Proliferation Of Esophageal Cancer Cells Research Articles

SOX2-Upregulated microRNA-30e Promotes Cell Invasion in Esophageal Cancer via USP4/SMAD4/CK2 Axis

Background: Both SOX2 and microRNA-30e (miR-30e) were reported to be involved in esophageal cancer (EC) progression. However, whether there is correlation between SOX2 and miR-30e in EC cells remains undiscovered. Herein, this study was intended to explore the correlation between SOX2 and miR-30e in EC and the effect of this correlation on EC cell invasion. Methods: Following collection of EC tissues and cells, miR-30e, SOX2, miR-30e, USP4, SMAD4, and CK2 expression was determined by RT-qPCR, western blot analysis and immunohistochemistry. Gain- and loss-of-function approaches were conducted in screened cell lines, followed by determination of cell proliferation, migration, invasion, and expression of epithelial-mesenchymal transition (EMT)-related genes. The relationship among miR-30e, USP4, SMAD4 and CK2 was assessed by dual-luciferase reporter assay, in vitro protein purification, immunoprecipitation, and ubiquitination experiments, and ChIP assay. The role of SOX2 and CK2 in EC in vivo was evaluated by tumor xenograft in nude mice. Results: EC tissues and cells contained high expression of SOX2, miR-30e, and CK2 as well as low expression of USP4 and SMAD4. miR-30e was upregulated by SOX2, and specifically targeted USP4. Additionally, USP4 induced the deubiquitination of SMAD4 to upregulate SMAD4. SMAD4 was enriched in CK2 promoter region and inhibited CK2 expression. Silencing of SMAD4 or USP4 or upregulation of SOX2, miR-30e or CK2 triggered enhancement in EC cell proliferation, migration, invasion, and EMT in vitro, and promotion of tumor growth and lung metastasis in vivo. Conclusion: Taken together, our work reveals a novel SOX2-mediated regulatory axis that plays critical roles in the invasiveness of EC cells. Funding Statement: This work was supported by Youth Foundation of the National Natural Science Foundation of China (No. 81702971), Key Scientific Research Projects of Colleges and Universities of Henan Province (No. 18A320054), Science and Technology Research Projects of Henan Province (No. 182102310116) and Youth Talent Lifting Projects of Henan Province (No. 2020HYTP050) Declaration of Interests: The authors declare that they have no competing interests. Ethics Approval Statement: The ethics committee of the First Affiliated Hospital of Zhengzhou University provided Ethical Approval for the experiments involved human beings in this study which were guided by the intentions of the Declaration of Helsinki. Ethical agreements were obtained from the donors or their relatives by written informed consent. The experiments involved animals were implemented in accordance with the principles embodied in the National Institutes of Health Guide for the Care and Use of Laboratory. Efforts were made to avoid all unnecessary distress to the animals.

Tripartite motif containing 59 (TRIM59) promotes esophageal cancer progression via promoting MST4 expression and ERK pathway

Objective: To detect the expression of tripartite motif containing 59 (TRIM59) in human esophageal cancer (EC) tissues and explore whether TRIM59 could affect the progression of EC.Methods: Quantitative PCR and immunohistochemistry assays were performed to detect the expression of TRIM59 in 40 human EC tissues and corresponding non-tumor tissues. The correlations between TRIM59 expression and clinical pathological features of patients with EC were also investigated. CCK-8, colony formation, wound closure, and transwell assays were performed to detect the effects of TRIM59 on EC cells in vitro., Immunoblotting assays were performed to detect the effects of TRIM59 on the expression of mammalian sterile-20-like kinase 4 (MST4) and ERK pathway.Results: We reported increased expression of TRIM59 in human EC tissues, and its expression was correlated with clinical features, including metastasis (p = .011*) and maximum diameter (p = .027*), in patients with EC. We further found that TRIM59 contributed to the proliferation and invasion of EC cells via regulating mammalian sterile-20-like kinase 4 (MST4) expression and ERK pathway.Conclusion: Our data confirmed the involvement of TRIM59 in EC progression and proposed that TRIM59 could serve as a promising therapeutic target for the treatment of EC.

Value of long non-coding RNA Rpph1 in esophageal cancer and its effect on cancer cell sensitivity to radiotherapy.

BACKGROUNDEsophageal cancer is a common digestive tract tumor that is generally treated with radiotherapy. Poor responses to radiotherapy in most patients generally result in local radiotherapy failure, so it is essential to find new radiosensitizers that can enhance the response of cancer cells to radiotherapy and improve the survival of esophageal cancer patients with radiation resistance. The long non-coding RNA (lncRNA) Rpph1 is highly expressed in human gastric cancer tissues, and represses breast cancer cell proliferation and tumorigenesis. However, the expression of lncRNA Rpph1 in esophageal cancer and its relationship with radio-sensitivity has not been studied.AIMTo explore the value of lncRNA Rpph1 in esophageal cancer and its effect on cancer cell sensitivity to radiotherapy.METHODSEighty-three patients with esophageal cancer admitted to Qilu Hospital of Shandong University and 90 healthy participants who received physical examinations were collected as research participants. The expression of Rpph1 was determined by qRT-PCR. siRNA-NC and siRNA-Rpph1 were transfected into esophageal cancer cell lines, and cells without transfection were designated as the blank control group. Cell survival was tested by colony formation assays, and the levels of proteins related to apoptosis and epithelial-mesenchymal transitions were determined by Western blot assays. Cell proliferation was assessed by MTT assays, cell apoptosis by flow cytometry, and cell migration by wound-healing assays. Changes in cell cycle distribution were monitored.RESULTSRpph1 was highly expressed in esophageal carcinoma, making it a promising marker for the diagnosis of esophageal cancer. Rpph1 could also be used to distinguish different short-term responses, T stages, N stages, and clinical stages of esophageal cancer patients. The results of 3-year overall survival favored patients with lower Rpph1 expression over patients with higher Rpph1 expression (P < 0.05). In vitro and in vivo experiments showed that silencing Rpph1 expression led to higher sensitivity of esophageal cancer cells to radiotherapy, stronger apoptosis in esophageal cancer cells induced by radiotherapy, higher expression of Bax and caspase-3, and lower expression of Bcl-2 (Bax, caspase-3, and Bcl-2 are apoptosis-related proteins). Additionally, silencing Rpph1 attenuated radiation-induced G2/M phase arrest, and significantly inhibited the expression of proteins involved in cell proliferation, migration, and epithelial-mesenchymal transition regulation in esophageal cancer cells.CONCLUSIONRpph1 is highly expressed in esophageal cancer. Silencing Rpph1 expression can promote cell apoptosis, inhibit cell proliferation and migration, and increase radio-sensitivity.

Dexmedetomidine suppresses the progression of esophageal cancer via miR-143-3p/epidermal growth factor receptor pathway substrate 8 axis

Esophageal cancer is one of the fatal cancers around the world. Dexmedetomidine (DEX) is widely used during anesthesia of esophageal cancer surgery. Nevertheless, the role of DEX in the progression of esophageal cancer remains barely known. The proliferation, apoptosis and metastasis of esophageal cancer cells were detected by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, flow cytometry, transwell migration and invasion assays and Western blot assay. The expression of miR-143-3p was measured by quantitative real-time PCR in esophageal cancer tissues and cells. The binding sites between miR-143-3p and epidermal growth factor receptor pathway substrate 8 (EPS8) were predicted by Starbase online software, and the combination was verified by dual-luciferase reporter assay. The murine xenograft model was established using KYSE150 cells to verify the function of DEX in vivo. DEX inhibited the proliferation and metastasis while accelerated the apoptosis of esophageal cancer cells. The abundance of miR-143-3p was lower in esophageal cancer tissues and cells than that in paring normal tissues and normal esophageal mucosal cells Het-1A. MiR-143-3p could be induced by DEX treatment in esophageal cancer cells, and miR-143-3p also suppressed the development of esophageal cancer. EPS8 was a functional target of miR-143-3p, and it played an oncogenic role in esophageal cancer. DEX inhibited the growth of tumor via miR-143-3p/EPS8 in vivo. DEX suppressed the growth and metastasis while facilitated the apoptosis of esophageal cancer cells through upregulating the abundance of miR-143-3p and reducing the level of EPS8 in vivo and in vitro, providing promising target for the treatment of esophageal cancer.

Imidazole-thiazolidinone inhibits oesophageal cancer cell proliferation via induction of apoptosis and cell cycle arrest at S phase

Purpose: To investigate the effect of imidazole-thiazolidinone on oesophageal cancer (OC) cell proliferation, and the mechanism of action involved.Methods: Human OC cells (HCE-6 and KYSE-1170) were cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10 % fetal bovine serum (FBS) and 1 % penicillin/streptomycin solution at 37 ˚C for 24 h in a humidified atmosphere of 5 % CO2 and 95 % air. After attaining 60 - 70 % confluency, the cells were treated with serum-free medium and graded concentrations of imidazolethiazolidinone (up to 160 μM) for 24 h. Normal cell culture without imidazole-thiazolidinone served as control. Cells in logarithmic growth phase were selected and used in this study. Cell proliferation and apoptosis were assessed using 3 (4,5 dimethyl thiazol 2 yl) 2,5 diphenyl 2H tetrazolium bromide (MTT), and flow cytometric assays, respectively. The levels of expression of apoptosis-related proteins were determined using Western blotting.Results: Treatment of HCE-6 and KYSE-1170 cells with imidazole-thiazolidinone for 48 h led to significant and dose-dependent reduction in their proliferation, as well as significant and dosedependent increase in the number of apoptotic cells (p &lt; 0.05). Light microscopy revealed significantreduction in HCE-6 cell count, detached cells, reduced cell size and irregular cytoplasmic vacuoles. Imidazole-thiazolidinone treatment significantly and dose-dependently decreased HCE-6 and KYSE-1170 cell migration, and arrested HCE-6 cell cycle at S phase (p &lt; 0.05). In HCE-6 cells, imidazolethiazolidinone treatment significantly and dose-dependently upregulated the expressions of cleaved caspase-3/8/9 and bax, but down-regulated bcl-2 expression significantly and dose-dependently (p &lt; 0.05). However, metalloproteinases 2 and 9 (MMP-2 and MMP-9) expressions in HCE-6 and KYSE-1170 cells were significantly and dose-dependently down-regulated by imidazole-thiazolidinone treatment (p &lt; 0.05).Conclusion: The results obtained in this study suggest that imidazole-thiazolidinone suppresses OC cell proliferation via induction of apoptosis and arrest of cell cycle at S phase.&#x0D; Keywords: Imidazole-thiazolidinone, Oesophageal cancer, Metastasis, Cell cycle arrest, Apoptosis

Regulatory mechanisms of PAR-2 on CyclinD1 in the proliferation of esophageal cancer cells

Regulatory mechanisms of PAR-2 on CyclinD1 in the proliferation of esophageal cancer cells

MicroRNA-301b and its target gene synaptosome-associated protein 91 as important modulators in esophageal cancer: functional experiments.

The aim of this study was to probe the influence of microRNA-301b (miR-301b) in esophageal cancer pathogenesis. Based on the data acquired from The Cancer Genome Atlas database, we found that miR-301b was highly expressed in esophageal cancer tissues and high expression of miR-301b was related to worse prognosis in patients with esophageal cancer. Quantitative real-time PCR revealed that the expression of miR-301b was higher in all examined esophageal cancer cell lines (ECA109, KY-SE150, TE-1, and NEC) than that in a human esophageal epithelial cell line (HEEC). Upregulation/downregulation of miR-301b facilitated/suppressed the growth, migration, and invasion of ECA109/KY-SE150 cells. Synaptosome-associated protein 91 (SNAP91) was proved to be one of the target genes of miR-301b and was negatively modulated by miR-301b. Besides, SNAP91 was lowly expressed in human esophageal cancer tissues and cell lines. Meanwhile, low expression of SNAP91 was concerned with poor prognosis in patients with esophageal cancer. Furthermore, we discovered that overexpression/depletion of SNAP91 suppressed/facilitated the proliferation of KY-SE150/ECA109 cells. MiR-301b and SNAP91 had little impact on HEEC cell proliferation and this degree of influence was negligible compared with their impacts on esophageal cancer cell proliferation. By rescue assay, we showed that overexpression of SNAP91 restrained the growth, migration, and invasion of ECA109 cells with overexpressed miR-301b while knockdown of SNAP91 showed the contrary effects on KY-SE150 cells with downregulated miR-301b. These consequences indicated that miR-301b played an important effect on esophageal cancer cells through regulating SNAP91, insinuating that miR-301b/SNAP91 might be novel potential targets for esophageal cancer therapy and prognosis.

Knockdown of MTA1 inhibits the proliferation and apoptosis of esophageal cancer cells

Objective: To investigate the effects of metastasis associated gene 1 (MTA1) expression on the proliferation and apoptosis of human esophageal cancer Eca109 cells. Methods: MTA1 siRNA was transfected into human esophageal cancer Eca109 cells, and the control group and blank group were set up. The expression of MTA1 in Eca109 cells with different treatment was detected by real-time fluorescent quantitative PCR (RT-PCR) and western blot. The proliferation of Eca109 cells was detected by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H tetrazolium bromide (MTT) assay. The cloning formation ability of Eca109 cells was detected by plate cloning assay. The apoptosis of Eca109 cells were detected by flow cytometry. The expression of proliferating cell nuclear antigen (PCNA) and apoptosis-related proteins, including cleaved caspase-3 and total caspase-3 protein in Eca109 cells were detected by western blot. Results: After 48 hours of transfection, RT-PCR result showed that the relative expression levels of MTA1 mRNA in Eca109 cells in the blank group, control group, and siRNA group were 1.00±0.10, 0.98±0.09 and 0.21±0.03, respectively. The expression of MTA1 mRNA in siRNA group was significantly inhibited (P<0.05), while no significant difference of MTA1 mRNA expression between the blank group and the control group has been found (P>0.05). Western blot results were consistent with those of RT-PCR. MTT array results showed that, compared with the blank group and transfection group, the absorbance values of Eca109 cells in siRNA group were dramatically reduced at 48, 72, and 96 h (all P<0.05). There were no significant differences of absorbance values between the blank group and the control group at 48, 72, and 96 h (all P>0.05). The results of the plate colony formation test showed that the number of colony formation in the blank group and control group were 58.64±6.86 and 60.02±7.04, respectively, significantly higher than 18.10±3.16 in siRNA group (P<0.05), while there was no significant difference between the blank group and control group (P>0.05). Flow cytometry results showed that the apoptosis rates in the blank group and control group were (2.13±0.54)% and (2.27±0.61)%, respectively, significantly lower than (32.61±5.28)% in siRNA group (P<0.05), while there was no significant difference between blank group and control group (P>0.05). Western blot results showed that the expression of PCNA protein was down-regulated while cleaved caspase-3 protein expression was upregulated in siRNA group, compared to the control group and blank group (P<0.05). Conclusions: Inhibition of MTA1 expression can inhibit the proliferation of Eca109 cells and induce apoptosis. This process may be related to the down-regulation of PCNA protein expression and activation of caspase-3 protein expression.

β-elemene suppresses the malignant behavior of esophageal cancer cells by regulating the phosphorylation of AKT

BackgroundEsophageal cancer is a digestive tract malignancy, ranking sixth among the world's deadliest tumor incidence. However, the pathogenesis of esophageal cancer is complex and the prognosis remains poor. Therefore, in-depth study of the pathogenesis and developing effective treatments are of great value for esophageal cancer. β-elemene is a natural monomeric compound derived from the Chinese herbal Curcuma wenyujin. β-elemene has been reported to have anti-tumor effects and used as an adjunct to clinical therapy for multiple cancers. This study aims to explore the effects of β-elemene on esophageal cancer and its related molecular mechanisms. MethodsTE-1 and KYSE-150 cells were used to evaluate the activity of β-elemene on esophageal cancerin vitro and in vivo. Western blot was performed for protein expression assessment. CCK8 assay and cell cycle analysis were used for proliferation testing. Flow cytometry was performed for apoptosis detection. Wound healing assay was subjected to assess the migration ability. Transwell chamber assay was applied to assess the invasion ability. HE staining, TUNEL staining and immunohistochemical staining were used to evaluate the changes in tumor tissues. ResultsWe found that β-elemene treatment suppressed proliferation, as well as induced apoptosis of esophageal cancer cells. In addition, β-elemene inhibited the migration and invasion ability of esophageal cancer cells. Furthermore, β-elemene exerted its effects against esophageal cancer by specifically regulating AKT signaling, thereby controlling the expression of PD-L1. Conclusionβ-elemene inhibits proliferation and metastasis of esophageal cancer cells by regulating the phosphorylation of AKT.

LINC00511 can promote the proliferation, migration and invasion of esophageal cancer cells through regulating microRNA-150-5p.

The aim of this study was to investigate the potential role of LINC00511 in esophageal cancer (ECa), and to explore its underlying mechanism through in vitro cell experiments. LINC00511 expression in ECa was analyzed by GEPIA database and verified by real-time fluorescence quantitative polymerase chain reaction (qPCR). The bioinformatics website was used to analyze the miRNAs that can bind to LINC00511, and the regulatory relationship between them was verified through Luciferase assay, qPCR as well as Western blotting analysis. Then, the impacts of LINC00511 and microRNA-150-5p on the proliferation or invasiveness of ECa cell lines Kyse30 and ECA109 were investigated by cell counting kit-8 (CCK-8) test and transwell experiment, respectively. Meanwhile, cell cycle and apoptosis were detected by flow cytometry. Analysis results of the GEPIA database revealed that LINC00511 had a significant high expression in ECa tissue samples in comparison with normal control ones, which is consistent with qPCR results. Meanwhile, a significant negative correlation was found between LINC00511 and microRNA-150-5p. In brief, LINC00511 was able to bind to microRNA-150-5p and inhibited its expression. Besides, overexpression of LINC00511 enhanced ECa cell proliferation and migration, accelerated cell cycle, and suppressed cell apoptosis, while transfection with microRNA-150-5p mimics caused the opposite effects. This study shows for the first time that LINC00511 modulates the progression of ECa by binding to microRNA-150-5p.

Epigenetic silencing of IGFBPL1 promotes esophageal cancer growth by activating PI3K-AKT signaling

BackgroundThere are seven insulin-like growth factor binding proteins (IGFBPs) that bind insulin-like growth factors (IGFs). IGFBP like protein1 (IGFBPL1) is a new member of this family. The function and mechanism of IGFBPL1 in esophageal cancer remains to be elucidated.MethodsEight esophageal cancer cell lines, 114 cases of esophageal dysplasia, and 501 cases of primary esophageal cancer samples were examined in this study. Methylation-specific polymerase chain reaction (MSP), immunohistochemistry, Western blot, flow cytometry, RNA interference assay, and xenograft mouse models were employed.ResultsThe expression of IGFBPL1was lost and complete methylation was found in KYSE150 and KYSE410 cells. Reduced expression and partial methylation of IGFBPL1 was found in Bic1, KYSE140, KYSE450, KYSE520, and COLO680N cells. High expression and unmethylation was detected in KYSE510 cells. Restoration of IGFBPL1 expression was found in KYSE150 and KYSE410 cells and the expression of IGFBPL1 was increased in Bic1, KYSE140, KYSE450, KYSE520, and COLO680N cells, after 5-AZA-2′-deoxycytidine treatment. IGFBPL1 was methylated in 47.3% (53/114) of esophageal dysplasia and 49.1% (246/501) of human primary esophageal squamous cell carcinoma (ESCC). Methylation of IGFBPL1 was significantly associated with TNM stage (p = 0.012), and tumor size (p = 0.009). IGFBPL1 inhibited esophageal cancer cell clonal formation and proliferation and induced cell apoptosis and G1/S phase arrest. Further study found that IGFBPL1 is involved in PI3K-AKT signaling and IGFBPL1 suppressed human ESCC xenografts growth in mice.ConclusionIGFBPL1 suppresses esophageal cancer cell growth by inhibiting PI3K-AKT signaling in vitro and in vivo. IGFBPL1 is a novel tumor suppressor in human esophageal cancer.

FOXM 1 induces Vasculogenic mimicry in esophageal cancer through \u03b2-catenin /Tcf4 signaling

ObjectiveTo investigate the role of FOXM1, β-catenin and TCF4 in esophageal cancer (EC) and their relationship to VM (Vasculogenic Mimicry).MethodsCCK-8 were performed to examine EC cell proliferation in FOXM1 silenced cells. EC cell migration and invasion were investigated through wound healing and Transwell assays, respectively. The formation of pipe like structures were assessed in 3D cultures. The expression of Foxm1, β-catenin, Tcf4 and E-cadherin were investigated through western blot, RT-qPCR and immunohistochemistry (IHC) staining. The relationship between FOXM1 expression, clinic-pathological features, and overall survival (OS) were further analyzed.ResultsA loss of FOXM1 expression correlated with the OS of ESCC patients. FOXM1 silencing led to a loss of cell growth and suppressed cell migration and invasion in ESCC cells. VM structures were identified in ESCC tissues and human EC cell lines. Mechanistically, FOXM1 was found to promote tumorigenesis through the regulation of β-catenin, Tcf4, and E-cadherin in EC cells, leading to the formation of VM structures.ConclusionsThese findings highlight FoxM1 as a novel therapeutic target in ESCC.

Effect of miR-196a inhibition on esophageal cancer growth in vitro.

Esophageal cancer has recent shown a higher incidence but lower 5-year survival rate after normal clinical treatment in China. The aim of this study was to observe whether the inhibition of miR-196a affects esophageal cancer cell growth by modulating the nuclear factor-κB target gene and to detect the possible cooperative therapeutic effects on esophageal cancer by knocking down miR-196a expression combined with the specific inhibitor of nuclear factor-κB target genes. Thus, anti-miR-196a or sotrastaurin, a protein kinase C (PKC) inhibitor, were used to alter PKC expression. We found that miR-196a knockdown or PKC inhibition by sotrastaurin changed PKC expression which then reduced esophageal cancer cell proliferation and downregulated proliferating cell nuclear antigen expression via the classical B-cell receptor-PKC nuclear factor-κB pathway but not the alternative pathway; in addition, miR-196a inhibition can increase the caspase level and induce esophageal cancer cell apoptosis. Our current results provided the evidence that miR-196a was related to the classical nuclear factor-κB pathway, and these new findings proved the potential therapeutic effect of miR-196a in targeted therapy for clinical esophageal cancer patients.

Negative regulation of CDK6 expression by microRNA-126-5p and its influence on the proliferation and invasion of esophageal cancer cells.

The present study aimed to investigate the expression of cyclin-dependent kinase 6 (CDK6) and microRNA-126-5p (miR-126-5p) in esophageal cancer tissues and cells, and their effect on esophageal cancer cell proliferation and invasion, and to explore the potential molecular mechanisms. The relative expression levels of CDK6 and miR-126-5p in esophageal cancer tissue, paracancerous tissue, and HEEC and EC109 cells were determined and compared using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). A miR-126-5p overexpression vector was constructed and a stable EC109 cell line expressing miR-126-5p was established. The EC109 cell line was transfected with a CDK6 small interfering RNA sequence. The rate of cell proliferation was determined using the WST-8 method, and cell invasion was determined using a Transwell assay. In addition, the relative expression levels of genes were determined using RT-qPCR; the relative expression levels of proteins were determined by western blot analysis; the binding sites of CDK6 and miR-126-5p were analyzed using TargetScan software; and the interaction of CDK6 and miR-126-5p was verified using dual-fluorescence reporter gene expression. Esophageal tissues and EC109 cells expressed higher levels of CDK6 but significantly lower levels of miR-126-5p compared with adjacent tissues and HEEC cells, and their correlation coefficient between esophageal tissues and matched adjacent tissues was -7.526. The overexpression of miR-126-5p and CDK6 knockdown in the EC109 cell line inhibited cell proliferation and invasion compared with the control and NC (negative control) groups. miR-126-5p overexpression reduced the relative expression level of CDK6, and CDK6 knockdown by siRNA increased the expression of miR-126-5p. miR-126-5p regulated CDK6 expression by binding to the 3'-untranslated region of its mRNA. Overexpression miR-126-5p inhibited the proliferation and migration of esophageal cancer cells by targeting CDK6 and negatively regulating its expression. These findings contribute to the understanding of the underlying molecular mechanism of esophageal cancer.

Silencing lncRNA AGAP2-AS1 Upregulates miR-195-5p to Repress Migration and Invasion of EC Cells via the Decrease of FOSL1 Expression

The interaction of long non-coding RNAs (lncRNAs), microRNAs (miRNAs), and mRNAs has been implicated in various types of cancers, including esophageal cancer (EC). The current study aimed to investigate the role of AGAP2-AS1/miR-195-5p/Fos-like antigen-1 (FOSL1) in EC progression. The expression of AGAP2-AS1, miR-195-5p, and FOSL1 in tumor tissues isolated from EC patients and EC cell lines was determined by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), the results of which illustrated that AGAP2-AS1 and FOSL1 were increased while miR-195-5p was reduced in EC. Next, the ectopic expression, knockdown, and reporter assay experiments were all employed to elucidate the mechanism of AGAP2-AS1/miR-195-5p/FOSL1 in the processes of EC cell proliferation, cell cycle, apoptosis, invasion, and migration as well as tumor growth. Knockdown of AGAP2-AS1 or overexpression of miR-195-5p reduced EC cell proliferation, migration, and invasion, blocked cell cycle entry, and elevated apoptosis. FOSL1 was found to be specifically targeted by miR-195-5p. AGAP2-AS1 was observed to upregulate FOSL1 by binding to miR-195-5p. Silencing of AGAP2-AS1 was observed to restrain the development of EC both in vitro and in vivo through upregulating miR-195-5p and downregulating FOSL1. Taken together, AGAP2-AS1 knockdown exercises suppressive effects on the development of EC through miR-195-5p-dependent downregulation of FOSL1. Therefore, targeting AGAP2-AS1 could be a future direction to develop a novel molecule-targeted therapeutic strategy for EC.

Circular RNA hsa_circ_0006168 contributes to cell proliferation, migration and invasion in esophageal cancer by regulating miR-384/RBBP7 axis via activation of S6K/S6 pathway.

Esophageal cancer (EC) ranks as the sixth leading cause of cancer-related mortality worldwide. Circular RNAs (circRNAs) are involved in the pathogenesis of different cancers. However, the regulatory mechanism of circ_0006168 in EC progression is still unclear. The expression of circ_0006168, microRNA (miR)-384, and retinoblastoma binding protein 7 (RBBP7) in tumors and cells was measured by quantitative Real Time-Polymerase Chain Reaction (qRT-PCR). The stability of circ_0006168 was analyzed after RNase R treatment. 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide (MTT) assay was conducted to evaluate cell viability. Transwell assay was applied to determine cell migration and invasion. Glucose consumption and lactate production were detected using glucose detection and lactic acid detection kits. The interaction between miR-384 and circ_0006168 or RBBP7 was certified by Dual-Luciferase reporter system. Protein expression of pyruvate kinase (PK), RBBP7, S6 ribosomal protein kinase (S6K), phosphorylated S6K (p-S6K), S6, phosphorylated S6 (p-S6) was analyzed by Western blot. Circ_0006168 and RBBP7 were over-expressed while miR-384 was low-expressed in EC tumors and cells. The repression of circ_0006168 attenuated cell proliferation, migration, invasion, and glycolysis in EC. Of note, circ_0006168 functioned as a sponge while RBBP7 acted as a target of miR-384 in EC. Rescue experiment revealed that miR-384 inhibitor abrogated circ_0006168 silencing-induced repression on cell proliferation, migration, and invasion in EC. Meanwhile, upregulation of RBBP7 restored the inhibition of miR-384 on EC cell progression. Moreover, circ_0006168 was able to improve RBBP7 level by interacting with miR-384. Also, circ_0006168 could activate S6K/S6 pathway by regulating RBBP7 expression. Abundance of circ_0006168 contributes to cell proliferation, migration, invasion, and glycolysis in EC by competitively sponging miR-384 to facilitate RBBP7 expression, representing prospective targets for EC therapy.

Effects of HOXA1 Gene Antisense Oligonucleotides on Growth of Esophageal Cancer Cells

To investigate the effect of antisense oligodeoxynucleotides (ASODN) of Homeobox A1 gene ( HOXA1) on proliferation, apoptosis, invasion and migration of esophageal carcinoma cells. The expression of HOXA1 protein in normal esophageal epithelial cells Het-1A and esophageal cancer TE-1, EC9706 and Eca109 cells was detected by Western blot. Screening of highly expressed of HOXA1 protein esophageal squamous cell carcinoma cells for follow-up experiments. HOXA1 antisense oligonucleotide (ASODN) chains, sense oligodeoxynucleotides (SODN) chain, and nonsense oligodeoxy nucleotides (N-ODN) chain were designed. The screened esophageal squamous cell carcinoma cells with high expression were divided into HOXA1 ASODN group (5, 10, 15 μmol/L HOXA1 ASODN transfected Eca109 cells), control group (conventional culture medium, no cell transfection), SODN group (cells transfected with 15 μmol/L of SODN) and N-ODN group (cells transfected with 15 μmol/L N-ODN). Cell viability, apoptosis rate and invasion and migration ability were detected by methylthiazolyldiphenyl-tetrazolium bromide (MTT) method, flow cytometry, transwell chamber respectively; The expression of HOXA1, phosphorylation serine/threonine kinase (p-AKT), proliferating cell nuclear antigen (PCNA), matrix metalloproteinase 2 (MMP-2) and B-cell lymphoma2 (Bcl-2) associated X protein (Bax) protein was detected by Western blot. Compared with normal esophageal epithelial cells Het-1A, the expression of HOXA1 protein in human esophageal squamous cell carcinoma cells TE-1, EC9706 and Eca109 was significantly higher ( P<0.05). The expression of HOXA1 protein was the highest in Eca109 cells, therefore, Eca109 cells were selected for follow-up experiments. The expression of HOXA1 protein in Eca109 cells transfected with HOXA1 ASODN was significantly decreased ( P<0.05). After transfection of Eca109 cells with HOXA1 ASODN, the viability of Eca109 cells decreased with the increase of concentration and time, the difference was significant compared with the control, SODN and N-ODN groups ( P<0.05). 15 μmol/L HOXA1 ASODN significantly inhibited cell viability. After 15 μmol/L HOXA1 ASODN was transfected into Eca109 cells, the invasion and migration abilities of cells were significantly decreased, the apoptosis rate was increased, the expressions of p-AKT, PCNA and MMP-2 were significantly decreased, and the expression of Bax was significantly increased ( P<0.05). Antisense oligodeoxynucleotides of HOXA1 gene can inhibit the proliferation, invasion and migration of esophageal cancer cells, and induce apoptosis. The mechanism is related to the inhibition of PI3K/AKT signaling pathway.

THAP11 mediates the proliferation and apoptosis of esophageal cancer cells via inhibiting ubiquitination of p53

To investigate the effects of thanatos-associated protein 11 (THAP11) on the proliferation and apoptosis of esophageal cancer cell and the underlying mechanism. Methods: Expression of THAP11 in human esophageal epithelial cells (Het-1A) and esophageal cancer cells (Eca109, TE-1, Ec 9706) were detected by Western blotting. Esophageal cancer TE-1 cells were divided into 3 groups: a normal control (NC) group, a negative control (LV-NC) group and a THAP11 (LV-THAP11) group. Then the cell proliferation were detected by MTT assay, cell apoptosis were detected by flow cytometry, caspase-3 and caspase-9 levels were detected by caspases kits. Ubiquitination of p53 was determined in esophageal cancer TE-1 cells. Results: Expression of THAP11 was reduced in esophageal cancer cells compared with human esophageal epithelial cells (P<0.05). After transfection with LV-THAP11 in TE-1 cells, cell viability was reduced (P<0.05), while apoptosis rate and caspase-3 and caspase-9 levels were increased (P<0.05), indicating that THAP11 inhibited growth of esophageal cancer cells. In addition, the THAP11 increased the levels of p53 (P<0.05) and inhibited the ubiquitination of p53 regulated by MDM2. Conclusion: THAP11 may inhibit the proliferation of esophageal cancer cells by inhibiting ubiquitination of p53.

Comprehensive multi-omics analysis identified core molecular processes in esophageal cancer and revealed GNGT2 as a potential prognostic marker.

BACKGROUNDEsophageal cancer is one of the most poorly diagnosed and fatal cancers in the world. Although a series of studies on esophageal cancer have been reported, the molecular pathogenesis of the disease remains elusive.AIMTo investigate comprehensively the molecular process of esophageal cancer.METHODSDifferential expression analysis was performed to identify differentially expressed genes (DEGs) in different stages of esophageal cancer from The Cancer Genome Atlas data. Exacting gene interaction modules were generated, and hub genes in the module interaction network were found. Further, through survival analysis, methylation analysis, pivot analysis, and enrichment analysis, some important molecules and related functions/pathways were identified to elucidate potential mechanisms in esophageal cancer.RESULTSA total of 7457 DEGs and 14 gene interaction modules were identified. These module genes were significantly involved in the positive regulation of protein transport, gastric acid secretion, insulin-like growth factor receptor binding, and other biological processes as well as p53 signaling pathway, epidermal growth factor signaling pathway, and epidermal growth factor receptor signaling pathway. Transcription factors (including hypoxia inducible factor 1A) and non-coding RNAs (including colorectal differentially expressed and hsa-miR-330-3p) that significantly regulate dysfunction modules were identified. Survival analysis showed that G protein subunit gamma transducin 2 (GNGT2) was closely related to survival of esophageal cancer. DEGs with strong methylation regulation ability were identified, including SST and SH3GL2. Furthermore, the expression of GNGT2 was evaluated by quantitative real time polymerase chain reaction, and the results showed that GNGT2 expression was significantly upregulated in esophageal cancer patient samples and cell lines. Moreover, cell counting kit-8 assay revealed that GNGT2 could promote the proliferation of esophageal cancer cell lines.CONCLUSIONThis study not only revealed the potential regulatory factors involved in the development of esophageal cancer but also deepens our understanding of its underlying mechanism.

Long non-coding RNA DLEU2 promotes the progression of esophageal cancer through miR-30e-5p/E2F7 axis

BackgroundEmerging evidences have proven the important roles of lncRNAs in tumorigenesis and cancer biology. However, the function of lncRNA DLEU2 in the progression of esophageal cancer (EC) has not been elaborated. In the present study, we aimed to investigate the effects of lncRNA DLEU2 on the progression of EC and the underlying mechanism. MethodsIn this study, lncRNA DLEU2 was silenced by siRNA interference in EC cell lines Eca-109 and KYSE-150, and its expression was up-regulated in TE-1 cells by transfection with pcDNA3.1-DLEU2, and its biological functions were examined. Then, bioinformatics analysis and dual-luciferase reporter assay were used to identify the binding miRNA of lncRNA DLEU2 and the target gene of miRNA. In addition, loss-of-function assays were performed to detect the biological functions of the target gene. At last, the rescue assays were used to investigate the relationship among lncRNA DLEU2, miRNA and target gene. ResultsWith the help of GEPIA analysis, we observed that lncRNA DLEU2 was up-regulated in EC tissues and associated with poor prognosis. Loss-of-function assay showed that silencing lncRNA DLEU2 inhibited the proliferation, migration and invasion of EC cells, and induced apoptosis by regulating the Bcl-2/Bax axis and Caspase cascade. Overexpression of lncRNA DLEU2 increased the proliferation, migration and invasion abilities of TE-1 cells, as well as decreased cell apoptosis. Bioinformatics analysis and dual-luciferase reporter assay verified that miR-30e-5p could directly bind with lncRNA DLEU2, and E2F7 was a direct target for miR-30e-5p in EC cells. Moreover, our data revealed that silencing E2F7 decreased the proliferation, migration and invasion abilities of EC cells, and induced apoptosis. Furthermore, the rescue assays demonstrated that the effects of lncRNA DLEU2 on the proliferation, migration and invasion of EC cells were reversed by miR-30e-5p inhibitor or up-regulation of E2F7. ConclusionsOur findings revealed the pro-oncogenic role of lncRNA DLEU2 and E2F7 in the progression of EC, suggesting that lncRNA DLEU2 exerts ceRNA functions in EC through regulating miR-30e-5p/E2F7 axis.