Invasion Of Esophageal Carcinoma Cells Research Articles

Baitouweng decoction suppresses growth of esophageal carcinoma cells through miR-495-3p/BUB1/STAT3 axis.

Esophageal carcinoma (EC) is one of the most prevalent cancers in human populations worldwide. Baitouweng decoction is one of the most important Chinese medicine formulas, with the potential to treat cancer. To investigate the role and mechanism of Baitouweng decoction on EC cells. Differentially expressed genes (DEGs) in EC tissues and normal tissues were screened by the cDNA microarray technique and by bioinformatics methods. The target genes of microRNAs were predicted based on the TargetScan database and verified by dual luciferase gene reporter assay. We used Baitouweng decoction to intervene EC cells, and detected the activity of EC9706 and KYSE150 cells by the MTT method. Cell cycle and apoptosis were measured by flow cytometry. The expression of BUB1 mRNA and miR-495-3p was measured by qRT-PCR. The protein levels of BUB1, STAT3, p-STAT3, CCNB1, CDK1, Bax, Caspase3, and Caspase9 were measured by Western blot analysis. The migration and invasion abilities of the cells were measured by wound-healing assay and Transwell invasion assay, respectively. DEGs identified are involved in biological processes, signaling pathways, and network construction, which are mainly related to mitosis. BUB1 was the key hub gene, and it is also a target gene of miR-495-3p. Baitouweng decoction could upregulate miR-495-3p and inhibit BUB1 expression. In vitro experiments showed that Baitouweng decoction significantly inhibited the migration and invasion of EC cells and induced apoptosis and G2/M phase arrest. After treatment with Baitouweng decoction, the expression of Bax, Caspase 3, and Caspase 9 in EC cells increased significantly, while the expression of BUB1, CCNB1, and CDK1 decreased significantly. Moreover, the STAT3 signaling pathway may play an important role in this process. Baitouweng decoction has a significant inhibitory effect on EC cell growth. BUB1 is a potential therapeutic target for EC. Further analysis showed that Baitouweng decoction may inhibit the growth of EC cells by upregulating miR-495-3p targeting the BUB1-mediated STAT3 signal pathway.

Abstract 4720: Effects of removing heat and nourishing yin prescription on malignant phenotype and PI3K/AKT signaling pathway of human esophageal carcinoma cells

Abstract Introduction: Esophageal carcinoma (EC) ranks eighth in the incidence of malignant tumors and the sixth mortality worldwide. This study aims to screen out the Chinese herbs with strong inhibitory effects on the proliferation of four kinds of EC cells EC9706, EC-1, TE-1 and Eca109 from 366 Chinese herbs produced in Henan Province, and study the effect of composed Removing Heat and Nourshing Yin Prescription(RHNYP) on the proliferation, migration, invasion and PI3K/AKT signaling pathway of four EC cells. Methods: MTT and orthogonal design was applied to screen out the best effective Chinese herbs and determine the optimal dosage ratio of prescription on EC9706, EC-1, TE-1, Eca109 cells from 366 Chinese herbs alcohol extracts, the effects of this prescription on the proliferation, migration, invasion, clone formation, cell cycle and apoptosis of four EC cells were investigated by MTT, RTCA, soft agarose colony forming test, flow cytometry and western blot analysis. Then exploring the effects of RHNYP on EC related signaling pathways and key genes expression by western blot and RT-PCR assay. Results: Rhizoma Fagopyri Dibotryis(Jinqiaomai), Rhizoma Anemarrhenae(Zhimu), Asparagus Cochinchinensis(Tiandong), and Carpesium Abrolanoides Linne(Heshi) were screened out and composed Removing Heat and Nourshing Yin Prescription(RHNYP) according to their efficacy and orthogonal design. RHNYP could obviously inhibit the proliferation of EC9706, TE-1, EC-1, ECa109 cells with dose-effect and time-dependent manner. IC50 values of Jinqiaomai, Zhimu, Tiandong, Heshi in RHNYP for EC9706 cell were 27.639μg/ml, 40.015μg/ml, 9.044μg/ml, 11.208μg/ml respectively; for EC-1 cell were 24.575μg/ml, 56.991μg/ml, 9.361μg/ml, 13.584μg/ml respectively; for TE-1 cell were 23.534μg/ml, 25.862μg/ml, 6.788μg/ml, 8.911μg/ml respectively; for ECa109 cell were 33.990μg/ml, 33.604μg/ml, 19.873μg/ml, 15.227μg/ml respectively. RHNYP could effectively inhibit the proliferation, migration, invasion, cloning ability, cell cycle and reduce apoptosis of four EC cells. PI3K/AKT/mTOR pathway had closely relationship with the occurrence and development of EC and RHNYP could effectively regulate the proteins expression on the pathway. CDCK3, KIF4A and RAD51AP1 genes were negatively correlated with the overall survival of EC patients and RHNYP could significantly inhibit their expression in four EC cells. Conclusion: RHNYP can effectively inhibit EC cell proliferation, migration, invasion and cell cycle, induce apoptosis, which may be related to the regulation of PI3K/AKT/mTOR pathway. Citation Format: Fuchun Si, Wenyu Zhao, Gao Si, Xuejie Song. Effects of removing heat and nourishing yin prescription on malignant phenotype and PI3K/AKT signaling pathway of human esophageal carcinoma cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4720.

Characterization of the Roles of Suppressor of Cytokine Signaling-3 in Esophageal Carcinoma.

This study was aimed to analyze the diagnostic, therapeutic, and prognostic value of the suppressor of cytokine signaling 3 (SOCS3) in pancancer, especially in esophageal carcinoma (ESCA), and investigate the role of SOCS3 in the tumorigenesis and progression of ESCA. We used a variety of bioinformatics methods to explore the expression of SOCS3 in 33 kinds of cancers and evaluate its potential role in the pathogenesis, prognosis, immune microenvironment, immune evasion, and therapeutic response of cancers. The results indicated that SOCS3 was upregulated in 10 cancers, downregulated in 12 cancers, and upregulated in ESCA. Mutation and amplification were the main causes of abnormal expression of SOCS3 in pancancer. In ESCA, expression of SOCS3 was negatively correlated with methylation. The analysis showed that ESCA patients with low SOCS3 levels had better overall survival. Furthermore, the SOCS3 level was positively related to the ESTIMATE score, immune score, stromal score, and negatively related to tumor purity. In ESCA, a significant association was found between SOCS3 and several immune checkpoint genes. In addition, SOCS3 was associated with sensitivity to 59 drugs. Next, the role of SOCS3 in ESCA was investigated in ECA109, EC9706 cells, and in xenografted mouse model. SOCS3 was confirmed to be upregulated in ESCA cells. Knockdown of SOCS3 decreased the proliferation, migration, and invasion of ESCA cells while increasing apoptosis. Meanwhile, downregulation of SOCS3 activated the nuclear factor kappa-B signaling pathway and inhibited ESCA tumorigenesis in vivo. In conclusion, high SOCS3 expression is closely related to the occurrence and progression of ESCA and can be used as a therapeutic target and prognostic biomarker for ESCA.

Tumor cell-derived exosomal lncRNA LOC441178 inhibits the tumorigenesis of esophageal carcinoma through suppressing macrophage M2 polarization.

Esophageal carcinoma (EC) is a highly malignant type of tumor. In a previous study, the authors found that long non-coding RNA (lncRNA) LOC441178 inhibited the tumorigenesis of EC. Moreover, exosomes derived from tumor cells containing lncRNAs were found to play a key role in the tumor environment; however, whether exosomes can affect the tumor microenvironment by carrying LOC441178 remains unclear. Thus, the present study aimed to clarify this. In order to assess the effects of exosomal LOC441178 in EC, cell invasion and migration were examined using the Transwell assay. Exosomes were identified using transmission electron microscopy, western blot analysis and nanoparticle tracking analysis. Furthermore, macrophage surface makers (CD206 and CD86) were analyzed using flow cytometry. Moreover, a subcutaneous xenograft mouse model was constructed to assess the role of TE-9 cells-derived exosomal LOC441178 in EC. The results revealed that LOC441178 overexpression notably suppressed the metastasis of EC cells. In addition, exosomes were successfully isolated from EC cells, and LOC441178 level was upregulated in exosomes derived from LOC441178-overexpressed EC cells. Exosomal LOC441178 also suppressed macrophage M2 polarization, and the polarized macrophages decreased EC cell invasion. Exosomes containing LOC441178 notably inhibited the growth of EC in mice. On the whole, the present study demonstrated that the delivery of LOC441178 by EC cell-secreted exosomes inhibited the tumorigenesis of EC by suppressing the polarization of M2 macrophages. These findings may provide a new theoretical basis for discovering new strategies against EC.

LncRNA miR205HG hinders HNRNPA0 translation: anti-oncogenic effects in esophageal carcinoma.

Esophageal carcinoma (ESCA) affects 4 450 000 people and causes approximately 400 000 deaths annually worldwide, making it the sixth most lethal and eighth most common cancer. Patients with ESCA are often diagnosed at the later stages in which cancer cell metastasis is the main factor contributing to the low 5‐year survival rate (< 20%) of this disease. Long noncoding RNAs (lncRNAs) are a group of regulatory RNAs with a length of > 200 nucleotides but which fail to encode proteins. In this study, by using real‐time quantitative PCR, we found that the expression of the miR205 host gene (miR205HG; a lncRNA) was downregulated in ESCA tumors when compared with normal esophageal tissues or adjacent normal tissues of tumors. Furthermore, we demonstrated that miR205HG modulates the expression of extracellular matrix‐related genes in ESCA cells. In the transwell assay, downregulation of miR205HG contributes to migration and invasion of ESCA cells. In relation to the mechanism, our data show that miR205HG interacts with heterogeneous nuclear ribonucleoprotein A0 (HNRNPA0) mRNA and then hamper its translation by interacting with lin‐28 homolog A (LIN28A). Altogether, we highlight that the miR205HG‐HNRNPA0 axis is implicated in the migration and invasion of ESCA cells and that these members of this pathway may serve as therapeutic targets to inhibit metastasis of ESCA.

MiR-145-5p Inhibits the Proliferation, Migration, and Invasion of Esophageal Carcinoma Cells by Targeting ABRACL.

Objective The study is aimed at investigating the regulatory relationship between miR-145-5p and ABRACL, and has tried at clarifying the mechanisms underlying the proliferation, migration, and invasion of esophageal carcinoma (EC) cells. Methods Gene expression data related to EC were accessed from TCGA database, and the “edgeR” package was used to screen differentially expressed genes. TargetScan, miRDB, and miRTarBase databases were used to predict potential targets for the target miRNA miR-145-5p. qRT-PCR and Western blot were performed to assess the expression of miR-145-5p and ABRACL in EC cells. Dual-luciferase reporter assay was performed to validate the targeting relationship between miR-145-5p and ABRACL. Functional experiments including CCK-8 assay, Transwell migration, and invasion assays were used to detect the proliferation, migration, and invasion of EC cells. Results The expression of miR-145-5p was significantly decreased in EC, while ABRACL was remarkably increased. In addition, there was a negative correlation identified between miR-145-5p and ABRACL mRNA. Overexpressing miR-145-5p was able to suppress cell proliferation, migration, and invasion, whereas silencing miR-145-5p posed an opposite effect. In the meantime, ABRACL was identified as a direct target of miR-145-5p by dual-luciferase reporter assay. Furthermore, miR-145-5p could inhibit the expression of ABRACL, in turn inhibiting the proliferation, migration, and invasion of EC cells. Conclusion miR-145-5p functions on the proliferation, migration, and invasion of EC cells via targeting ABRACL, and it may be a novel therapeutic target in EC treatment.

Long non-coding RNA DDX11-AS1 promotes esophageal carcinoma cell proliferation and migration through regulating the miR-514b-3p/RBX1 axis

ABSTRACT Esophageal carcinoma (ESCA) is one of the most aggressive malignancies with extremely high morbidity and mortality. At present, limited advancement in ESCA treatment has achieved. Therefore, it is urgent to explore the pathogenesis and progression mechanism of ESCA to provide the basis for the formulation of novel therapeutic strategies. Previous studies have found that long non-coding RNA (lncRNA) DDX11-AS1 expression enhances the paclitaxel resistance of ESCA cells. However, the mechanisms underlying the drug resistance conferred by lncRNA DDX11-AS1 in ESCA remains to be elucidated. Our research aims to clarify the role and mechanism of lncRNA DDX11-AS1 in regulating the progression of ESCA. We found that the expression of lncRNA DDX11-AS1 in ESCA tissues and cell lines was significantly upregulated. Subsequently, silencing lncRNA DDX11-AS1 significantly inhibited the proliferation, migration and invasion of ESCA cells, and induced the level of cell apoptosis. In terms of mechanism, our data showed that miR-514b-3p/RING box protein 1 (RBX1) axis played a crucial role in the oncogenic function of lncRNA DDX11-AS1. LncRNA DDX11-AS1 expression impaired the inhibitory function of miR-514b-3p on RBX1 through sponging effect. Taken together, our data support the notion that lncRNA DDX11-AS1 promotes the progression of ESCA through miR-514b-3p/RBX1 axis. Our research uncovers the novel regulatory role of lncRNA DDX11-AS1 in ESCA and lays a theoretical basis for developing novel treatment strategy of ESCA.

PRR11 and SKA2 promote the proliferation, migration and invasion of esophageal carcinoma cells.

Proline-rich protein 11 (PRR11) together with its upstream adjacent gene, spindle and kinetochore associated 2 (SKA2), represent a classic, head-to-head gene pair. The role of the PRR11 and SKA2 gene pair has been described in various types of cancer, including breast cancer, non-small cell lung cancer, hepatocellular carcinoma and ovarian carcinoma. However, its role in esophageal carcinoma (ESCC) remains unclear. The mRNA expression levels of PRR11 and SKA2 were examined in ESCC surgical specimens. In addition, the role of PRR11 and SKA2 in the proliferation and migratory and invasive capacities of EC9706 and EC109 cell lines was examined. The results from the present study demonstrated that PRR11 and SKA2 expression levels were upregulated in ESCC tissues compared with adjacent normal tissues. Furthermore, PRRl1 and SKA2 knockdown significantly inhibited the proliferation and migratory and invasive capacities of ESCC cells. Conversely, PRRl1 and SKA2 overexpression significantly promoted the proliferation and migratory and invasive capacities of ESCC cell lines via activation of the AKT signaling pathway and certain markers of epithelial-mesenchymal transition, including Snail and N-cadherin. The results from the present study suggested that the PRR11 and SKA2 gene pair may represent a potential target in the diagnosis and treatment of ESCC.

Expression of lncRNA AK058003 in esophageal carcinoma and analysis of its intervention effect.

The aim of this study was to investigate the expression of long non-coding ribonucleic acid (lncRNA) AK058003 in esophageal carcinoma (EC) tissues, and to analyze its intervention effect. The expression of lncRNA AK058003 in EC tissues and para-carcinoma tissues from 130 EC patients was detected via quantitative Polymerase Chain Reaction (qPCR). EC cell lines were selected for exogenous interference in lncRNA AK058003. Subsequently, the expression of lncRNA AK058003 in normal esophageal epithelial cell line (Het-1A) and EC cell lines (EC109, EC9706, KYSE-150, KYSE-30, and TE-1) was detected by qPCR. EC9706 cell lines with the highest expression of lncRNA AK058003 were selected and transfected with lncRNA AK058003 siRNA and lncRNA AK058003 control, respectively. After transfection, the expression of lncRNA AK058003 was determined using PCR. The changes in cell growth and proliferation were analyzed via cell growth curve and cell cycle assay. Meanwhile, the changes in cell migration and invasion were analyzed through wound healing assay. Protein expressions of matrix metalloproteinase-1 (MMP1) and MMP2 were determined by Western blot. Clinical data were collected from EC patients, and the association between lncRNA AK058003 expression and tumor-node-metastasis (TNM) stage was finally analyzed. LncRNA AK058003 was highly expressed EC tissues compared with para-carcinoma tissues (p<0.01). Compared with Het-1A cells, the expression of lncRNA AK058003 was significantly higher in EC109, EC9706, KYSE-150, KYSE-30, and TE-1 cells, with highest level in EC9706 cells (p<0.05). The expression of lncRNA AK058003 remarkably declined in lncRNA AK058003 siRNA group compared with lncRNA AK058003 control group (p<0.001). Compared with lncRNA AK058003 control group, the proliferation of EC cells was significantly weakened in lncRNA AK058003 siRNA group, with the greatest difference at 3 d. Flow cytometry results revealed that cell cycle was arrested in G0/G1 phase in lncRNA AK058003 siRNA group. Wound healing assay indicated that the intercellular distance became large, and cell migration ability was evidently enhanced in lncRNA AK058003 siRNA group with time (p<0.05). Besides, the protein expressions of MMP1 and MMP2 were remarkably lower in lncRNA AK058003 siRNA group than those in lncRNA AK058003 control group. This indicated remarkably declined invasion and metastasis ability. In addition, the postoperative prognosis was significantly worse in patients with higher expression of lncRNA AK058003 (p<0.05). All these findings suggested that lncRNA AK058003 could serve as a biomarker for EC prognosis. LncRNA AK058003 is highly expressed in EC patients, which promotes proliferation, migration, invasion, and metastasis of EC cells. In addition, the postoperative prognosis of EC patients with high expression of lncRNA AK058003 is relatively poor.

RETRACTED: Silencing lncRNAs PVT1 Upregulates miR-145 and Confers Inhibitory Effects on Viability, Invasion, and Migration in EC

Long non-coding RNA (lncRNA) plasmacytoma variant translocation 1 (PVT1) is correlated to various malignant tumors. Consequently, we explored effects of lncRNA PVT1 on esophageal carcinoma (EC) targeting microRNA-145 (miR-145). EC tissues, adjacent normal tissues, and EC-related cell lines were collected and cultured. Expression of lncRNA PVT1, miR-145, fascin-1 (FSCN1), and related genes with intervening expression of PVT1 and miR-145 was determined. Bioinformatic website, dual-luciferase reporter assay, and RNA immunoprecipitation (RIP) were carried to verify target relationship among lncRNA PVT1, FSCN1, and miR-145. Scratch test, Transwell assay, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and flow cytometry were performed for detection of migration, invasion, viability, and apoptosis of transfected cells, respectively. Finally, tumor formation in nude mice was measured. After database analysis, lncRNA PVT1, miR-145, and FSCN1 were selected for study. lncRNA PVT1 and FSCN1 can bind to miR-145. After overexpressing miR-145 or inhibiting lncRNA PVT1, EC cell viability, migration, and invasion were inhibited, while volume and weight of tumor formation in nude mice decreased. Expression of lncRNA PVT1, FSCN1, Bcl-2, CD147, VEGFR2, and MTA1 decreased and expression of miR-145 and Bax increased. Silencing lncRNA PVT1 can upregulate miR-145, which is a tumor suppressor in EC via knockdown of FSCN1. Thus, we might provide a potential theoretical basis for EC treatment.

Retraction Note: RNAi targeting CXCR4 inhibits proliferation and invasion of esophageal carcinoma cells

The Editor-in-Chief has retracted this article [1] because Fig. 3 shows overlap with Fig. 6 in [2], Fig. 2b in [3] and Fig. 6a in [4]. An investigation by Zhengzhou University has confirmed that these figures overlap. The data reported in this article are therefore unreliable.

MiRNA‑30a‑3p inhibits metastasis and enhances radiosensitivity in esophageal carcinoma by targeting insulin‑like growth factor 1 receptor.

It has been demonstrated that microRNAs (miRNAs) serve important roles in various biological processes, such as tumorigenesis. In the present study, the role of miR-30a-3p in the pathogenesis of esophageal carcinoma (EC) was investigated. Reverse transcription-quantitative polymerase chain reaction was performed to determine the levels of miR-30a-3p expression in EC tissues and cell lines. Then, the effects of miR-30a-3p on the migration, invasion and radiosensitivity of EC cells were investigated using scratch-wound, Transwell and radiosensitivity assays, respectively. A dual-luciferase reporter assay was performed to determine potential interactions between miR-30a-3p and the 3′-untranslated region (3′-UTR) of insulin-like growth factor 1 receptor (IGF-1R). The results demonstrated that the levels of miR-30a-3p expression in EC tissues and cell lines were significantly decreased compared with those in paired healthy tissues and a human esophageal epithelial cell line. Upregulation of miR-30a-3p expression significantly suppressed migration, invasion and epithelial-mesenchymal transition (EMT), and enhanced radiosensitivity in EC cells. Analysis of luciferase activity demonstrated that miR-30a-3p interacted with the 3′-UTR of IGF-1R, and knockdown of IGF-1R induced similar effects on the migration, invasion, EMT and radiosensitivity of EC cells. The results indicated that miR-30a-3p suppressed metastasis and enhanced the radiosensitivity of EC cells via downregulation IGF-1R, suggesting that miR-30a-3p may be a potential therapeutic target in the treatment of EC.

The 19q13 KRAB Zinc-finger protein ZFP82 suppresses the growth and invasion of esophageal carcinoma cells through inhibiting NF-κB transcription and inducing apoptosis.

To study the epigenetic alternations and biological functions of ZFP82 in esophageal squamous cell carcinoma. Analysis of ZFP82 expression was carried out by quantitative real-time PCR. Cell function was tested by MTS cell proliferation assay, transwell assay and flow cytometry. Gene mechanisms were studied by reverse transcription-PCR (RT-PCR), quantitative real-time PCR, luciferase reporter assay and Western blot. ZFP82 promoter methylation was downregulated in esophageal squamous cell carcinoma. ZFP82 ectopic expression suppressed cell function and regulated NF-κB phosphorylation and genes involved in cell cycle arrest and apoptosis. Moreover, ZFP82 methylation was correlated with age, tumor stage and outcome. ZFP82 is a tumor suppressor and is disrupted by promoter CpG methylation during esophageal carcinogenesis.

The targeted regulation of Gli1 by miR-361 to inhibit epithelia-mesenchymal transition and invasion of esophageal carcinoma cells.

Epithelia-mesenchymal transition (EMT) is critical for invasion and metastasis of esophageal carcinoma. Gli1, a transcriptional factor in Hedgehog pathway, is correlated with EMT, invasion and metastasis of tumors. However, its role in esophageal cancer is still unknown. Bioinformatics analysis revealed relationship between microRNA (miR)-361 and 3'-UTR of Gli1 gene. This study thus investigated the role of miR-361 and Gli1 in invasion and metastasis of esophageal cancer. Both tumor and adjacent tissues were collected from 58 esophageal cancer patients to test the expressions of miR-361 and Gli1, the relationship of which was confirmed by dual-luciferase reporter gene assay. Cultured esophageal cancer cells EC9706 were transfected with mimic NC, miR-361 mimic, si-NC, si-Gli1, miR-361 mimics+si-Glil, pQC or pQC-FU-Gli1. Transwell and colony formation assays were performed for cell invasion and attachment-independent growth. Expressions of Gli1, Snail, E-cadherin and N-cadherin proteins were revealed by Western blotting. The expression of Gli1 was significantly elevated in esophageal cancer tissues, along with lower miR-361 expression which was correlated with TNM stage. MiR-361 inhibited the expression of Gli1 via targeting on 3'-UTR of Gli1 gene. The transfection of miR-361 mimics and/or si-Gli1 significantly suppressed the growth of malignant cells. The over-expression of miR-361 and/or silencing of Gli1 decreased intracellular expression of Gli1, Snail and N-cadherin, and increased E-cadherin expression to suppress EMT and invasion of tumor cells while the opposite effects were obtained by over-expression of Gli1. Abnormal elevation of Gli1 and decrease of miR-361 were found in esophageal cancer tissues. MiR-361 weakened invasion of cancer cells and impeded EMT process via the inhibition of Gli1.

Metformin inhibits the migration and invasion of esophageal squamous cell carcinoma cells by downregulating the protein kinase B signaling pathway.

Previous studies have suggested that metformin, a biguanide family member widely used as an oral antidiabetic drug, may inhibit proliferation and induce apoptosis in certain types of cancer cell. However, the molecular mechanisms underlying metformin-associated anticancer effects, and in particular antimetastatic effects, remain to be fully understood. The present study assessed the efficacy of metformin in inhibiting the migration and invasion of the esophageal carcinoma cell line EC109, and evaluated the effect of metformin on the protein kinase B (AKT) signaling pathway. EC109 cells were treated with 0, 5, 10 or 20 mM metformin during the logarithmic growth phase. A Transwell assay and western blot analysis revealed that metformin inhibited the migration and invasion of EC109 cells, nuclear factor-κB activation, matrix metallopeptidase 9 and N-cadherin expression in a phosphorylated-AKT dependent manner. These results suggested that metformin inhibits the migration and invasion of human esophageal carcinoma cells by suppressing AKT phosphorylation and regulating the expression of migration- and invasion-associated genes.

Downregulation of MACC1 inhibits the viability, invasion and migration and induces apoptosis in esophageal carcinoma cells through the phosphatase and tensin homolog/phosphoinositide 3-kinase/protein kinase B signaling pathway.

As an oncogene, MACC1 serves an important function in cancer progression and metastasis. However, the effect of MACC1 in esophageal carcinoma (EC) remains to be fully understood. The present study assessed the association between MACC1 expression and the progression of EC cells. A small interfering (si)RNA was delivered into EC cells to downregulate MACC1 expression. The MTT assay demonstrated that EC cell viability was reduced by siRNA-MACC1. Decreasing MACC1 expression increased the apoptotic rate of EC cells compared with control cells. Transwell and Matrigel assays demonstrated that EC cell migration and invasion, respectively, were downregulated by siRNA-MACC1. Furthermore, knocking down MACC1 suppressed the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) signaling pathway by upregulating the expression of phosphatase and tensin homolog (PTEN), a tumor suppressor. The results of the present study revealed that MACC1 expression affected cellular functions of the EC cells through the PTEN/PI3K/Akt signaling pathway. Therefore, MACC1 may potentially serve as a novel biomarker and therapeutic target for EC.

Inhibition of hypoxia-inducible factor-1α by PX-478 as a potential targeted therapy in ESCC.

e14083 Background: Hypoxia is a unique microenvironment in solid tumors, including ESCC. We aim to investigate the interaction between hypoxia-inducible factor-1α (HIF-1α), COX-2 and programmed cell death ligand-1 (PD-L1) and uncover the role of HIF-1α inhibitor PX-478 as a potential targeted therapy in ESCC. Methods: Immunohistochemical staining was performed to investigate the levels of HIF-1α, COX-2 and PD-L1 from 133 pT3N0M0 ESCC patients after radical resection. The prognostic value of the expression of HIF-1α, COX-2 and PD-L1 and the correlation with clinicopathologic features was evaluated. Knockdown assay, CCK-8 assay, Western blot, real-time polymerase chain reaction (RT-PCR), flow cytometry and Transwell migration assays were used in cells experiment. Results: HIF-1α and PD-L1 are independent prognostic factors in pT3N0M0 ESCC. Further data showed that HIF-1α plays an important role in regulation of COX-2 and PD-L1 expression. Our in vitro studies demonstrated that HIF-1α inhibitor, PX-478, induced G2 phase arrest, increased apoptosis, and inhibited migration and invasion of esophageal carcinoma cells, and thus significantly inhibit ESCC cells proliferation. Conclusions: Our results provide new insight into the potential role of HIF-1α inhibitors, PX-478 and open up the possibility of PX-478 for targeted therapy of ESCC.

Decreased expression of CD63 tetraspanin protein predicts elevated malignant potential in human esophageal cancer.

The tetraspanin CD63 has been described to have critical roles in multiple biological processes, including tumorigenesis and metastasis in several types of cancer. However, its role in esophageal carcinoma (EC) has not been reported. In the current study, immunohistochemistry was used to investigate CD63 expression in 106 esophageal cancer samples, 49 adjacent esophagus tissues and 17 normal esophagus mucosa tissues. The results revealed that the overexpression of CD63 was observed in esophageal cancer samples and negatively correlated with tumor stage and lymph node metastasis. To further evaluate the role of CD63 in esophageal carcinoma, the invasiveness of EC cells was analyzed using matrigel invasion assays and wound healing assays in vitro. Furthermore, it was found that CD63 knockdown increased the invasiveness of TE-1 cells through the upregulation of matrix metalloproteinase (MMP) expression via promoting epithelial-mesenchymal transition. The current data therefore suggested that low levels of CD63 expression may be involved in the tumor progression of esophageal carcinoma.

Influence of lymphatic endothelial cells on proliferation and invasiveness of esophageal carcinoma cells in vitro and lymphangiogenesis in vivo.

The aim of the study was to investigate the interaction between esophageal carcinoma cells with different differentiation degree and esophageal carcinoma-related lymphatic endothelial cells. Different lymphatic endothelial cell conditioned mediums were used to cultivate well-differentiated esophageal carcinoma EC9706 cells and poorly differentiated esophageal carcinoma KYSE150 cells, and immunocytochemistry and Western blot analyses were applied to detect the expression of MMP-9 protein and TIMP-2 protein in each group; in situ hybridization and RT-PCR methods were used to detect the expression of MMP-9 and TIMP-2 mRNA in each group; CCK-8 method was used to detect cell proliferation in each group; and transwell method was utilized to detect cell invasiveness in each group. Through constructing the transplanted tumor model of esophageal carcinoma of nude mice, the D2-40 and LYVE-1 immunohistochemical staining was performed on transplanted tumors and surrounding tissues, lymphatic microvessels were marked, and lymphatic microvessel density (LMVD) was measured. The expression of MMP-9 protein and mRNA in experimental group was significantly higher than that in control groups (P<0.05); TIMP-2 protein and mRNA expression in experimental group was significantly lower than that in control groups (P<0.05); cell proliferation ability and invasiveness ability in experimental group were significantly higher than those in control groups (P<0.05); LMVD-marked D2-40 and LMVD-marked LYVE-1 of transplanted tumor tissue in the experimental group were significantly higher than those in control groups (P<0.05). The esophageal squamous carcinoma-related lymphatic microvessel could promote the proliferation and invasive ability of esophageal squamous carcinoma cells in vitro. It had different effects on esophageal carcinoma cells with different differentiation degree and had more obvious influence on poorly differentiated esophageal carcinoma cells, which may be related to the up-regulated MMP-2 expression and down-regulated TIMP-2 expression of esophageal carcinoma cells. The esophageal squamous carcinoma-related lymphatic microvessel endothelial cells could promote the growth of esophageal carcinoma-transplanted tumor of nude mice and lymphangiogenesis.

HIF-1α induces VE-cadherin expression and modulates vasculogenic mimicry in esophageal carcinoma cells.

To investigate whether hypoxia inducible factor (HIF)-1α modulates vasculogenic mimicry (VM) by upregulating VE-cadherin expression in esophageal squamous cell carcinoma (ESCC). Esophageal squamous cancer cell lines Eca109 and TE13 were transfected with plasmids harboring small interfering RNAs targeting HIF-1α or VE-cadherin. The proliferation and invasion of esophageal carcinoma cells were detected by MTT and Transwell migration assays. The formation of tubular networks of cells was analyzed by 3D culture in vitro. BALB/c nude mice were used to observe xenograft tumor formation. The relationship between the expression of HIF-1α and VE-cadherin, ephrinA2 (EphA2) and laminin5γ2 (LN5γ2) was measured by Western blot and real-time polymerase chain reaction. Knockdown of HIF-1α inhibited cell proliferation (32.3% ± 6.1% for Eca109 cells and 38.6% ± 6.8% for TE13 cells, P < 0.05). Both Eca109 and TE13 cells formed typical tubular networks. The number of tubular networks markedly decreased when HIF-1α or VE-cadherin was knocked down. Expression of VE-cadherin, EphA2 and LN5γ2 was dramatically inhibited, but the expression of matrix metalloproteinase 2 had no obvious change in HIF-1α-silenced cells. Knockdown of VE-cadherin significantly decreased expression of both EphA2 and LN5γ2 (P < 0.05), while HIF-1α expression was unchanged. The time for xenograft tumor formation was 6 ± 1.2 d for Eca109 cells and Eca109 cells transfected with HIF-1α Neo control short hairpin RNA (shRNA) vector, and 8.4 ± 2.1 d for Eca109 cells transfected with an shRNA against HIF-1α. Knockdown of HIF-1α inhibited vasculogenic mimicry (VM) and tumorigenicity in vivo. HIF-1α may modulate VM in ESCC by regulating VE-cadherin expression, which affects VM formation through EphA2 and LN5γ2.