Expression Of Epithelial-mesenchymal Transition Biomarkers Research Articles

P4HA2 involved in SLUG-associated EMT predicts poor prognosis of patients with KRAS-positive colorectal cancer.

This study aimed to examine the immunohistochemical expression of epithelial-mesenchymal transition biomarkers: P4HA2 and SLUG in colorectal carcinoma (CRC) specimens, then to assess their relation to clinicopathological features including KRAS mutations and patients' survival, and finally to study the correlation between them in CRC. The result of this study showed that SLUG and P4HA2 were significantly higher in association with adverse prognostic factors: presence of lympho-vascular invasion, perineural invasion, higher tumor budding, tumor stage, presence of lymph node metastasis, and presence of distant metastasis. CRC specimens with KRAS mutation were associated with significant higher SLUG and P4HA2 expression. High expression of both SLUG and P4HA2 was significantly unfavorable prognostic indicator as regards overall survival (OS) and disease-free survival (DFS). In KRAS mutated cases, high P4HA2 expression was the only significant poor prognostic indicator as regarding DFS. In conclusions, our data highlight that both SLUG and P4HA2 expression may serve as potentially important poor prognostic biomarkers in CRC and targeting these molecules may be providing a novel therapeutic strategy. In KRAS mutation group, high P4HA2 expression is the only independent prognostic factor for tumor recurrence, so it can be suggested to be a novel target for therapy.

Low expression of RACK1 is associated with metastasis and worse prognosis in cholangiocarcinoma

BackgroundCholangiocarcinoma is a poorly prognostic malignant tumor, and the metastatic stage of cancer is not an early stage when diagnosed. Lymph node metastasis is common in the early stage. Ribosomal receptor for activated C-kinase 1 (RACK1) has found involved in the oncogenesis of various tumors and in the epithelial-mesenchymal transition (EMT). Nevertheless, its role in cholangiocarcinoma remains unknown. Material and methodsThe possible correlation between RACK1 and tumor prognosis was analyzed in cholangiocarcinoma patients. The GEO and TCGA databases were used to evaluate the level of RACK1 in cholangiocarcinoma. The RBE and HCCC-9810 cell lines were used to examine the effects of RACK1 in the behavior of tumor cells in vitro. ResultsThe Kaplan-Meier analysis indicated that low expression of RACK1 was associated with poor prognosis and RACK1 was negatively related to lymph node metastasis, which were verified in databases TCGA and GEO; downregulation of RACK1 via RNA interference correlated with changes in the expression of EMT biomarkers and promoted the migration of cholangiocarcinoma cell lines. ConclusionThe protein expression of RACK1 is significantly higher in cholangiocarcinoma tissues than in peritumoral tissues, however, the high RACK1 expression indicates better overall survival and less risk for lymph node metastasis. In vitro, RACK1 may suppress the migratory ability of cholangiocarcinoma cells by inhibiting EMT.

EMT and Cancer Cell Stemness Associated With Chemotherapeutic Resistance in Esophageal Cancer.

Drug resistance often occurs after chemotherapy in esophageal cancer patients, leading to cancer metastasis and recurrence. However, the relationship among cancer cell migration, recurrence and drug resistance in esophageal cancer drug-resistant cells has not been clearly explained. In this study, we constructed paclitaxel (PTX)-resistant esophageal cancer cells to explore the causes of drug resistance and poor prognosis after chemotherapy in esophageal cancer. Colony formation assay was used to evaluate the difference of colony formation between parental cells and drug resistance cells. Microsphere formation assay was used to examine the phenotype of stem cells. Wound healing and Transwell assays were used to detect the migration ability of drug-resistant cells. Western blotting and immunofluorescence assays were used to explore the mechanisms. Finally, we used nude mouse xenograft model to explore the tumor characteristics and the expression of relative proteins to verify our findings in vivo. Our study demonstrated that the cancer cell stemness characteristics exist in drug-resistant esophageal cancer cells, that expressed the biomarkers of stem cells and were prone to epithelial-mesenchymal transition (EMT). Our results suggested that the expression of EMT biomarkers and stemness-related proteins increased in esophageal cancer cells after continuously using chemotherapeutic drugs for a period of time. This study indicated that simultaneously targeting EMT and stemness could be a better strategy for the treatment of esophageal cancer drug resistance.

EphA5 knockdown enhances the invasion and migration ability of esophageal squamous cell carcinoma via epithelial-mesenchymal transition through activating Wnt/\u03b2-catenin pathway

BackgroundThe erythropoietin-producing hepatocellular (Eph) receptor A5 (EphA5) has been found to be overexpressed in some malignant tumors and is associated with disease prognosis. However, the role of EphA5 in esophageal squamous cell carcinoma (ESCC) is not clear.MethodsIn the present study, we measured the expression of EphA5 in ESCC tissues and cell lines including KYSE150 and KYSE450 cells. siRNA transfection was used to interfere with EphA5 expression in ESCC cell lines. Cell viability, colony formation, scratch and invasion assays were performed to explore the roles of EphA5 in ESCC cell lines. Flow cytometry analysis was performed to investigate whether EphA5 could affect the cell apoptosis and cycle. The biomarkers related to epithelial-mesenchymal transition (EMT) and molecules associated with Wnt/β‑catenin signaling were also measured by western blot and immunofluorescence.ResultsThe protein and mRNA expression of EphA5 were significantly higher in fresh ESCC tissues and cell lines compared with normal control groups and human normal esophageal epithelial cells (HEEC). The cell viability assay and colony formation assay revealed that EphA5 knockdown enhanced the proliferation of KYSE150 and KYSE450 cells in vitro. The invasion and migration of ESCC cells were accelerated after EphA5 knockdown. The expression of EMT biomarkers was altered in ESCC cells transfected with siRNA targeting EphA5. Moreover, EphA5 downregulation enhanced the protein levels of β‑catenin and p-GSK-3βSer9, which play a key role in the Wnt/β‑catenin pathway.ConclusionsEphA5 knockdown promotes the proliferation of esophageal squamous cell carcinoma,enhances invasion and migration ability via epithelial-mesenchymal transition through activating Wnt/β‑catenin pathway.

Inhibitor of growth 4 (ING4) inhibits hypoxia-induced EMT by decreasing HIF-1α and snail in HK2 cells

Renal fibrosis is a common mechanism that leads to all kidney diseases and Epithelial-mesenchymal transition (EMT) is considered as one of the potential mechanisms of renal fibrosis. Inhibitor of growth 4 (ING4) was reported to involve in several diseases; especially it was negatively correlated with lung fibrogenesis parameters. However, the role of ING4 and underlying mechanisms in EMT are still unknown. In this study, we used a UUO rat model to mimic renal fibrosis, which was examined by Masson and HE staining analysis. To explore the effects of ING4 on hypoxia-induced EMT, HK2 cells were treated with hypoxia to induce EMT and ING4 was over-expressed in hypoxia-treated HK2 cells by transfection of pEGFP-N1-ING4. MTT assay was used to describe the cell viability of HK2 cells under the hypoxic condition. The expression levels of ING4, hypoxia-inducible factor-1α (HIF-1α), and EMT markers (E-cadherin, N-cadherin and vimentin) were examined in vivo and in vitro by western blot, qRT-PCR, immunohistochemical staining or Immunofluorescence. Our results showed that, in a UUO rat model, ING4 was decreased and EMT was developed with reduction in E-cadherin and increase in N-cadherin and vimentin, suggesting a significant association between ING4 expression and EMT. Under hypoxia, E-cadherin was down-regulated and N-cadherin and vimentin were up-regulated, indicating that hypoxia induced EMT in HK2 cells. Nonetheless, changes in the expression of EMT biomarkers were inhibited by over-expression of ING4. Moreover, over-expressing ING4 decreased the expression of HIF-1α and snail in HK2 cells. These findings suggest that ING4 may inhibit hypoxia-induced EMT via decreasing HIF-1α and snail in HK2 cells, indicating the potential of ING4 as a therapeutic target for renal fibrosis.

Impact of EMT in stage IIIB/IV NSCLC treated with erlotinib and bevacizumab when compared with cisplatin, gemcitabine and bevacizumab.

The aim of the present study was to assess the expression of epithelial-mesenchymal transition biomarkers (E-cadherin and vimentin) and their potential significance as prognostic markers in patients with stage IIIB/IV non-squamous non-small cell lung cancer (NSCLC) enrolled in the INNOVATIONS trial, receiving treatment with either erlotinib/bevacizumab (EB) or cisplatin/gemcitabine/bevacizumab (PGB). The tumor tissues of 104 patients were retrospectively analyzed using immunohistochemistry to assess the expression of E-cadherin and vimentin. The distribution between the treatment arms was 46 patients in the EB-arm and 58 in the PGB-arm. Comparing the treatment arms according to E-cadherin and vimentin expression, the analysis revealed that progression-free survival (PFS) was increased in the PGB treatment group when compared with EB treatment in patients with low expression of E-cadherin [hazard ratio (HR)=0.353; 95% confidence interval (CI) 0.189- 0.658; log-rank P=0.0007] and in those with high expression of vimentin [HR=0.276 (95% CI, 0.115- 0.659), log-rank P=0.0021]. In patients that exhibited high E-cadherin and were negative for vimentin, there was no difference in the PFS between the PGB and EB treatment groups. In conclusion, in non-squamous NSCLC with downregulated E-cadherin and upregulated vimentin, the efficacy of chemotherapy with PGB was superior compared with EB; but the same effect was not observed in patients with high E-cadherin and low vimentin. Although increased PFS was observed in patients with PGB treatment compared with EB treatment in the whole analysis populations, in the subgroup of patients with the mesenchymal phenotype, no prognostic or predictive value of either biomarker could be identified. The potential role of bevacizumab in overcoming chemotherapy resistance in the population with the mesenchymal phenotype has to be further explored.

Small interfering RNA-mediated knockdown of Twist attenuates the aggressive phenotypes of human endometrial carcinoma Ishikawa cells.

The present study aimed to determine the effect of Twist downregulation on the proliferation, apoptosis and migration of human endometrial carcinoma Ishikawa cells. Endogenous expression of the Twist transcription factor was knocked down by delivery of Twist-targeting small interfering RNA (siRNA). Changes in the expression of epithelial-mesenchymal transition biomarkers, namely epithelial (E)-cadherin, neural (N)-cadherin and Twist, were determined by western blot analysis. Cell cycle distribution and apoptosis were evaluated by flow cytometry. Cell proliferation and migration were analyzed using cell-counting and wound-healing assays, respectively. Transfection with Twist siRNA led to a significant reduction in the expression of Twist and N-cadherin (P<0.05), while significantly increasing the expression of E-cadherin, relative to negative control transfectants (all P<0.05). Proliferation was also significantly decreased in Ishikawa cells transfected with Twist siRNA (P<0.05), which was accompanied by an increased rate of apoptosis and cell cycle arrest at S-phase. In addition, Twist downregulation led to a significant reduction in cell migration (P<0.05). These data suggest that Twist serves a role in the regulation of cell proliferation and migration in Ishikawa cells and may represent a potential target for the treatment of human endometrial carcinoma.

WITHDRAWN: MiR-143 suppresses the epithelial-mesenchymal transition of spinal glioblastoma through down-regulation of ERK5

Abstract The spinal glioblastoma (GBM) represents the one of most common tumors in humans. However, the biological processes and molecular mechanisms of spinal GBM are still unclear. It is known that miR-143 participates in the development of various tumor progressions. The present study was to evaluate the level of miR-143 in spinal GBM tissues and cells. We further investigated that the molecular mechanisms of miR-143 in U87 and U251 cell lines. Here, our data showed that the expression levels of miR-143 were significantly reduced in spinal GBM tissues and cell lines. Accordingly, the expression levels of the extracellular signal regulated kinase 5 (ERK5) were significantly increased in spinal GBM tissues and cell lines. Ectopic expression of miR-143 in U87 and U251 cells resulted in decreased cell growth and enhanced cell apoptosis, and inhibited the expression of epithelial-mesenchymal transition (EMT) biomarkers. Further study characterized the 3' untranslated region (3'-UTR) of ERK5 gene as a direct target of miR-143 in U87 and U251 cells as determined by luciferase reporter assays. In addition, the ectopic expression of miR-143 led to the down-regulation of epidermal growth factor receptor (EGFR), while the over-expression of ERK5 reversed the miR-143-inhibited EGFR expression, and promoted cell growth and the expression of EMT biomarkers. In conclusion, this study demonstrated that miR-143 plays a crucial role in regulating the EMT of GBM by directly targeting ERK5, and miR-143 may act as a potential therapeutic target in spinal GBM patients.

Increased Epithelial Expression of CTGF and S100A7 with Elevated Subepithelial Expression of IL-1β in Trachomatous Trichiasis.

PurposeTo characterize the histological appearance and expression of pro-inflammatory mediators, growth factors, matrix metalloproteinases and biomarkers of epithelial-mesenchymal transition (EMT) in healthy control and trachomatous trichiasis (TT) conjunctival tissue.MethodsConjunctival biopsies were taken from 20 individuals with TT and from 16 individuals with healthy conjunctiva, which served as controls. Study participants were of varying ethnicity and were living in a trachoma-endemic region of northern Tanzania. Formalin-fixed paraffin-embedded tissue sections were stained using hematoxylin and eosin or by immunohistochemistry using antibodies against: IL-1β, IL-6, IL-17A, IL-22, CXCL5, S100A7, cleaved caspase 1 (CC1), PDGF, CTGF, TGFβ2, MMP7, MMP9, E-cadherin, vimentin, and αSMA.ResultsTissue from TT cases had a greater inflammatory cell infiltrate relative to controls and greater disruption of collagen structure. CTGF and S100A7 were more highly expressed in the epithelium and IL-1β was more highly expressed in the substantia propria of TT cases relative to controls. Latent TGFβ2 was slightly more abundant in the substantia propria of control tissue. No differences were detected between TT cases and controls in the degree of epithelial atrophy, the number of myofibroblasts or expression of EMT biomarkers.ConclusionsThese data indicate that the innate immune system is active in the immunopathology of trachoma, even in the absence of clinical inflammation. CTGF might provide a direct link between inflammation and fibrosis and could be a suitable target for therapeutic treatment to halt the progression of trachomatous scarring.

Concomitant expression of epithelial-mesenchymal transition biomarkers in breast ductal carcinoma: Association with progression

Epithelial to mesenchymal transition (EMT) is a process implicated in cancer progression in which the underlying cellular changes have been identified mainly using in vitro models. We determined the expression of some putative EMT biomarkers including E-cadherin, beta-catenin, zinc finger factor Snail (Snail), transforming growth factor beta1 (TGFbeta1), TGFbeta type II receptor (TBRII) and the HGF receptor (c-met) and their possible correlation to progression and overall survival in a series of breast ductal carcinoma in situ (DCIS) and invasive ductal carcinomas (IDC). Biomarkers were immunohistochemically determined in 55 IDC specimens from which 21 had lymph node metastases and in 95 DCIS specimens, 46 of these cases associated to invasive carcinoma, in a tissue microarray (TMA). Positive cytoplasmic staining of TGFbeta1 (78.2%), c-met (43.6%), Snail (34.5%), TBRII (100%), membranous E-cadherin (74.5%) and membranous/cytoplasmic beta-catenin (71%) were detected in the IDC samples. Metastatic lymph node samples displayed similar frequencies. A significant increase of c-met and TGFbeta1 positivity along DCIS to IDC progression was noted but only TGFbeta1 positivity was associated with presence of lymph node metastases and advanced stages in IDC. The evaluation of the other EMT markers in DCIS did not show differences in positivity rate as compared to invasive carcinomas. DCIS either pure or associated to IDC showed similar expression of the analyzed biomarkers. All the carcinomas exhibited positive expression of TBRII. Associations between the markers, determined by Spearman's correlation coefficient, showed a significant association between TGFbeta1 and respectively E-cadherin, beta-catenin and c-met in DCIS cases, but in invasive carcinomas only cadherin and catenin were positively correlated. Kaplan-Meier survival curves revealed that none of the EMT biomarkers analyzed were correlated with survival, which was significantly determined only by clinical and hormone receptor parameters.