ESL attenuates BLM-induced IPF in mice: Dual mediation of the TLR4/NF-κB and TGF-ß1/PI3K/Akt/FOXO3a pathways.

BACKGROUNDS: Idiopathic pulmonary fibrosis (IPF) is a persistent and advanced pulmonary ailment. The roles of innate immunity and adaptive immunity are pivotal in the evolution of IPF. An ill-adjusted interaction between epithelial cells and immune cells is responsible for initiating the epithelial-mesenchymal transition (EMT) process and sustaining chronic inflammation, thereby fostering fibrosis progression. The intricacy of IPF pathogenesis has hindered the availability of efficacious agents. Elephantopus scaber Linn. (ESL) is a canonical Chinese medicine with significant immunoregulatory effects, and its aqueous extract has been proven to attenuate IPF symptoms in bleomycin (BLM)-induced mice. However, the underlying mechanism through which ESL relieves IPF remains unclear. AIM: To validate whether ESL reverses IPF by mediating the immune response and EMT. METHODS: Ultra-performance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS/MS) and UPLC were used to identify the components and determine the concentrations of the specific compounds in the ESL. Network pharmacology and molecular docking were applied to predict the potential mechanism underlying the anti-IPF effect of ESL. BLM-induced IPF mice were used to validate the anti-IPF effect of ESL, and lung tissue was collected to test putative pathways involved in inflammation and EMT via immunohistochemistry (ICH), real-time quantitative polymerase chain reaction (RT-qPCR) and Western blotting. RESULTS: Sixty-one compounds were identified, and thirteen main ingredients were quantified in the ESL. In silico experiments predicted that the IPF-mediated reversal of adverse effects by ESL would be related to interruption of the Toll-like receptor 4 (TLR4)/nuclear factor-k-gene binding (NF-ĸB) inflammatory pathway and the transforming growth factor-beta l (TGF-ß1)/phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/forkhead box O3 (FOXO3a) fibrosis pathway. In vivo experiments showed that ESL alleviates BLM-induced lung inflammation and fibrosis by reducing neutrophil aggregation and fibroblast foci, similar to the effects of the positive control drug pirfenidone (PFD). ESL markedly inhibited the transcription of TNF-α, IL-1ß, and IL-6, which are downstream genes of the NF-κB signaling pathway. Furthermore, the protein levels of TLR4 and p-NF-κB were correspondingly inhibited in response to ESL treatment. Additionally, ESL reverses BLM-induced changes in the expression of EMT-related biological characteristic indicators (collagen I [COLIA1], E-cadherin, and alpha smooth muscle actin [α-SMA]) at the messenger ribonucleic acid (mRNA) level and markedly inhibits the expression of EMT-related upstream proteins (TGF-ß1, p-PI3K, p-Akt, and p-FOXO3a). CONCLUSION: Our research suggested that ESL attenuates BLM-induced IPF through mediating the EMT process via the TGF-ß1/PI3K/Akt/FOXO3a signaling pathway and inhibiting inflammation through the TLR4/NF-κB signaling pathway, highlighting that ESL can serve as an immunoregulator for relieving the abnormal immune response and reversing the EMT in IPF.

Network Pharmacological Analysis and Experimental Validation of the Effects of Silybin on Proliferation, Migration, and Immunotherapy of Papillary Thyroid Cancer.

AIM: The study aimed to use network pharmacology research and in vitro experiments to investigate the material basis and molecular mechanisms of silybin in the treatment of papillary thyroid carcinoma. BACKGROUND: Papillary thyroid cancer (PTC) has a decent prognosis; however, recurrence and metastasis are the leading causes of death in patients with PTC. A key research focus in thyroid cancer treatment is the inhibition of PTC proliferation, invasion, and migration. Silybin, the major active element in the traditional Chinese herb silymarin, has been used to treat a range of diseases, including cancer, but no study has been undertaken to determine whether it can help prevent PTC. OBJECTIVE: In this study, we attempted to determine through network pharmacology and in vitro experiments if silybin inhibits the development of papillary thyroid cancer by inhibiting cell cycle and invasive migration. METHODS: To predict the probable targets and underlying mechanisms of silybin against PTC, a network pharmacology research was performed. In vitro experiments were conducted to further evaluate silybin's anti-cancer properties and priority targets against PTC. RESULTS: The datasets revealed a total of 489 silybin targets acting on PTC, and functional enrichment analysis suggested that the target genes were enriched in functions and pathways related to PTC development, invasion, migration, and immunotherapy. By constructing these target PPI networks, the seven hub genes, fibronectin 1 (FN1), tissue inhibitor of metalloproteinases 1 (TIMP1), N-cadherin (CDH2), collagen type III alpha 1 chain (COL3A1), cyclin D1 (CCND1), AP-1 transcription factor subunit (JUN), and hepatocyte growth factor receptor (MET) were found. These hub genes were determined to be highly linked to a worse clinicopathological form, a higher risk of metastatic recurrence, and a worse prognosis of PTC. The common immunological checkpoint gene expression levels were positively correlated with the expression levels of the hub genes. Silybin decreased the proliferative and metastatic capacity of PTC cells, according to in vitro investigations. When PTC was treated with silybin, the FN1/AKT signaling pathway was blocked, CCND1 expression was reduced, and CDH2, Vimentin, Snail, Slug and PD-L1 expressions were dramatically reduced, while E-cadherin expression was significantly elevated. CONCLUSION: These findings provide preliminary evidence that silybin inhibits PTC cell proliferation, metastasis, and invasion by altering the FN1/AKT signaling pathway and inhibiting the EMT process. Silybin can reverse immunosuppression in papillary thyroid cancer by affecting immunological checkpoint gene expression levels. These studies provide a theoretical and experimental scientific basis for the potential anticancer effects of silybin on PTC.

IGF2 is upregulated by its antisense RNA to potentiate pancreatic cancer progression.

Pancreatic cancer is a deadly cancer. More and more long noncoding RNAs (lncRNAs) have received confirmation to be dysregulated in tumors and exert the regulatory function. Studies have suggested that lncRNA insulin-like growth factor 2 antisense RNA (IGF2-AS) participates in the development of some cancers. Thus, we attempted to clarify its function in pancreatic cancer. Reverse-transcription quantitative polymerase chain reaction was applied for testing IGF2-AS expression in pancreatic cancer cells. Colony formation and Transwell wound experiments were applied for determining cell proliferative, migratory, and invasive capabilities. The alteration of epithelial-mesenchymal transition (EMT)-related gene level was tested via western blot. The mice model was established for measuring the tumor growth and metastasis. RIP validated the interaction of RNAs. IGF2-AS displays high expression in pancreatic cancer cells. IGF2-AS depletion repressed PC cell proliferative, migratory, invasive capabilities, and EMT process. Furthermore, pancreatic cancer tumor growth and metastasis were also inhibited by IGF2-AS depletion. Additionally, IGF2-AS positively regulated IGF2 level via recruiting HNRNPC. IGF2 overexpression counteracted the functions of IGF2-AS deficiency on pancreatic cancer cell behaviors. Moreover, IGF2R deletion was found to inhibit the positive effect of IGF2 on pancreatic cancer progression. IGF2-AS potentiates pancreatic cancer cell proliferation, tumor growth, and metastasis by recruiting HNRNPC via the IGF2-IGF2R regulatory pathway. These discoveries might offer a novel insight for treatment of PC, which may facilitate targeted therapies of PC in clinical practice.

The inhibitory effect of LINC00261 upregulation on the pancreatic cancer EMT process is mediated by KLF13 via the mTOR signaling pathway

PurposeThe long noncoding RNA LINC00261 was reported to be involved in carcinogenesis and has been validated as a tumor suppressor in pancreatic cancer (PC); however, how LINC00261 is regulated has not been fully examined. Here, we attempted to investigate the upstream and downstream targets of LINC00261 in PC.MethodsLINC00261 expression in PC tissues was examined by the Gene Expression Omnibus (GEO) datasets and the Gene Expression Profiling Interactive Analysis (GEPIA) database. The quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays were performed to detect the expression level of LINC00261 in PC cells. The location of LINC00261 in PC cells was identified by RNA fluorescence in situ hybridization (RNA-FISH). Cell Counting Kit-8 (CCK-8), cell apoptosis assay, transwell invasion and migration assays testified the critical role of LINC00261 in PC. The luciferase reporter assay was applied to confirm the binding of LINC00261 to its upstream transcription factor KLF13. The changes in LINC00261 related target protein levels were analyzed by Western blotting assay.ResultsLINC00261 was significantly lower in PC tissues and was mainly concentrated in the nucleus. Overexpression of LINC00261 inhibited the invasion and migration of PC cells. Mechanistically, transcription factor KLF13 was confirmed to inhibit the epithelial-mesenchymal transition (EMT) process of PC cells by promoting the transcription of LINC00261 and suppressing the expression of metastasis-associated proteins, such as matrix metalloproteinase MMP2 and vimentin, thus inhibiting the metastasis of PC.ConclusionLINC00261 regulates PC cell metastasis through the “KLF13-LINC00261-mTOR-P70S6K1-S6” signaling pathway, which provides a significant set of potential PC therapeutic targets.

USP20 regulates the stability of EMT transcription factor SOX4 and influences colorectal cancer metastasis.

BACKGROUND: Colorectal cancer (CRC) is a familiar malignancy accompanied by higher morbidity and mortality. The deubiquitination enzyme USP20 has been discovered to be one key factor in several cancers progression. SOX4 is a critical transcription factor to regulate the expression of various genes, and participates into the occurrence and progression of cancers. In this study, it was aimed to illustrate the role of USP20 and the regulatory relationship between USP20 and SOX4 in CRC. METHODS: The protein expressions of USP20, SOX4, E-cadherin, N-cadherin, Snail and slug were tested through western blot. The cell proliferation ability was verified through CCK-8 assay. The migration and invasion abilities were detected through Transwell assay. The mRNA expression of SOX4 was confirmed through RT-qPCR. The interaction between USP20 and SOX4 was notarized through Co-IP assay. RESULT: Our study demonstrated that USP20 displayed higher expression, and facilitated CRC progression through regulating cell proliferation, migration, invasion and EMT process markers. USP20 was found to modulate SOX4 protein expression. Next, it was verified that USP20 regulated SOX4 degradation through deubiquitination. Finally, through rescue assays, we revealed that USP20 mediated SOX4 expression to accelerate CRC progression. CONCLUSIONS: In this study, USP20 regulated the stability of EMT transcription factor SOX4 and aggravated colorectal cancer metastasis. This finding might highlight the function of USP20 in the treatment of CRC.

The inhibitory effect of LINC00261 upregulation on the pancreatic cancer EMT process is mediated by KLF13 via the mTOR signaling pathway.

PURPOSE: The long noncoding RNA LINC00261 was reported to be involved in carcinogenesis and has been validated as a tumor suppressor in pancreatic cancer (PC); however, how LINC00261 is regulated has not been fully examined. Here, we attempted to investigate the upstream and downstream targets of LINC00261 in PC. METHODS: LINC00261 expression in PC tissues was examined by the Gene Expression Omnibus (GEO) datasets and the Gene Expression Profiling Interactive Analysis (GEPIA) database. The quantitative reverse transcription polymerase chain reaction (qRT-PCR) assays were performed to detect the expression level of LINC00261 in PC cells. The location of LINC00261 in PC cells was identified by RNA fluorescence in situ hybridization (RNA-FISH). Cell Counting Kit-8 (CCK-8), cell apoptosis assay, transwell invasion and migration assays testified the critical role of LINC00261 in PC. The luciferase reporter assay was applied to confirm the binding of LINC00261 to its upstream transcription factor KLF13. The changes in LINC00261 related target protein levels were analyzed by Western blotting assay. RESULTS: LINC00261 was significantly lower in PC tissues and was mainly concentrated in the nucleus. Overexpression of LINC00261 inhibited the invasion and migration of PC cells. Mechanistically, transcription factor KLF13 was confirmed to inhibit the epithelial-mesenchymal transition (EMT) process of PC cells by promoting the transcription of LINC00261 and suppressing the expression of metastasis-associated proteins, such as matrix metalloproteinase MMP2 and vimentin, thus inhibiting the metastasis of PC. CONCLUSION: LINC00261 regulates PC cell metastasis through the "KLF13-LINC00261-mTOR-P70S6K1-S6" signaling pathway, which provides a significant set of potential PC therapeutic targets.

Thioredoxin Reductase-1 as a Potential Biomarker in Fibroblast-Associated HCT116 Cancer Cell Progression and Dissemination in a Zebrafish Model.

The tumor microenvironment, especially that of fibroblasts, strongly promotes colorectal cancer (CRC) progression. Progressive cancers usually accumulate high reactive oxygen species (ROS), leading to oxidative stress. The stress relates to the expression of thioredoxin reductase-1 (TrxR-1), which is an oxidative stress sensitivity molecule. This study aimed to investigate TrxR-1 expression as an indication of colon-fibroblast-inducing colorectal cancer progression and metastasis. We found that the high proliferative fibroblast-cultured media (FCM) contained pro-inflammatory cytokines that have a high ability to influence HCT116 and CRC cell progression, when compared with complete media (CM) as a control in terms of growth (CM = 100.00%, FCM = 165.96%), migration (CM = 32.22%, FCM = 83.07%), invasion (CM = 130 cells/field, FCM = 449 cells/field), and EMT transformation while decreasing E-cadherin expression (CM = 1.00, FCM = 0.69) and shape factor (CM = 0.94, FCM = 0.61). In addition, the overexpression of TrxR-1 is associated with cellular oxidant enchantment in FCM-treated cells. A dot plot analysis showed a strong relation between the EMT process and the overexpression of TrxR-1 in FCM-treated cells (CM = 13/100 cells, FCM = 45/100 cells). The cancer transplantation of the adult zebrafish model illustrated a significantly higher number of microtumors in FCM-treated cells (CM = 4.33 ± 1.51/HPF, FCM = 25.00 ± 13.18/HPF) disseminated in the intraperitoneal cavity with TrxR-1 positive cells. The overexpression of TrxR-1 indicated fibroblast-associated CRC progression in HCT116 cells and the zebrafish model. Therefore, TrxR-1 could be applied as a novel biomarker for colorectal cancer progression and prognostic evaluation.

PADI1 contributes to EMT in PAAD by activating the ERK1/2-p38 signaling pathway.

BACKGROUND: Peptidylarginine deiminase 1 (PADI1) has been reported to promote tumorigenesis in breast cancer. However, the functional role of PADI1 in pancreatic ductal adenocarcinoma (PAAD) has remained elusive until now. METHODS: The expression pattern of PADI1 in PAAD tissues and normal tissues was analyzed using The Cancer Genome Atlas (TCGA) dataset. A Kaplan-Meier curve analysis was performed to evaluate the prognostic value of PADI1 in PAAD patients. PADI1 was knocked down in CFPAN-1 and HPAC cells, and overexpressed in PANC-1 and Bxpc-3 cells by RNA interference. A wound-healing assay was performed to analyze relative cell migration distance. Cell migration and invasion were assessed by a Transwell assay. Related protein expression levels were measured by western blot and immunofluorescence. RESULTS: The bioinformatics analysis showed that PADI1 was overexpressed in PAAD tissues and associated with a poor survival prognosis. The knockdown of PADI1 suppressed cell migration and invasion, and activated the ERK1/2-p38 signaling pathway in CFPAN-1 and HPAC cells. The overexpression of PADI1 produced the opposite results in PANC-1 and Bxpc-3 cells. Additionally, treatment with an MEK1/2 inhibitor significantly attenuated the effects of PADI1 knockdown on cell migration, invasion, the epithelial-mesenchymal transition (EMT) process, and p-ERK1/2 and p38 expression in CFPAN-1 and HPAC cells. CONCLUSIONS: Our data suggested that PADI1 may function as an oncogene in regulating metastasis in vitro in PAAD.

Human/eukaryotic ribosomal protein L14 (RPL14/eL14) overexpression represses proliferation, migration, invasion and EMT process in nasopharyngeal carcinoma.

Although human/eukaryotic ribosomal protein L14 (RPL14/eL14) is known to be associated with a variety of cancers, its role in nasopharyngeal carcinoma (NPC) remains unclear. The aim of this study was to explore the impact of RPL14(eL14) in NPC. The results of quantitative real-time polymerase chain reaction (qRT-PCR), western blot, and immunohistochemical staining revealed that the expression of RPL14(eL14) significantly reduced in NPC tissues and cells. Furthermore, the protein expression of RPL14(eL14) was linked to NPC-related clinical pathological features, including the T and N classification of Tumor Node Metastasis (TNM) staging (all p < 0.05). Cell counting kit-8 (CCK-8) assay and colony formation assay revealed that RPL14(eL14) overexpression repressed NPC cell proliferation. In cell cycle assay, RPL14(eL14) overexpression significantly blocked NPC cells in S phase. Overexpression of RPL14(eL14) repressed cell migration and invasion in NPC as shown by transwell assay and cell scratch healing assay. In addition, RPL14(eL14) was closely correlated with the expression of epithelial-mesenchymal transition (EMT) biomarkers, including E-cadherin, N-cadherin, and vimentin as detected by western blot. In conclusion, our results revealed that RPL14(eL14) may be considered as an antioncogene in NPC, which greatly suppresses cancer progression.

Limb-bud and heart (LBH) inhibits cellular migration, invasion and epithelial-mesenchymal transition in nasopharyngeal carcinoma via downregulating αB-crystallin expression.

Limb-bud and heart (LBH) gene has received increasing attention in recent cancer studies. Here we investigated the role of the LBH gene in regulating the metastasis capacity and epithelial-mesenchymal transition (EMT) of nasopharyngeal carcinoma (NPC) cells, and its potential mechanism. The expressions of LBH and αB-crystallin (CRYAB) were modulated by lentiviral infection, or plasmid/siRNA transfection, and the phosphorylation of p38 was suppressed by an inhibitor, to explore their functions in modulating NPC cell phenotypes, as well as the relationships of these factors with each other. Cellular proliferation, migration and invasion were examined by RTCA system, Transwell assays and Matrigel Transwell assays respectively. The EMT progression was indicated by RT-qPCR and Western blotting measuring the expressions of EMT biomarkers. NPC xenografts were constrcucted, and formed tumors were sectioned for morphology and immunohistofluorescence. The interaction between LBH and CRYAB was examined by colocalization and Fluorescence resonance energy transfer (FRET) analysis. We reached the conclusion that LBH inhibits the proliferation, migration, invasion and EMT of NPC cells, and its effects were partially achieved by suppressing p38 phosphorylation, which subsequently downregulates the mRNA expression and phosphorylation of CRYAB, while CRYAB directly interacts with LBH in NPC cells. This LBH-related pathway we revealed provides a novel therapeutic target for nasopharyngeal carcinoma research.

Gene expression profile in metastatic and non-metastatic parathyroid carcinoma.

Parathyroid carcinoma (PC) is one of the rarest and aggressive malignancies of the endocrine system. In some instances, the histological diagnosis remains uncertain unless there is evidence of gross local invasion or secondary spread. The identification of molecular markers could improve the diagnostic accuracy of these lesions. The expression of 740 genes involved in the tumor progression processes was assessed in 8 parathyroid adenomas (PAs), 17 non-metastatic and 10 metastatic PCs using NanoString technology. Clustering analysis and Ingenuity Pathway Analysis (IPA) were interrogated to compare the gene expression profiles among the three analyzed groups and to evaluate the potential role of differentially expressed genes, respectively. The 103 differentially expressed genes between metastatic PCs and PAs are able to discriminate perfectly the two groups from a molecular point of view. The molecular signatures identified in non-metastatic PCs vs PAs and in metastatic PCs vs non-metastatic PCs comparisons, although with some exceptions, seem to be histotype-specific IPA reveals that hepatic fibrosis/hepatic stellate cell activation and GP6 signaling pathway are involved in malignant behavior of parathyroid tumors, whereas the activation of the HOTAIR regulatory pathway are involved in the metastatization process. Our investigation identified differentially expressed genes in non-metastatic PCs mainly encoding ECM proteins and in metastatic PCs driving endothelial-to-mesenchymal transition or encoding mediators of angiogenesis. The identified genes might be promising molecular markers potentially useful in the clinical practice for the early diagnosis and prognosis of PC.

Long non-coding RNA AGAP2-AS1 accelerates cell proliferation, migration, invasion and the EMT process in colorectal cancer via regulating the miR-4,668-3p/SRSF1 axis.

BACKGROUND: Colorectal cancer (CRC) is a frequently occurring tumor. Although a number of long noncoding RNAs have been researched in CRC, the expression, function and mechanism of AGAP2-AS1 remains poorly investigated. METHODS: Gene expression was analyzed by a quantitative reverse transcriptase-polymerase chain rreaction and western blot analyses. Cell counting kit-8, colony formation and Transwell assays were utilized to explore the functional role of AGAP2-AS1 in CRC. Luciferase reporter, RNA pull down and RNA immunoprecipitation assays were implemented to verify relationships between RNA molecules. RESULTS: In the present study, AGAP2-AS1 was unveiled as highly expressed in CRC cell lines compared to normal cells. AGAP2-AS1 knockdown suppressed cell proliferation, migration, invasion and the epithelial-to-mesenchymal transition process. Interestingly, AGAP2-AS1 sponges miR-4,668-3p to release SRSF1 in CRC. Furthermore, in the rescue functional assay, miR-4,668-3p down-regulation exacerbated the malignant behaviors of AGAP2-AS1-depleted CRC cells, whereas such effects were further offset by SRSF1 knockdown. CONCLUSIONS: AGAP2-AS1 facilitates cell proliferation, motility and EMT in CRC via targeting the miR-4,668-3p/SRSF1 axis. AGAP2-AS1 or SRSF1 may have potential as underlying therapeutic targets for CRC patients.

Punicalagin inhibits the viability, migration, invasion, and EMT by regulating GOLPH3 in breast cancer cells.

Breast cancer (BC) is one of the most common malignancies worldwide. Punicalagin (PN), which is a type of polyphenol, has been reported to act as a tumor suppressor. This study aimed to investigate the effects of PN on cellular process in BC and its molecular mechanism. The effects of various doses of PN on cell viability, migration, and invasion capacities of MCF-7 and MDA-MB-231 cells were detected by CCK-8, wound-healing, and Transwell assays. Golgi phosphoprotein 3 (GOLPH3) was then transfected into the cells with or without PN treatment, and GPLPH3 expression level was examined by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, and expressions of epithelial-mesenchymal transition (EMT)-related protein matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), E-Cadherin, and N-Cadherin were measured by Western blot. High dose of PN treatment (50 µM or higher) significantly inhibited viability, migration, and invasion of MCF-7 and MDA-MB-231 cells, while overexpressed GOLPH3 promoted cell viability, migration, and invasion, and partially reversed the effects of PN treatment on the BC cells. PN inhibited the expressions of GOLPH3, MMP-2, MMP-9, and N-Cadherin, and promoted E-Cadherin expression, while overexpression of GOLPH3 partly reversed above effects attributing to PN. Thus, PN suppresses cell viability and metastasis via regulating GOLPH3 in BC, which provides a possible therapeutic direction to the treatment of BC.

SMAD3 inducing the transcription of STYK1 to promote the EMT process and improve the tolerance of ovarian carcinoma cells to paclitaxel.

OBJECTIVE: To figure out the relationship between SMAD3 and serine-threonine tyrosine kinase (STYK1) in ovarian carcinoma cell's paclitaxel resistance. METHODS: The quantitative reverse transcription-polymerase chain reactpostion and Western blot analysis were used to analyze RNA and protein content of SMAD3 and STYK1, respectively. The chromatin immunoprecipitation assay was used to confirm the binding site of SMAD3 to the STYK1 promoter region. Transwell assay was used to detect cell invasion and migration, and Western Blot was used to detect the marker proteins (vimentin and E-cadherin) of epithelial-mesenchymal transition (EMT) process. MTT and apoptosis assay were used to, respectively, measure cell vitality and apoptosis. In vivo experiments, rats were subcutaneously implanted with A2780 cells to establish an animal model of ovarian cancer and the survival curve was drawn. RESULTS: Upregulating SMAD3 induced the expression of STYK1 in ovarian cancer cell lines. STYK1 is a direct transcriptional target of SMAD3. Upregulating STYK1 improved the paclitaxel resistance of ovarian carcinoma cells. Upregulating STYK1 promoted cell invasion, migration, and the EMT process, and SMAD3 had the same effect with STYK1 on cell invasion, cell migration, and the EMT process. The animal assay showed that downregulating STYK1 inhibited the EMT process and the paclitaxel resistance, further promoting the treatment of cervical cancer. CONCLUSION: SMAD3 combined with the promoter region of STYK1 to promote the transcription process of STYK1, thereby promoting the EMT process and paclitaxel resistance of ovarian cancer cells.

Iron chelator-induced up-regulation of Ndrg1 inhibits proliferation and EMT process by targeting Wnt/ß-catenin pathway in colon cancer cells.

Di-2-pyridylketone-4,4-dimethyl-3-thiosemicarbazone (Dp44mT), as the novel iron chelator, has been reported to inhibit the tumorigenesis and progression of various cancer cells. However, whether Dp44mT has anticancer effects in colon cancer cells is still unknown. Here, we investigated the antitumor action of Dp44mT in colon cancer and its underlying mechanisms, and the connections between Dp44mT and N-myc downstream-regulated genes 1(Ndrg1). We used cell viability, migration and invasion assay, flow cytometry, western blot and qRT-PCR to examine the anticancer effects of Dp44mT and Ndrg1. We found that Dp44mT suppressed cell viability, migration, invasion and induced apoptosis of colon cancer cells and over-expression of Ndrg1 also suppressed cell viability, migration, invasion and induced apoptosis of colon cancer cells. Dp44mT attenuated the TGF-ß1-induced EMT in colon cancer cells, and Dp44mT could up-regulate Ndrg1 expression level. Overexpression of Ndrg1 attenuates the TGF-ß1-induced EMT, Dp44mT and Ndrg1 suppressed EMT through activation of Wnt/ß catenin signaling pathway. In conclusion, our data demonstrated that Dp44mT/Ndrg1 have effective anticancer capability in colon cancer cells and that may represent a promising treatment strategy for human colon cancer.

MiRNA-545 negatively regulates the oncogenic activity of EMS1 in gastric cancer.

Gastric cancer (GC) is a common malignant tumor of the digestive system. In addition, GC metastasis is an extremely complicated process. In this article, high expression levels of EMS1 mRNA and protein were found to be positively correlated with an enhanced malignant potential of GC cells and a poor clinical prognosis of GC patients. Interestingly, the expression levels of EMS1 mRNA and protein in GC cells were inhibited by microRNA-545 (miR-545), which was identified by a bioinformatics analysis. The expression level of miR-545 in carcinoma tissues was significantly lower than that in para-carcinoma tissues. The proliferation and epithelial-mesenchymal transition (EMT) of GC cells were suppressed by exogenous oligonucleotides of miR-545 mimics. In addition, the expression levels of EMT-associated markers were altered with the expression of miR-545. Notably, the growth rates of tumors in nude mice were seriously restrained by an intratumoral injection of oligonucleotides of the miR-545 mimics. These results suggest a negative regulatory role of miR-545 on the oncogenic activity of EMS1. In addition, EMS1 and miR-545 may be potential biomarkers for GC diagnosis. Synthesized oligonucleotides of miR-545 mimics may be developed as important gene medicines for GC therapy in the future.

7,12-Dimethylbenz[α]anthracene increases cell proliferation and invasion through induction of Wnt/ß-catenin signaling and EMT process.

7,12-Dimethylbenz[α]anthracene (DMBA) is a hazardous component present in polluted environments. DMBA has been used as an experimental tool for in vivo tumor formation owing to its carcinogenic effects, but the detailed molecular mechanism of DMBA has not been fully established. To comprehend the carcinogenic mechanism of DMBA, we explored its effects in the breast cancer cell lines, MCF-7 and MDA-MB-231, and the cervical cancer cell line, HeLa. Cell viability assay and measurement of a proliferation marker showed that DMBA markedly increased cancer cell proliferation. Furthermore, morphological observations and wound healing assays in nontumorigenic MCF-10A cells and trans-well invasion assays in cancer cells following DMBA treatment revealed that DMBA induced cell migration and invasion. To reveal the molecular mechanism of DMBA, we investigated the effects of DMBA on the epithelial-mesenchymal transition (EMT) process and Wnt/ß-catenin signaling, a critical pathway for cell proliferation that was reported to correlate with the EMT process, by using quantitative RT-PCR (qPCR), western blot analysis, and confocal microscopy. Consequently, we found that DMBA increased cancer cell proliferation and invasion through the promotion of EMT-inducing factors and ß-catenin. Especially, it was revealed in promoter activity assay using mutated luciferase vectors on transcription factor-binding sites that TWIST1 is promoted by DMBA through induction of STAT3-mediated promoter activation. To further elucidate the detailed mechanism of DMBA, we aimed to identify the key regulator of its carcinogenic action. DMBA was shown to significantly upregulate the expression of specificity protein 1 (Sp1), a transcription factor, and the carcinogenic effects of DMBA were blocked via the suppression or interruption of Sp1 activity. In conclusion, our data suggested that DMBA induced carcinogenic effects through activation of Wnt/ß-catenin signaling and the EMT process by upregulating Sp1 activity.

N-cadherin promotes thyroid tumorigenesis through modulating major signaling pathways.

Epithelial-mesenchymal transition (EMT), a crucial step in disease progression, plays a key role in tumor metastasis. N-cadherin, a well-known EMT marker, acts as a major oncogene in diverse cancers, whereas its functions in thyroid cancer remains largely unclear. This study was designed to explore the biological roles and related molecular mechanism of N-cadherin in thyroid tumorigenesis. Quantitative RT-PCR (qRT-PCR) and immunohistochemistry assays were used to evaluate N-cadherin expression. A series of in vitro studies such as cell proliferation, colony formation, cell cycle, apoptosis, migration and invasion assays were performed to determine the effect of N-cadherin on malignant behavior of thyroid cancer cells. Our results showed that N-cadherin was significantly upregulated in papillary thyroid cancers (PTCs) as compared with non-cancerous thyroid tissues. N-cadherin knockdown markedly inhibited cell proliferation, colony formation, cell migration and invasion, and induced cell cycle arrest and apoptosis. On the other hand, ectopic expression of N-cadherin promoted thyroid cancer cell growth and invasiveness. Mechanically, our data demonstrated that tumor-promoting role of N-cadherin in thyroid cancer was closely related to the activities of the MAPK/Erk, the phosphatidylinositol-3-kinase (PI3K)/Akt and p16/Rb signaling pathways in addition to affecting the EMT process. Altogether, our findings suggest that N-cadherin promotes thyroid tumorigenesis by modulating the activities of major signaling pathways and EMT process, and may represent a potential therapeutic target for this cancer.

Stable knockdown of protein kinase CK2-alpha (CK2α) inhibits migration and invasion and induces inactivation of hedgehog signaling pathway in hepatocellular carcinoma Hep G2 cells.

Protein kinase CK2-alpha (CK2α), one isoform of the catalytic subunits of serine/threonine kinase CK2, has been indicated to participate in tumorigenesis of various malignancies, including hepatocellular carcinoma (HCC). In the present study, in order to explore the potential role of CK2α in human HCC, we employed short hairpin RNA (shRNA)-mediated RNA interference (RNAi) technology to inhibit the endogenous CK2α expression in HCC cells and established a Hep G2 cell line with stable knockdown of CK2α. Results from wound healing and transwell invasion assays indicated that stable knockdown of CK2α markedly inhibited Hep G2 cell migration and invasion as compared with those transfected with a negative control plasmid. This alteration was accompanied with expression down-regulation of matrix metalloproteinase (MMP)-2, MMP-9, Snail, Slug, Vimentin, and up-regulation of epithelial cadherin (E-cadherin). Moreover, CK2α silencing also induced inactivation of Hedgehog signaling pathway by inhibiting Gli1 and Patched homolog 1 (PTCH1) expressions in HCC cells. Collectively, these results demonstrate that knockdown of CK2α can suppress cell migration and invasion, reduces expression of MMPs, inhibits epithelial-mesenchymal transition (EMT) process and induces inactivation of Hedgehog pathway in HCC cells in vitro. Our study provides in vitro evidence to demonstrate that the pathogenesis of human HCC may be correlated with the high expression of CK2α.