. TERRA and TERT are transiently downregulated during TGFβ-induced EMT in NMuMG cells, correlating with Snail1 expression. Global transcriptome analysis indicates that ectopic expression of TERRA affects the transcription of some genes induced during EMT, such as fibronectin, whereas that of TERT does not modify those genes. We propose that Snail1 repression of TERRA is required not only for telomere maintenance but also for the expression of a subset of mesenchymal genes. PMID:29059385

Evidence for epithelial-mesenchymal transition in cancer stem-like cells derived from carcinoma cell lines of the cervix uteri.

PubMed

Lin, Jiaying; Liu, Xishi; Ding, Ding

2015-01-01

The cancer stem cell (CSC) paradigm is one possible way to understand the genesis of cancer, and cervical cancer in particular. We quantified and enriched ALDH1(+) cells within cervical cancer cell lines and subsequently characterized their phenotypical and functional properties like invasion capacity and epithelial-mesenchymal transition (EMT). ALDH1 expression in spheroid-derived cells (SDC) and the parental monolayer-derived cell (MDC) line was compared by flow-cytometry. Invasion capability was evaluated by Matrigel assay and expression of EMT-related genes Twist 1, Twist 2, Snail 1, Snail 2, Vimentin and E-cadherin by real-time PCR. ALDH1 expression was significantly higher in SDC. ALDH1(+) cells showed increased colony-formation. SDC expressed lower levels of E-cadherin and elevated levels of Twist 1, Twist 2, Snail 1, Snail 2 and Vimentin compared to MDC. Cervical cancer cell lines harbor potential CSC, characterized by ALDH1 expression as well as properties like invasiveness, colony-forming ability, and EMT. CSC can be enriched by anchorage-independent culture techniques, which may be important for the investigation of their contribution to therapy resistance, tumor recurrence and metastasis.

Forkhead box K2 inhibits the proliferation, migration, and invasion of human glioma cells and predicts a favorable prognosis.

PubMed

Wang, Bo; Zhang, XueBin; Wang, Wei; Zhu, ZhiZhong; Tang, Fan; Wang, Dong; Liu, Xi; Zhuang, Hao; Yan, XiaoLing

2018-01-01

Forkhead box K2 (FOXK2) is a member of the forkhead box family of transcription factors. Recently, researchers discovered that overexpression of FOXK2 inhibits the proliferation and metastasis of breast cancer, non-small cell lung cancer, and colorectal cancer, and is related to the clinical prognosis. However, in hepatocellular carcinoma, FOXK2 results in the opposite phenotypes. Currently, the contribution of FOXK2 to glioma pathogenesis is not clear. We evaluated the expression of FOXK2 in 151 glioma patients using immunohistochemistry assays. The associations among the expression of FOXK2, clinicopathological parameters, and the prognosis of glioma patients were statistically analyzed. We downregulated and upregulated the level of FOXK2 in glioma cells by transfections with small interfering RNA and plasmids. Then, we investigated the effects on tumor cell behavior in vitro by Cell Counting Kit-8 assays, colony-formation assay, transwell assay, and the epithelial-to-mesenchymal transition (EMT) biomarker levels. The clinical data showed that expression of FOXK2 gradually decreased with increasing World Health Organization (WHO) grades and a low level of FOXK2 indicates a poor prognosis. FOXK2 expression is negatively correlated with Ki67 expression and the WHO degree but is not correlated with other clinicopathological parameters, including sex, age, Karnofsky Performance Status, tumor diameter, O -6-methylguanine-DNA methyltransferase, and glutathione S -transferase pi. FOXK2 knockdown enhances glioma cell proliferation, migration, invasion, and EMT process, and, in contrast, FOXK2 overexpression inhibits glioma cell proliferation, migration, invasion, and the EMT process. Expression of FOXK2 gradually decreases with increasing WHO grades. FOXK2 inhibits tumor proliferation, migration, and invasion. FOXK2 is a critical mediator of the EMT process.

MicroRNA-630 Suppresses Epithelial-to-Mesenchymal Transition by Regulating FoxM1 in Gastric Cancer Cells.

PubMed

Feng, Jing; Wang, Xiaojuan; Zhu, Weihua; Chen, Si; Feng, Changwei

2017-06-01

In the present study, we investigated the functional role of microRNA (miR)-630 in epithelial-to-mesenchymal transition (EMT) of gastric cancer (GC) cells, as well as the regulatory mechanism. Cells of human GC cell line SGC 7901 were transfected with miR-630 mimic or miR-630 inhibitor. The transfection efficiency was confirmed by qRT-PCR. Cell migration and invasion were determined by Transwell assay. Protein expression of E-cadherin, vimentin, and Forkhead box protein M1 (FoxM1) was tested by Western blot. Moreover, the expression of FoxM1 was elevated or suppressed, and then the effects of miR-630 abnormal expression on EMT and properties of migration and invasion were examined again, as well as protein expression of Ras/phosphoinositide 3-kinase (PI3K)/AKT related factors. The results showed that (i) the EMT and properties of migration and invasion were statistically decreased by overexpression of miR-630 compared to the control group but markedly increased by suppression of miR-630. However, (ii) abnormal expression of FoxM1 reversed these effects in GC cells. Moreover, (iii) expression of GTP-Rac1, p-PI3K, and p-AKT was decreased by miR-630 overexpression but increased by FoxM1 overexpression. (iv) The decreased levels of GTP-Rac1, p-PI3K, and p-AKT induced by miR-630 overexpression were dramatically elevated by simultaneous overexpression of FoxM1. In conclusion, our results suggest that miR-630 might be a tumor suppressor in GC cells. MiR-630 suppresses EMT by regulating FoxM1 in GC cells, supposedly via inactivation of the Ras/PI3K/AKT pathway.

Implication of Highly Cytotoxic Natural Killer Cells for Esophageal Squamous Cell Carcinoma Treatment.

PubMed

Lim, Kee Siang; Mimura, Kosaku; Kua, Ley-Fang; Shiraishi, Kensuke; Kono, Koji

2018-04-20

Esophageal squamous cell carcinoma (ESCC) is an aggressive upper gastrointestinal cancer and effective treatments are limited. Previous studies reported that natural killer (NK) cells expanded by coculturing with K562-mb15-41BBL feeder cells, a genetically modified K562 leukemia cell line that expresses membrane-bound interleukin (IL)-15 and 41BBL ligand, were highly proliferative and highly cytotoxic. Here, we investigated the potential of expanded NK cells for ESCC treatment. We analyzed both genetic and surface expression levels of NKG2D ligands (NKG2DLs) in ESCC using publicly available microarray data sets and ESCC cell lines. The cytotoxicity of resting and of IL-2-activated NK cells against ESCC cell lines was compared with that of expanded NK cells. We then also investigated the effect of epithelial mesenchymal transition (EMT) inducers, GSK3β inhibitor and epidermal growth factor, on NKG2DLs expressions. As a result, MICA and MICB were significantly overexpressed in ESCC compared with adjacent normal tissues and surface NKG2DLs were expressed in ESCC cell lines. Expanded NK cells were much potent than IL-2-activated and resting NK cells against ESCC cell lines. Blocking of NKG2D with anti-NKG2D monoclonal antibody dampened expanded NK cell cytotoxicity, suggesting that the NKG2DLs-NKG2D interaction is crucial for NK cells to eliminate ESCC cells. EMT inducers concurrently induced EMT and NKG2DLs expression in ESCC cell lines rendering transitioned cells more sensitive to expanded NK cells. In conclusion, expanded NK cells were highly cytotoxic against NKG2DLs-expressing ESCC cells, particularly the EMT phenotype. These results provide a strong rationale for clinical use of these NK cells in ESCC patients.

Cadherin Expression, Vectorial Active Transport, and Metallothionein Isoform 3 Mediated EMT/MET Responses in Cultured Primary and Immortalized Human Proximal Tubule Cells

PubMed Central

Slusser, Andrea; Bathula, Chandra S.; Sens, Donald A.; Somji, Seema; Sens, Mary Ann; Zhou, Xu Dong; Garrett, Scott H.

2015-01-01

Background Cultures of human proximal tubule cells have been widely utilized to study the role of EMT in renal disease. The goal of this study was to define the role of growth media composition on classic EMT responses, define the expression of E- and N-cadherin, and define the functional epitope of MT-3 that mediates MET in HK-2 cells. Methods Immunohistochemistry, microdissection, real-time PCR, western blotting, and ELISA were used to define the expression of E- and N-cadherin mRNA and protein in HK-2 and HPT cell cultures. Site-directed mutagenesis, stable transfection, measurement of transepithelial resistance and dome formation were used to define the unique amino acid sequence of MT-3 associated with MET in HK-2 cells. Results It was shown that both E- and N-cadherin mRNA and protein are expressed in the human renal proximal tubule. It was shown, based on the pattern of cadherin expression, connexin expression, vectorial active transport, and transepithelial resistance, that the HK-2 cell line has already undergone many of the early features associated with EMT. It was shown that the unique, six amino acid, C-terminal sequence of MT-3 is required for MT-3 to induce MET in HK-2 cells. Conclusions The results show that the HK-2 cell line can be an effective model to study later stages in the conversion of the renal epithelial cell to a mesenchymal cell. The HK-2 cell line, transfected with MT-3, may be an effective model to study the process of MET. The study implicates the unique C-terminal sequence of MT-3 in the conversion of HK-2 cells to display an enhanced epithelial phenotype. PMID:25803827

Intratumoral bidirectional transitions between epithelial and mesenchymal cells in triple-negative breast cancer.

PubMed

Yamamoto, Mizuki; Sakane, Kota; Tominaga, Kana; Gotoh, Noriko; Niwa, Takayoshi; Kikuchi, Yasuko; Tada, Keiichiro; Goshima, Naoki; Semba, Kentaro; Inoue, Jun-Ichiro

2017-06-01

Epithelial-mesenchymal transition (EMT) and its reverse process, mesenchymal-epithelial transition MET, are crucial in several stages of cancer metastasis. Epithelial-mesenchymal transition allows cancer cells to move to proximal blood vessels for intravasation. However, because EMT and MET processes are dynamic, mesenchymal cancer cells are likely to undergo MET transiently and subsequently re-undergo EMT to restart the metastatic process. Therefore, spatiotemporally coordinated mutual regulation between EMT and MET could occur during metastasis. To elucidate such regulation, we chose HCC38, a human triple-negative breast cancer cell line, because HCC38 is composed of epithelial and mesenchymal populations at a fixed ratio even though mesenchymal cells proliferate significantly more slowly than epithelial cells. We purified epithelial and mesenchymal cells from Venus-labeled and unlabeled HCC38 cells and mixed them at various ratios to follow EMT and MET. Using this system, we found that the efficiency of EMT is approximately an order of magnitude higher than that of MET and that the two populations significantly enhance the transition of cells from the other population to their own. In addition, knockdown of Zinc finger E-box-binding homeobox 1 (ZEB1) or Zinc finger protein SNAI2 (SLUG) significantly suppressed EMT but promoted partial MET, indicating that ZEB1 and SLUG are crucial to EMT and MET. We also show that primary breast cancer cells underwent EMT that correlated with changes in expression profiles of genes determining EMT status and breast cancer subtype. These changes were very similar to those observed in EMT in HCC38 cells. Consequently, we propose HCC38 as a suitable model to analyze EMT-MET dynamics that could affect the development of triple-negative breast cancer. © 2017 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

Data-Driven Discovery of Extravasation Pathway in Circulating Tumor Cells

PubMed Central

Yadavalli, S.; Jayaram, S.; Manda, S. S.; Madugundu, A. K.; Nayakanti, D. S.; Tan, T. Z.; Bhat, R.; Rangarajan, A.; Chatterjee, A.; Gowda, H.; Thiery, J. P.; Kumar, P.

2017-01-01

Circulating tumor cells (CTCs) play a crucial role in cancer dissemination and provide a promising source of blood-based markers. Understanding the spectrum of transcriptional profiles of CTCs and their corresponding regulatory mechanisms will allow for a more robust analysis of CTC phenotypes. The current challenge in CTC research is the acquisition of useful clinical information from the multitude of high-throughput studies. To gain a deeper understanding of CTC heterogeneity and identify genes, pathways and processes that are consistently affected across tumors, we mined the literature for gene expression profiles in CTCs. Through in silico analysis and the integration of CTC-specific genes, we found highly significant biological mechanisms and regulatory processes acting in CTCs across various cancers, with a particular enrichment of the leukocyte extravasation pathway. This pathway appears to play a pivotal role in the migration of CTCs to distant metastatic sites. We find that CTCs from multiple cancers express both epithelial and mesenchymal markers in varying amounts, which is suggestive of dynamic and hybrid states along the epithelial-mesenchymal transition (EMT) spectrum. Targeting the specific molecular nodes to monitor disease and therapeutic control of CTCs in real time will likely improve the clinical management of cancer progression and metastases. PMID:28262832

Reactivation of epigenetically silenced miR-124 reverses the epithelial-to-mesenchymal transition and inhibits invasion in endometrial cancer cells via the direct repression of IQGAP1 expression.

PubMed

Dong, Peixin; Ihira, Kei; Xiong, Ying; Watari, Hidemichi; Hanley, Sharon J B; Yamada, Takahiro; Hosaka, Masayoshi; Kudo, Masataka; Yue, Junming; Sakuragi, Noriaki

2016-04-12

Overexpression of IQGAP1 and microRNA (miRNA) dysregulation are frequent in human tumors, but little is known about the role of IQGAP1 and its relationship to miRNA in endometrial carcinogenesis. We demonstrate that IQGAP1 activates the epithelial-mesenchymal transition (EMT) program and that miR-124 directly represses IQGAP1 expression in endometrial cancer (EC) cells. The overexpression of IQGAP1 stimulates EMT features and enhances migration, invasion and proliferation of EC cells, whereas knocking down IQGAP1 expression reverses EMT and inhibits these malignant properties. Using miRNA microarray profiling, we identified 29 miRNAs (let-7b, let-7f, miR-10b, miR-15b, miR-23a, miR-24, miR-25, miR-27a, miR-29b, miR-30a-5p, miR-34a, miR-124, miR-127, miR-130b, miR-148a, miR-155, miR-191*, miR-194, miR-224, miR-362, miR-409-3p, miR-422b, miR-424, miR-453, miR-497, miR-518d, miR-518f*, miR-526a and miR-656) that are significantly down-regulated in an in vitro-selected highly invasive derivative cell line (HEC-50-HI) relative to the parental HEC-50 cells. We further identified miR-124 as a direct regulator of IQGAP1 in EC cells. Enforced expression of miR-124 suppresses EC cell invasion and proliferation. The expression of IQGAP1 mRNA was significantly elevated in EC tissues, while the expression of miR-124 was decreased. The downregulation of miR-124 correlates with a poor survival outcome for patients with EC. Treating EC cells with the demethylating agent 5-aza-2'-deoxycytidine increased miR-124 expression and down-regulated IQGAP1 levels. Our data suggest that IQGAP1 promotes EMT, migration and invasion of EC cells. MiR-124, a novel tumor suppressor miRNA that is epigenetically silenced in EC, can reverse EMT and the invasive properties, by attenuating the expression of the IQGAP1 oncogene.

Reactivation of epigenetically silenced miR-124 reverses the epithelial-to-mesenchymal transition and inhibits invasion in endometrial cancer cells via the direct repression of IQGAP1 expression

PubMed Central

Watari, Hidemichi; Hanley, Sharon J.B.; Yamada, Takahiro; Hosaka, Masayoshi; Kudo, Masataka; Yue, Junming; Sakuragi, Noriaki

2016-01-01

Overexpression of IQGAP1 and microRNA (miRNA) dysregulation are frequent in human tumors, but little is known about the role of IQGAP1 and its relationship to miRNA in endometrial carcinogenesis. We demonstrate that IQGAP1 activates the epithelial–mesenchymal transition (EMT) program and that miR-124 directly represses IQGAP1 expression in endometrial cancer (EC) cells. The overexpression of IQGAP1 stimulates EMT features and enhances migration, invasion and proliferation of EC cells, whereas knocking down IQGAP1 expression reverses EMT and inhibits these malignant properties. Using miRNA microarray profiling, we identified 29 miRNAs (let-7b, let-7f, miR-10b, miR-15b, miR-23a, miR-24, miR-25, miR-27a, miR-29b, miR-30a-5p, miR-34a, miR-124, miR-127, miR-130b, miR-148a, miR-155, miR-191*, miR-194, miR-224, miR-362, miR-409-3p, miR-422b, miR-424, miR-453, miR-497, miR-518d, miR-518f*, miR-526a and miR-656) that are significantly down-regulated in an in vitro-selected highly invasive derivative cell line (HEC-50-HI) relative to the parental HEC-50 cells. We further identified miR-124 as a direct regulator of IQGAP1 in EC cells. Enforced expression of miR-124 suppresses EC cell invasion and proliferation. The expression of IQGAP1 mRNA was significantly elevated in EC tissues, while the expression of miR-124 was decreased. The downregulation of miR-124 correlates with a poor survival outcome for patients with EC. Treating EC cells with the demethylating agent 5-aza-2′-deoxycytidine increased miR-124 expression and down-regulated IQGAP1 levels. Our data suggest that IQGAP1 promotes EMT, migration and invasion of EC cells. MiR-124, a novel tumor suppressor miRNA that is epigenetically silenced in EC, can reverse EMT and the invasive properties, by attenuating the expression of the IQGAP1 oncogene. PMID:26934121

Salt-induced epithelial-to-mesenchymal transition in Dahl salt-sensitive rats is dependent on elevated blood pressure.

PubMed

Wang, Y; Mu, J J; Liu, F Q; Ren, K Y; Xiao, H Y; Yang, Z; Yuan, Z Y

2014-02-01

Dietary salt intake has been linked to hypertension and cardiovascular disease. Accumulating evidence has indicated that salt-sensitive individuals on high salt intake are more likely to develop renal fibrosis. Epithelial-to-mesenchymal transition (EMT) participates in the development and progression of renal fibrosis in humans and animals. The objective of this study was to investigate the impact of a high-salt diet on EMT in Dahl salt-sensitive (SS) rats. Twenty-four male SS and consomic SS-13(BN) rats were randomized to a normal diet or a high-salt diet. After 4 weeks, systolic blood pressure (SBP) and albuminuria were analyzed, and renal fibrosis was histopathologically evaluated. Tubular EMT was evaluated using immunohistochemistry and real-time PCR with E-cadherin and alpha smooth muscle actin (α-SMA). After 4 weeks, SBP and albuminuria were significantly increased in the SS high-salt group compared with the normal diet group. Dietary salt intake induced renal fibrosis and tubular EMT as identified by reduced expression of E-cadherin and enhanced expression of α-SMA in SS rats. Both blood pressure and renal interstitial fibrosis were negatively correlated with E-cadherin but positively correlated with α-SMA. Salt intake induced tubular EMT and renal injury in SS rats, and this relationship might depend on the increase in blood pressure.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liao, Xiao-hui; Zhang, Ling, E-mail: lindazhang8508@hotmail.com; Chen, Guo-tao

Tubular epithelial-to-mesenchymal transition (EMT) plays a crucial role in the progression of renal tubular interstitial fibrosis (TIF), which subsequently leads to chronic kidney disease (CKD) and eventually, end-stage renal disease (ESRD). We propose that augmenter of liver regeneration (ALR), a member of the newly discovered ALR/Erv1 protein family shown to ameliorate hepatic fibrosis, plays a similar protective role in renal tubular cells and has potential as a new treatment option for CKD. Here, we showed that recombinant human ALR (rhALR) inhibits EMT in renal tubular cells by antagonizing activation of the transforming growth factor-β1 (TGF-β1) signaling pathway. Further investigation revealedmore » that rhALR suppresses the expression of TGF-β receptor type II (TβR II) and significantly alleviates TGF-β1-induced phosphorylation of Smad2 and nuclear factor-κB (NF-κB). No apparent adverse effects were observed upon the addition of rhALR alone to cells. These findings collectively suggest that ALR plays a role in inhibiting progression of renal tubular EMT, supporting its potential utility as an effective antifibrotic strategy to reverse TIF in CKD. - Highlights: • ALR is involved in the pathological progression of renal EMT in NRK-52E cells. • ALR suppresses the expression of TβRII and the phosphorylation of Smad2 and NF-κB. • ALR plays a role in inhibiting progression of renal tubular EMT.« less

Doxycycline reverses epithelial-to-mesenchymal transition and suppresses the proliferation and metastasis of lung cancer cells

PubMed Central

Liu, Yan-rong; Liu, Hui-juan; Zhao, Dong; Chen, Shuang; Xiao, Ting; Meng, Jing; Jing, Xue-shuang; Wang, Jing; Sun, Bo; Dai, Ting-ting; Yang, Cheng; Sun, Tao; Zhou, Hong-gang

2015-01-01

The gelatinase inhibitor doxycycline is the prototypical antitumor antibiotic. We investigated the effects of doxycycline on the migration, invasion, and metastasis of human lung cancer cell lines and in a mouse model. We also measured the effect of doxycycline on the transcription of epithelial-mesenchymal transition (EMT) markers, and used immunohistochemistry to determine whether EMT reversal was associated with doxycycline inhibition. Doxycycline dose-dependently inhibited proliferation, migration, and invasion of NCI-H446 human small cell lung cancer cells. It also suppressed tumor growth from NCI-H446 and A549 lung cancer cell xenografts without altering body weight, inhibited Lewis lung carcinoma cell migration, and prolonged survival. The activities of the transcription factors Twist1/2, SNAI1/2, AP1, NF-κB, and Stat3 were suppressed by doxycycline, which reversed EMT and inhibited signal transduction, thereby suppressing tumor growth and metastasis. Our data demonstrate functional targeting of transcription factors by doxycycline to reverse EMT and suppress tumor proliferation and metastasis. Thus, doxycycline selectively targets malignant tumors and reduces its metastatic potential with less cytotoxicity in lung cancer patients. PMID:26512779

Mechanisms of the epithelial-to-mesenchymal transition in sea urchin embryos

PubMed Central

Katow, Hideki

2015-01-01

Sea urchin mesenchyme is composed of the large micromere-derived spiculogenetic primary mesenchyme cells (PMC), veg2-tier macromere-derived non-spiculogenetic mesenchyme cells, the small micromere-derived germ cells, and the macro- and mesomere-derived neuronal mesenchyme cells. They are formed through the epithelial-to-mesenchymal transition (EMT) and possess multipotency, except PMCs that solely differentiate larval spicules. The process of EMT is associated with modification of epithelial cell surface property that includes loss of affinity to the apical and basal extracellular matrices, inter-epithelial cell adherens junctions and epithelial cell surface-specific proteins. These cell surface structures and molecules are endocytosed during EMT and utilized as initiators of cytoplasmic signaling pathways that often initiate protein phosphorylation to activate the gene regulatory networks. Acquisition of cell motility after EMT in these mesenchyme cells is associated with the expression of proteins such as Lefty, Snail and Seawi. Structural simplicity and genomic database of this model will further promote detailed EMT research. PMID:26716069

Activation of IRE1α-XBP1 pathway induces cell proliferation and invasion in colorectal carcinoma

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jin, Chun; Jin, Zhao; Chen, Nian-zhao

2016-01-29

Cell proliferation and tumor metastasis are considered as the main reasons for death in colorectal carcinoma (CRC). IRE1α-XBP1 pathway is the most conserved UPR pathways, which are activated during ER stress caused by the accumulation of unfolded or misfolded protein in the lumen of ER. Here, we demonstrated the critical role of IRE1α-XBP1 pathway and underlying molecular mechanism in cell proliferation and tumor metastasis in CRC. By the use of tissue microarray analysis of samples from 119 patients with CRC, IRE1α was determined to be an independent predictor of overall survival as higher expression of IRE1α in CRC patients showedmore » lower survival rates (p = 0.0041). RNA interference and ectopic expression of IRE1α were applied to determine the molecular effects of IRE1α in CRC cells. The silencing of IRE1α inhibited the proliferation and blocked the invasion of CRC cells in vitro, while ectopic expression of IRE1α in turn promoted cell proliferation and invasion. IRE1α-XBP1 pathway regulated the mitosis of CRC cells through the directly binding of XBP1s to Cyclin D1 promoter to activate Cyclin D1 expression. Our results reveal that IRE1α-XBP1 pathway plays an important role in tumor progression and epithelial-to-mesenchymal transition (EMT), and IRE1α could be employed as a novel prognostic marker and a promising therapeutic target for CRC. - Highlights: • IRE1 was determined to be an independent predictor of overall survival in CRC patient. • IRE1-XBP1 pathway promoted CRC cell proliferation through regulating Cyclin D1 expression. • IRE1-XBP1 pathway played important role in EMT of CRC cells.« less

NR2F2 inhibits Smad7 expression and promotes TGF-β-dependent epithelial-mesenchymal transition of CRC via transactivation of miR-21.

PubMed

Wang, Hao; Nie, Lei; Wu, Lei; Liu, Qiufang; Guo, Xueyan

2017-03-25

Metastasis is one of the most decisive factors influencing CRC patient prognosis and current studies suggest that a molecular mechanism known as EMT broadly regulates cancer metastasis. NR2F2 is a key molecule in the development of CRC, but the roles and underlying mechanisms of NR2F2 in TGF-β induced EMT in CRC remain largely unknown. In the current study, we were interested to examine the role of NR2F2 in the TGF-β-induced EMT in CRC. Here, we found NR2F2 was upregulated in CRC cells and promotes TGF-β-induced EMT in CRC. Using comparative miRNA profiling TGF-β pre-treated CRC cells in which NR2F2 had been knocked down with that of control cells, we identified miR-21 as a commonly downregulated miRNA in HT29 cells treated with TGF-β and NR2F2 siRNA, and its downregulation inhibiting migration and invasion of CRC cells. Moreover, we found NR2F2 could transcriptional activated miR-21 expression by binding to miR-21 promoter in HT29 by ChIP and luciferase assay. In the last, our data demonstrated that Smad7 was the direct target of miR-21 in CRC cells. Thus, NR2F2 could promote TGF-β-induced EMT and inhibit Smad7 expression via transactivation of miR-21, and NR2F2 may be a new common therapeutic target for CRC. Copyright © 2017 Elsevier Inc. All rights reserved.

The inhibition of lung cancer cell migration by AhR-regulated autophagy

PubMed Central

Tsai, Chi-Hao; Li, Ching-Hao; Cheng, Yu-Wen; Lee, Chen-Chen; Liao, Po-Lin; Lin, Cheng-Hui; Huang, Shih-Hsuan; Kang, Jaw-Jou

2017-01-01

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that is highly expressed in multiple organs and tissues. Whereas AhR mediates the metabolism of xenobiotic and endogenous compounds, its novel function in cancer epithelial-mesenchymal transition (EMT) remains controversial. Autophagy also participates in tumour progression through its functions in cell homeostasis and facilitates adaptation to EMT progression. In the present study, we found that AhR-regulated autophagy positively modulates EMT in non-small cell lung cancer cells. The motility of A549, H1299, and CL1-5 cells were correlated with different AhR expression levels. Invasive potential and cell morphology also changed when AhR protein expression was altered. Moreover, AhR levels exerted a contrasting effect on autophagy potential. Autophagy was higher in CL1-5 and H1299 cells with lower AhR levels than in A549 cells. Both AhR overexpression and autophagy inhibition decreased CL1-5 metastasis in vivo. Furthermore, AhR promoted BNIP3 ubiquitination for proteasomal degradation. AhR silencing in A549 cells also reduced BNIP3 ubiquitination. Taken together, these results provide a novel insight into the cross-linking between AhR and autophagy, we addressed the mechanistic BNIP3 modulation by endogenous AhR, which affect cancer cell EMT progression. PMID:28195146

EML4-ALK induces epithelial–mesenchymal transition consistent with cancer stem cell properties in H1299 non-small cell lung cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Guo, Fuchun; Liu, Xiaoke, E-mail: liuxk57@163.com; Qing, Qin, E-mail: qinqingscu@126.com

2015-04-10

The echinoderm microtubule-associated protein-like 4(EML4) – anaplastic lymphoma kinase (ALK) fusion gene has been identified as a driver mutation in non-small-cell lung cancer (NSCLC). However, the role of EML4-ALK in malignant transformation is not entirely clear. Here, for the first time, we showed that H1299 NSCLC cells stably expressing EML4-ALK acquire EMT phenotype, associated with enhanced invasive migration and increased expression of EMT-inducing transcription factors. H1299-EML4-ALK cells also displayed cancer stem cell-like properties with a concomitant up-regulation of CD133 and enhanced ability of mammospheres formation. Moreover, we found that inhibition of ERK1/2 reversed EMT induced by EML4-ALK in H1299 cells.more » Taken together, these results suggested that EML4-ALK induced ERK activation is mechanistically associated with EMT phenotype. Thus, inhibition of ERK signaling pathway could be a potential strategy in treatment of NSCLC patients with EML4-ALK translocation. - Highlights: • EML4-ALK induced epithelial–mesenchymal transition in H1299 cells. • Expression of EML4-ALK promotes invasion and migration in vitro. • EML4-ALK enhanced sphere formation and stem cell-like properties in H1299 cells. • Blockage of ERK1/2 reverse Epithelial–Mesenchymal transition induced by EML4-ALK.« less