BackgroundEpithelial-mesenchymal transition (EMT) is a dedifferentiation process that mainly involves in mesenchymal marker upregulation, epithelial maker downregulation and cell polarity loss. Related hypoxia factors play a crucial role in EMT, however, it remains few evidence to clarify the role of HIF-2α in EMT in pancreatic cancer.MethodIn this study, we investigated the expression of HIF-2α and E-cadherin by immunohistochemistry in 70 pancreatic cancer patients, as well as the correlation to the clinicopathologic characteristics. Then we regulated the expression of HIF-2α in pancreatic cancer cells to examine the role of HIF-2α on invasion and migration in vitro. Finally, we tested the relation of HIF-2α and EMT related proteins by Western blot and determined whether HIF-2α regulated EMT through Twist regulating the expression of E-cadherin by Chromatin immunoprecipitation (ChIP) assay.ResultsWe found that HIF-2α protein was expressed positively in 67.1 % (47/70) of pancreatic cancer tissues and 11.4 % (8/70) of adjacent non-tumor pancreatic tissues, and there was a significant difference in the positive rate of HIF-2α protein between two groups (χ2 = 45.549, P < 0.05). In addition, the staining for HIF-2α was correlated with tumor differentiation (P < 0.05), clinical stage (P < 0.05) and lymph node metastasis (P < 0.05), while E-cadherin expression was only correlated with lymph node metastasis (P < 0.05). HIF-2α promoted cell migration, invasion in vitro, and regulated the expression of E-cadherin and MMPs, which are critical to EMT. Our further ChIP assay suggested that only Twist2 could bind to the promoter of E-cadherin in -714 bp region site, but there is no positive binding capacity in -295 bp promoter region site of E-cadherin. Clinical tissues IHC staining showed that Twist2 and E-cadherin expression had an obviously negative correlation in pancreatic cancer. Nevertheless, it had no obvious correlation between Twist1 and E-cadherin.ConclusionThese findings indicated that HIF-2α promotes EMT in pancreatic cancer by regulating Twist2 binding to the promoter of E-cadherin, which meant that HIF-2α and this pathway may be effective therapeutic targets for pancreatic cancer.

No abstract

The expression of microRNA-125b (miR-125b) has been investigated in many human cancers. It has been demonstrated to be downregulated in certain types of cancer, such as bladder cancer, thyroid anaplastic carcinomas, squamous cell carcinoma of the tongue, hepatocellular carcinoma, ovarian and breast cancer. In the present study, we examined the effects of miR-125b on bladder cancer cell migration and invasion. Following transfection of miR-125b, the expression of miR-125b was analyzed in T24 and EJ bladder cancer cells. Additionally, cell migration, cell invasion and luciferase assays, as well as western blot analysis were conducted in the bladder cancer cells. In this study, we demonstrated that miR-125b inhibited cell migration and invasion in T24 and EJ cells. We also provided the first evidence that miR-125b may directly target matrix metalloproteinase 13 (MMP13) in bladder cancer. Our study provided evidence that miR-125b suppresses cell migration and invasion by targeting MMP13 in bladder cancer cell lines. These results suggested that miR-125b could be used for the development of new molecular markers and therapeutic approaches to inhibit bladder cancer metastasis.

MicroRNAs (miRNAs or miRs) are believed to have great potential for use as molecular targets in the diagnosis and treatment of cancer. In this study, we demonstrate that miR-375 is downregulated in pancreatic carcinoma (PC) tissues and PC cell lines. We found that miR-375 negatively regulates the expression of 3-phosphoinositide-dependent protein kinase 1 (PDK1) by directly targeting the 3'UTR of the PDK1 transcript. To investigate the biological roles and the potential mechanisms of action of miR‑375, we induced either the up- or downregulation of miR-375 expression by transfecting various PC cells with miR-375 mimics or an inhibitor. Our results revealed that the upregulation of miR-375 inhibited cell growth and induced cell apoptosis, while the downregulation of miR-375 with the inhibitor had the opposite effect. In addition, our data demonstrate that miR-375 suppresses the malignant behavior of PC cells through the Akt signaling pathway rather than mitogen-activated protein kinase (MAPK) signaling pathways. Taken together, our findings indicate that targeting miR-375 by a genetic approach may provide a novel strategy for the treatment of PC.

The expression of microRNA-99a (miRNA-99a) has been investigated in a number of human cancers. It has been reported to be downregulated in several types of cancer, including ovarian carcinoma, squamous cell carcinoma of the tongue, squamous cell lung carcinoma, hepatocellular carcinoma, bladder cancer, prostate cancer and childhood adrenocortical tumors. In the present study, the effects of miRNA-99a on bladder cancer cell proliferation, migration and invasion were examined. Following transfection of miRNA-99a, cell viability, cell migration assay, cell invasion, western blot analysis and luciferase assays were conducted in bladder cancer cell lines. It was found that miRNA-99a inhibits cell proliferation, migration and invasion in T24 and EJ cells. Additionally, this study provided the first evidence that miRNA-99a is likely to directly target fibroblast growth factor receptor 3 in bladder cancer. The study provided evidence that miRNA-99a suppresses cell proliferation, migration and invasion by targeting growth factor receptor 3 in bladder cancer cell lines. These results indicated that it could be investigated as a target for therapeutic drugs designed to treat bladder cancer.

microRNA (miRNA)-133b has been revealed to be downregulated in head and neck/oral, bladder, human non-small cell lung, colorectal and esophageal squamous cell cancer. The present study examined the expression of miR-133b in renal cell carcinoma (RCC) cell lines and the effects of miRNA-133b on RCC cell proliferation, migration and invasion. Quantitative polymerase chain reaction was used to detect the expression of miR-133b in RCC cell lines. Following transfection of miR-133b, the expression of miR-133b was examined and a cell viability assay, cell migration assay, cell invasion assay, western blot analysis and luciferase assay were conducted in RCC cell lines. The present study revealed that miRNA‑133b was downregulated and inhibited cell proliferation, migration and invasion in 786-O and A498 cells. In addition, to the best of our knowledge, the present study provided the first evidence that miRNA-133b may directly target matrix metallopeptidase 9 (MMP-9) in RCC. The present study also provided evidence that miRNA-133b suppresses cell proliferation, migration and invasion by targeting MMP-9 in RCC cell lines. These results suggested that miRNA-133b may be used for the development of novel molecular markers and therapeutic approaches to inhibit the metastasis of RCC.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most complicated and fatally pathogenic human malignancies. Therefore, there is an urgent need to improve our understanding of the underlying molecular mechanism that drives the initiation, progression, and metastasis of PDAC. The aim of the present study was to identify the key genes and signaling pathways associated with PDAC using bioinformatics analysis. Four transcriptome microarray datasets (GSE15471, GSE55643, GSE62165 and GSE91035) were acquired from Gene Expression Omnibus datasets, which included 226 PDAC samples and 65 normal pancreatic tissue samples. We screened differentially expressed genes (DEGs) with GEO2R and investigated their biological function by Gene Ontology (GO) and Kyoto Encyclopedia of Genes (KEGG) analysis. The overall survival data was obtained from UALCAN, which calculated the data shared with The Cancer Genome Atlas. In addition, a protein-protein interaction (PPI) network of the DEGs was constructed by STRING and Cytoscape software. The four sets of DEGs exhibited an intersection consisting of 205 genes (142 up-regulated and 63 down-regulated), which may be associated with PDAC. GO analysis showed that the 205 DEGs were significantly enriched in the plasma membrane, cell adhesion molecule activity and the Energy pathways, and glycine, serine, threonine metabolism were the most enriched pathways according to KEGG pathway analysis. Kaplan-Meier survival analysis revealed that 22 of 205 common genes were significantly associated with the overall survival of pancreatic cancer patients. In the PPI network and sub-network, DKK1 and HMGA2 were considered as hub genes with high connectivity degrees. DKK1 and HMGA2 are strongly associated with WNT3A and TP53 separately, which indicates that they may play an important role in the Wnt and P53 signaling pathways. Using integrated bioinformatics analysis, we identified DKK1 and HMGA2 as candidate genes in PDAC, which may improve our understanding of the mechanisms of the pathogenesis and integration; the two genes may be therapeutic targets and prognostic markers for PDAC.

Bcl-2 interacting protein 3 (BNIP3) is involved in various cellular processes and is considered a key regulator of hypoxia-induced apoptosis. In the present study, the expression of BNIP3 in pancreatic cancer tissues, the correlation with clinicopathological characteristics and prognosis and the regulation of this protein in pancreatic cancer cell lines with regard to the induction of apoptosis were investigated. BNIP3 expression was significantly lower in pancreatic cancer tissues compared with normal epithelia and was associated with tumor size, clinical stage, and lymph node metastasis. The expression of BNIP3 correlated positively to the proapoptotic protein Bax and negatively to the antiapoptotic protein Bcl-2, whereas the induction of apoptosis by BNIP3 was independent of caspase 3 and 9 activation. The restoration of BNIP3 expression in pancreatic cancer cells in vitro, caused loss of ΔΨm, increase in ROS production, and apoptosis induction. The opposite effect was observed in pancreatic cancer cells, following BNIP3 silencing by RNAi. The absence of BNIP3 expression in pancreatic cancer cells was related to gene methylation that suppressed binding of HIF-1α to the BNIP3 promoter, whereas 5-Aza-2′-deoxycytidine (Aza-dC) treatment restored BNIP3 expression and sensitized pancreatic cancer cells to BNIP3-induced apoptosis. The findings indicated that BNIP3 was significantly downregulated in pancreatic cancer resulting in reduced apoptosis induction. Silencing of BNIP3 expression was associated with methylation of the hypoxia-responsive element (HRE) site that in turn inhibited the binding of HIF-1α to the BNIP3 promoter. The data suggest that BNIP3 reactivation is a potential target for therapeutic intervention against pancreatic cancer.