Overexpression of Snail is associated with lymph node metastasis and poor prognosis in patients with gastric cancer
ABSTRACT: Epithelial-mesenchymal transition (EMT) is in the highlights as a significant role in tumor progression and invasion. Snail was known as the one of regulators of EMT in various malignant tumors. The aim of this study was to investigate the effect of Snail on invasiveness/migratory ability of gastric cancer cell lines and clinicopathological and prognostic significance of Snail over-expression using immunohistochemistry in tissue microarray of 314 gastric adenocarcinomas (GC). Differential gene expression in Snail over-expressed GC was investigated using cDNA microarray analysis. Silencing of Snail by ShRNA induced decreased of invasion, migration of gastric cancer cell lines. In contrast, over-expression of Snail induced increased invasiveness and migratory ability of gastric cancer cell lines in accordance increase of VEGF and MMP11. Furthermore, the over-expression of Snail (≥75% nuclear staining of Snail) was significantly associated with tumor progression (p<0.0001), lymph node metastases (p=0.002), lymphovascular invasion (p=0.002), and perineural invasion (p=0.002) in 314 GC patient. Snail over-expression was also associated with poor prognosis (shorter survival) in GC patients (p=0.023). cDNA microarray revealed 213 differential expressed genes in Snail over-expressed GC tissues, including genes related to metastasis, invasion. Based on above results, it was suggested that Snail plays a significant role in invasiveness/migratory ability of GCs. In addition, Snail might be used to predictive biomarker for evaluation of prognosis or aggressiveness in GCs. 48 primary gastric adenocarcinoma fresh frozen tissues were used for microarray. All the tissues were obtained after curative resection after pathologic confirm at Pusan National University Hospital (PNUH, Busan, Korea) and Cheonnam University Hospital (CNUH, Cheonnam, Korea). Microarray experiment and data analysis were done at Cancer research institute, PNUH, Busan, Korea.
Project description:Epithelial-mesenchymal transition (EMT) is in the highlights as a significant role in tumor progression and invasion. Snail was known as the one of regulators of EMT in various malignant tumors. The aim of this study was to investigate the effect of Snail on invasiveness/migratory ability of gastric cancer cell lines and clinicopathological and prognostic significance of Snail over-expression using immunohistochemistry in tissue microarray of 314 gastric adenocarcinomas (GC). Differential gene expression in Snail over-expressed GC was investigated using cDNA microarray analysis. Silencing of Snail by ShRNA induced decreased of invasion, migration of gastric cancer cell lines. In contrast, over-expression of Snail induced increased invasiveness and migratory ability of gastric cancer cell lines in accordance increase of VEGF and MMP11. Furthermore, the over-expression of Snail (?75% nuclear staining of Snail) was significantly associated with tumor progression (p<0.0001), lymph node metastases (p=0.002), lymphovascular invasion (p=0.002), and perineural invasion (p=0.002) in 314 GC patient. Snail over-expression was also associated with poor prognosis (shorter survival) in GC patients (p=0.023). cDNA microarray revealed 213 differential expressed genes in Snail over-expressed GC tissues, including genes related to metastasis, invasion. Based on above results, it was suggested that Snail plays a significant role in invasiveness/migratory ability of GCs. In addition, Snail might be used to predictive biomarker for evaluation of prognosis or aggressiveness in GCs. 48 primary gastric adenocarcinoma fresh frozen tissues were used for microarray. All the tissues were obtained after curative resection after pathologic confirm at Pusan National University Hospital (PNUH, Busan, Korea) and Cheonnam University Hospital (CNUH, Cheonnam, Korea). Microarray experiment and data analysis were done at Cancer research institute, PNUH, Busan, Korea.
Project description:Epithelial-mesenchymal transition (EMT) plays a significant role in tumor progression and invasion. Snail is a known regulator of EMT in various malignant tumors. This study investigated the role of Snail in gastric cancer.We examined the effects of silenced or overexpressed Snail using lenti-viral constructs in gastric cancer cells. Immunohistochemical analysis of tissue microarrays from 314 patients with gastric adenocarcinoma (GC) was used to determine Snail's clinicopathological and prognostic significance. Differential gene expression in 45 GC specimens with Snail overexpression was investigated using cDNA microarray analysis.Silencing of Snail by shRNA decreased invasion and migration in GC cell lines. Conversely, Snail overexpression increased invasion and migration of gastric cancer cells, in line with increased VEGF and MMP11. Snail overexpression (?75% positive nuclear staining) was also significantly associated with tumor progression (P?<?0.001), lymph node metastases (P?=?0.002), lymphovascular invasion (P?=?0.002), and perineural invasion (P?=?0.002) in the 314 GC patients, and with shorter survival (P?=?0.023). cDNA microarray analysis revealed 213 differentially expressed genes in GC tissues with Snail overexpression, including genes related to metastasis and invasion.Snail significantly affects invasiveness/migratory ability of GCs, and may also be used as a predictive biomarker for prognosis or aggressiveness of GCs.
Project description:Epidermal growth factor-like domain-containing protein 7 (EGFL7) is upregulated in human epithelial tumors and so is a potential biomarker for malignancy. Indeed, previous studies have shown that high EGFL7 expression promotes infiltration and metastasis of gastric carcinoma. The epithelial-mesenchymal transition (EMT) initiates the metastatic cascade and endows cancer cells with invasive and migratory capacity; however, it is not known if EGFL7 promotes metastasis by triggering EMT. We found that EGFL7 was overexpressed in multiple human gastric cancer (GC) cell lines and that overexpression promoted cell invasion and migration as revealed by scratch wound and transwell migration assays. Conversely, shRNA-mediated EGFL7 knockdown reduced invasion and migration. Furthermore, EGFL7-overexpressing cells grew into larger tumors and were more likely to metastasize to the liver compared to underexpressing CG cells following subcutaneous injection in mice. EGFL7 overexpression protected GC cell lines against anoikis, providing a plausible mechanism for this enhanced metastatic capacity. In excised human gastric tumors, expression of EGFL7 was positively correlated with expression levels of the mesenchymal marker vimentin and the EMT-associated transcription repressor Snail, and negatively correlated with expression of the epithelial cell marker E-cadherin. In GC cell lines, EGFL7 knockdown reversed morphological signs of EMT and decreased both vimentin and Snail expression. In addition, EGFL7 overexpression promoted EGF receptor (EGFR) and protein kinase B (AKT) phospho-activation, effects markedly suppressed by the EGFR tyrosine kinase inhibitor AG1478. Moreover, AG1478 also reduced the elevated invasive and migratory capacity of GC cell lines overexpressing EGFL7. Collectively, these results strongly suggest that EGFL7 promotes metastasis by activating EMT through an EGFR-AKT-Snail signaling pathway. Disruption of EGFL7-EGFR-AKT-Snail signaling may a promising therapeutic strategy for gastric cancer.
Project description:Gastric cancer (GC) is among the most fatal cancers in China. MicroRNAs (miRNAs) are versatile regulators during GC development and progression. miR-491-5p has been demonstrated to act as a tumor suppressor in several types of cancer. However, the role of miR-491-5p in GC metastasis remains unknown. Here, we found that miR-491-5p was significantly decreased in GC tissues compared with adjacent non-cancerous tissues, and low miR-491-5p level was associated with large tumor size. Overexpression of miR-491-5p significantly suppressed GC cell epithelial-to-mesenchymal transition (EMT) and tumor metastasis in vitro and in vivo. Mechanistically, SNAIL was identified as a direct target of miR-491-5p. The silencing of SNAIL phenocopied the tumor suppressive function of miR-491-5p, whereas re-expression of SNAIL in GC cells rescued the EMT markers and cell migratory ability that were inhibited by miR-491-5p. In addition, miR-491-5p inhibited FGFR4 indirectly. Inhibition of FGFR4 also decreased the SNAIL level and impaired EMT and cell migration. Taken together, these findings indicate that downregulation of miR-491-5p promoted GC metastasis by inducing EMT via regulation of SNAIL and FGFR4.
Project description:Rho GDP dissociation inhibitor 2 (RhoGDI2) expression correlates with tumor growth, metastasis, and chemoresistance in gastric cancer. Here, we show that RhoGDI2 functions in the epithelial-mesenchymal transition (EMT), which is responsible for invasiveness during tumor progression. This tumorigenic activity is associated with repression of E-cadherin by RhoGDI2 via upregulation of Snail. Overexpression of RhoGDI2 induced phenotypic changes consistent with EMT in gastric cancer cells, including abnormal epithelial cell morphology, fibroblast-like properties, and reduced intercellular adhesion. RhoGDI2 overexpression also resulted in decreased expression of the epithelial markers E-cadherin and ?-catenin and increased expression of the mesenchymal markers vimentin and fibronectin. Importantly, RhoGDI2 overexpression also stimulated the expression of Snail, a repressor of E-cadherin and inducer of EMT, but not other family members such as Slug or Twist. RNA interference-mediated knockdown of Snail expression suppressed RhoGDI2-induced EMT and invasion, confirming that the effect was Snail-specific. These results indicate that RhoGDI2 plays a critical role in tumor progression in gastric cancer through induction of EMT. Targeting RhoGDI2 may thus be a useful strategy to inhibit gastric cancer cell invasion and metastasis.
Project description:MicroRNAs (miRNAs) deregulation is frequent in human gastric cancers (GCs), but the role of specific miRNAs involved in this disease remains elusive. MiR-22 was previously reported to act as tumor suppressors or oncogenes in diverse cancers. However, their accurate expression, function and mechanism in GC are largely unclear. Here, we found that the expression of miR-22 was significantly reduced in clinical GC tissues compared with paired adjacent normal tissues, and was significantly correlated with a more aggressive phenotype of GC in patients, and miR-22 low expression correlated with poor overall survival. The introduction of miR-22 markedly suppressed GC cell growth, migration and invasion, and inhibition of miR-22 promoted GC cell proliferation, migration and invasion in vitro. We further demonstrated that miR-22 acted as tumor suppressors through targeting extracellular matrix (ECM) remodeling member matrix metalloproteinase 14 (MMP14) and epithelial-to-mesenchymal transition (EMT) inducer Snail in GC. Moreover, ectopic expression of MMP14 or Snail restored inhibitory effects of miR-22 on cell migration and invasion in GC cells, and a negative relationship between the miR-22 expression and MMP14 or Snail mRNA levels was observed in GC. Finally, overexpression of miR-22 suppressed tumor growth, peritoneal dissemination and pulmonary metastasis in vivo. Taken together, we identified that miR-22 is a potent tumor suppressor in GC. MiR-22 downregulation promotes GC invasion and metastasis by upregulating MMP14 and Snail, and then inducing ECM remodeling and EMT. These findings provide a better understanding of the development and progression of GC and may be an important implication for future therapy of the GC.
Project description:<h4>Background</h4>Over-expression of long non-coding RNA HOTAIR has been reported in several types of cancer. Yet its involvement in gastric cancer (GC) has not been well understood. The aim of present study was to examine the expression pattern of HOTAIR in GC patients, then, explore its role in promoting cancer invasion and underlying molecular mechanism.<h4>Methods</h4>The expression level of HOTAIR in the tumor specimens of GC patients was quantified by Realtime RT-PCR. The correlation between HOTAIR level and clinicopathological factors as well as prognosis was then examined. Down-regulation of HOTAIR by RNA interference was applied to investigate its roles in tumor invasiveness via the view of Epithelial-to-mesenchymal transition (EMT).<h4>Results</h4>The expression level of HOTAIR in cancer tissues was higher than that in adjacent noncancerous tissues. Expression level of HOTAIR was significantly correlated with lymph node metastasis and TNM stage. Furthermore, high expression level of HOTAIR was a predictor of poor over-all survival in GC patients. In vitro, inhibition of HOTAIR in GC cells could reduce invasiveness, as well as the expression of MMP1 and MMP3. In addition, suppression of HOTAIR could reverse EMT process.<h4>Conclusions</h4>HOTAIR could act as a potential predictor for over-all survival in patients with GC. Inhibition of HOTAIR could reduce invasiveness and reverse EMT process in GC cells, indicating the potential role of HOTAIR in GC diagnostics and therapeutics.
Project description:Aberrant expression of neuropilin and tolloid-like 2 (NETO2) has been observed during the progression of some human carcinomas. However, the expression pattern and clinical relevance of NETO2 in gastric cancer (GC) remain to be elucidated. In this study, we found that NETO2 expression was higher in GC tissues compared with paired non-cancerous tissues. Moreover, the expression of NETO2 was positively correlated with clinical stage, invasion depth, lymph node metastasis, and tumor size, but inversely correlated with overall and disease-free survival rates. Cox regression analysis identified NETO2 as an independent prognostic indicator for GC patients. Overexpression of NETO2 facilitated migration and invasion of GC cells in vitro and metastasis in vivo in association with induction of epithelial-mesenchymal transition. Conversely, knockdown of NETO2 had the opposite effects. Mechanistically, silencing NETO2 reduced the phosphorylation of PI3K, AKT, and NF-?B p65 as well as the expression of Snail, whereas NETO2 overexpression achieved the opposite results. Furthermore, we identified TNFRSF12A as a mediator for NETO2 to activate PI3K/AKT/NF-?B/Snail axis. Collectively, our results demonstrate that NETO2 promotes invasion and metastasis of GC cells and represents a novel prognostic indicator as well as a potential therapeutic target in GC.
Project description:Background:Gastric cancer (GC) is one of the most common gastrointestinal malignancies worldwide. Emerging evidence indicates that hyperglycemia promotes tumor progression, especially the processes of migration, invasion and epithelial-mesenchymal transition (EMT). However, the underlying mechanisms of GC remain unclear. Method:Data from the Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases were used to detect the expression of glycolysis-related enzymes and EMT-related transcription factors. Small interfering RNA (siRNA) transfection was performed to decrease ENO1 expression. Immunohistochemistry (IHC), Western blot and qRT-PCR analyses were used to measure gene expression at the protein or mRNA level. CCK-8, wound-healing and Transwell assays were used to assess cell proliferation, migration and invasion. Results:Among the glycolysis-related genes, ENO1 was the most significantly upregulated in GC, and its overexpression was correlated with poor prognosis. Hyperglycemia enhanced GC cell proliferation, migration and invasion. ENO1 expression was also upregulated with increasing glucose concentrations. Moreover, decreased ENO1 expression partially reversed the effect of high glucose on the GC malignant phenotype. Snail-induced EMT was promoted by hyperglycemia, and suppressed by ENO1 silencing. Moreover, ENO1 knockdown inhibited the activation of transforming growth factor ? (TGF-?) signaling pathway in GC. Conclusions:Our results indicated that hyperglycemia induced ENO1 expression to trigger Snail-induced EMT via the TGF-?/Smad signaling pathway in GC.
Project description:Trametes robiniophila Murr. (Huaier) is a widely used anti-cancer agent in China. Strong evidence for the anti-proliferative activity of Huaier has been reported; however, its anti-metastatic potential against gastric cancer (GC) as well as its underlying mechanism of action are unknown. Here, we show that treatment with an aqueous Huaier extract over a range of concentrations significantly suppressed both the invasiveness and migratory ability of GC cells. Huaier could also partly reverse the epithelial-mesenchymal transition (EMT), as characterized by increased expression of the epithelial marker E-cadherin and decreased expression of the mesenchymal markers N-cadherin and vimentin. In addition, Huaier-treated cells expressed lower levels of Twist compared to untreated controls, and overexpression of Twist via transfection could partially abolish the anti-metastatic activity of Huaier. Furthermore, elevated Twist expression was correlated with an advanced TNM stage, a high rate of lymph node metastasis, and reduced disease-free survival in GC patients. These findings reveal a novel anti-metastatic mechanism for Huaier, which inhibits the EMT by targeting Twist, suggesting its potential application against a GC relapse.