Journal: Translational Oncology

Article Title: Anticancer Activity of a Broccoli Derivative, Sulforaphane, in Barrett Adenocarcinoma: Potential Use in Chemoprevention and as Adjuvant in Chemotherapy 1

doi:

Figure Lengend Snippet: Effect of SFN on BEAC cell survival. BEAC cells were cultured in the medium containing no SFN or various concentrations of SFN. Cells were harvested at different time points as indicated and proliferative potential was assessed by trypan blue exclusion and/or proliferation assay, based on the production of a yellow product (formazan) after reduction of a highly water-soluble tetrazolium salt by dehydrogenases in viable cells. The growth curves show the mean of three independent experiments, with SEM. (A) Barrett adenocarcinoma (FLO-1) cells treated with various concentrations of SFN. (B) BEAC (OE33) cells treated with various concentrations of SFN. (C) Photomicrograph of BEAC (FLO-1 and OE33) cells treated with 3 µM SFN for 72 hours. (D) Photomicrograph of normal diploid fibroblasts and primary normal esophageal epithelial cells (ScienCell Research Laboratories) treated with 3 µM SFN for 72 hours. (E) FLO-1 cells were treated with SFN for 48 hours, detached floating cells from the medium and the attached cells (by trypsinization) were collected separately and evaluated for number and viability using trypan blue exclusion. The number of cells detached after treatment with various concentrations of SFN is expressed as percent of untreated FLO-1 cells. “Total” represents the total number of detached cells whereas “Dead” reflects the fraction of dead cells in detached cell population. (F) Panel (I): FLO-1 cells were incubated with various concentrations of SFN for 48 hours, and the expression of caspase 8 was detected by Western blot analysis, using anti-caspase 8 mouse monoclonal antibody (Cell Signaling, Danvers, MA). Panel (II): Bar graph showing caspase 8 expression relative to β-actin.

Article Snippet: Normal primary human esophageal epithelial cells (HEEC) were purchased from ScienCell Research Laboratories (Carlsbad, CA) and have been described previously [ 30 ].

Techniques: Cell Culture, Proliferation Assay, Incubation, Expressing, Western Blot

Journal: Cell Transplantation

Article Title: LINC00461 Promoted Endometrial Carcinoma Growth and Migration by Targeting MicroRNA-219-5p/Cyclooxygenase-2 Signaling Axis

doi: 10.1177/0963689721989616

Figure Lengend Snippet: LINC00461 was elevated in EC tissues and cell models. (A) Relative LINC00461 levels in EC tissues (tumor, n = 45) and control tissues (normal, n = 45), determined using qRT-PCR. (B) LINC00461 levels in EC tissues (tumor, n = 45) and control tissues (normal, n = 45), determined using ISH. (C) Kaplan–Meier curves of overall survival of 45 EC patients, stratified by LINC00461 expressions. (D) Relative LINC00461 levels in various EC cell lines (KLE, Ishikawa, HEC1-A, HEC-1-B, and AN3-CA) and hEECs, determined by qRT-PCR (mean ± SEM, ** P < 0.01). EC: endometrial carcinoma; hEEC: human endometrial epithelial cell; ISH: in situ hybridization; qRT-PCR: quantitative real-time polymerase chain reaction; SEM: standard error of the mean.

Article Snippet: Human endometrial epithelial cells (hEEC, catalog No. PCS-100-011) and EC cell lines including KLE (catalog No. CRL-1622), HEC1-A (catalog No. HTB-112), HEC-1-B (catalog No. HTB-113), and AN3-CA, (catalog No. HTB-111) were purchased from American Type Culture Collection (ATCC, Rockville, MD, USA).

Techniques: Control, Quantitative RT-PCR, In Situ Hybridization, Real-time Polymerase Chain Reaction

Journal: BMC Cancer

Article Title: The role of DNMT1/hsa-miR-124-3p/BCAT1 pathway in regulating growth and invasion of esophageal squamous cell carcinoma

doi: 10.1186/s12885-019-5815-x

Figure Lengend Snippet: The expression and hypermethylation of hsa-miR-124-3p in ESCC tissues and cells. The mRNA expression of hsa-miR-124-3p in normal tissues and ESCC tissues ( a ) and its expression in HEEC, KYSE-150 and Eca109 cells ( b ) were determined by qRT-PCR analysis. The methylation status of hsa-miR-124-3p in normal tissues and ESCC tissues ( c ) and its status in HEEC, KYSE-150 and Eca109 cells ( d ) were determined by bisulfite Sanger sequencing and methylation specific PCR respectively. hsa-miR-124-3p expression in tumor tissues was shown in the patents with different TNM stages ( e ) and different differentiation ( f ). Experiments were performed in triplicate and each value represents mean ± SD. *** P < 0.001

Article Snippet: Human normal esophageal cell line HEEC (Het-1A, ATCC® CRL-2692TM) was obtained from the America Type Culture Collection in May 2017, and two human esophageal carcinoma cell lines, KYSE-150 (TCHu236) and Eca-109 (TCHu69), were obtained from Shanghai Institute of Biological Sciences, Chinese Academy of Sciences in May 2017.

Techniques: Expressing, Quantitative RT-PCR, Methylation, Sequencing

Journal: BMC Cancer

Article Title: The role of DNMT1/hsa-miR-124-3p/BCAT1 pathway in regulating growth and invasion of esophageal squamous cell carcinoma

doi: 10.1186/s12885-019-5815-x

Figure Lengend Snippet: The expression of BCAT1 in ESCC tissues and cells. The mRNA expression of BCAT1 in normal tissues and ESCC tissues ( a ) and its expression in HEEC, KYSE-150 and Eca109 cells ( b ) were determined by qRT-PCR analysis. c The protein expression of BCAT1 in normal tissues and ESCC tissues was determined by immunohistochemistry staining analysis. d The Kaplan-Meier survival curve of ESCC patients with different levels of BCAT1 expression. e The correlation between hsa-miR-124-3p and BCAT1 gene expression in ESCC tissues was assessed by qRT-PCR ( n = 50; p < 0.0001). The experiments were performed in triplicate and each value represents mean ± SD. * p < 0.05, *** p < 0.001

Article Snippet: Human normal esophageal cell line HEEC (Het-1A, ATCC® CRL-2692TM) was obtained from the America Type Culture Collection in May 2017, and two human esophageal carcinoma cell lines, KYSE-150 (TCHu236) and Eca-109 (TCHu69), were obtained from Shanghai Institute of Biological Sciences, Chinese Academy of Sciences in May 2017.

Techniques: Expressing, Quantitative RT-PCR, Immunohistochemistry, Staining, Gene Expression

Journal: BMC Cancer

Article Title: The role of DNMT1/hsa-miR-124-3p/BCAT1 pathway in regulating growth and invasion of esophageal squamous cell carcinoma

doi: 10.1186/s12885-019-5815-x

Figure Lengend Snippet: The regulatory role of DNMT1 inhibitor on expression of hsa-miR-124-3p and BCAT1 in ESCC cells. a - b The mRNA expression of DNMT1 in normal tissues and ESCC tissues ( a ) and its expression in HEEC, KYSE-150 and Eca109 cells ( b ) were determined by qRT-PCR analysis. c The methylation status of hsa-miR-124-3p in KYSE-150 and Eca109 cells treated with DNMT1 inhibitor 5 μM 5-Azacitidine (5-Aza) or its control (PBS), was detected by methylation specific PCR. d The mRNA expression of DNMT1, hsa-miR-124-3p and BCAT1 in KYSE-150 and Eca109 cells treated with 5-Aza or PBS were determined by qRT-PCR analysis. e The protein expression of DNMT1 and BCAT1 were detected by western blotting assay. The target protein expression relative to GAPDH expression was displayed on the right. The experiments were performed in triplicate and each value represents mean ± SD. *** P < 0.001

Article Snippet: Human normal esophageal cell line HEEC (Het-1A, ATCC® CRL-2692TM) was obtained from the America Type Culture Collection in May 2017, and two human esophageal carcinoma cell lines, KYSE-150 (TCHu236) and Eca-109 (TCHu69), were obtained from Shanghai Institute of Biological Sciences, Chinese Academy of Sciences in May 2017.

Techniques: Expressing, Quantitative RT-PCR, Methylation, Control, Western Blot

Journal: Journal of Inflammation Research

Article Title: TNF/NF-κB Signaling Pathway is Involved in Mechanisms of Tongluo Decoction on Rat Models with Sequelae of Pelvic Inflammatory Disease

doi: 10.2147/JIR.S518734

Figure Lengend Snippet: TNF, IKBKB, and NFKB1 mediated the effect of Tongluo Decoction (TLD) on the necroptosis of human endometrial epithelial cells (HEECs). ( A and B ) ( C ) The expression of TNF, IKBKB, and NFKB1 mRNAs in HEECs by quantitative real-time PCR, after TLD serum treatment, with or without overexpression of TNF, IKBKB and NFKB1. ( D ) The necroptosis of HEECs characterized by number of PI + cell ratio using flow cytometry with Annexin V/PI staining. ( E – H ) The protein levels of RIPK1, RIPK3, MLKL, and cas8 in HEECs by Western blot, after TLD serum treatment, with or without overexpression of TNF, IKBKB and NFKB1. ** P < 0.01; *** P < 0.001.

Article Snippet: Human endometrial epithelial cells (HEECs) were obtained from Procell (Wuhan, China), and THP-1 cells were obtained from the American Type Culture Collection (Manassas, USA).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Over Expression, Flow Cytometry, Staining, Western Blot

Journal: Molecular Therapy. Nucleic Acids

Article Title: SOX2-Upregulated microRNA-30e Promotes the Progression of Esophageal Cancer via Regulation of the USP4/SMAD4/CK2 Axis

doi: 10.1016/j.omtn.2020.10.027

Figure Lengend Snippet: SOX2 Is Highly Expressed in EC Tissues and Cells, which Is Negatively Correlated with EC Patients’ Prognosis (A) mRNA expression of SOX2 in EC tissues and adjacent normal tissues detected by qRT-PCR (n = 30). (B) Quantitation of SOX2 protein in EC tissues and adjacent normal tissues by immunohistochemical assay. (C) Kaplan-Meier analysis showing the correlation of SOX2 expression with OS and DFS of EC patients. (D) SOX2 expression determined by qRT-PCR and western blot analysis in EC cell lines (Eca109, EC9706, KYSE150, and TE-1) and normal esophageal epithelial cells (HEEC). ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. Paired t test was used for comparison between data of EC tissues and adjacent normal tissues, and one-way ANOVA was used for data comparison among multiple groups. Data are shown as mean ± standard deviation of three technical replicates.

Article Snippet: Human esophageal epithelial cell line HEEC and human esophageal cancer cell lines (Eca109, EC9706, KYSE150, and TE-1) were attained from the Cell Bank, China Center for Type Culture Collection (Wuhan, Hubei, P.R.

Techniques: Expressing, Quantitative RT-PCR, Quantitation Assay, Immunohistochemical staining, Western Blot, Comparison, Standard Deviation

Journal: Molecular Therapy. Nucleic Acids

Article Title: SOX2-Upregulated microRNA-30e Promotes the Progression of Esophageal Cancer via Regulation of the USP4/SMAD4/CK2 Axis

doi: 10.1016/j.omtn.2020.10.027

Figure Lengend Snippet: SOX2 Promotes Proliferation, Migration, and Invasion, as well as EMT of Eca109 Cells via miR-30e Upregulation In Vitro (A) Expression of miR-30e in EC tissues and adjacent normal tissues (n = 30) determined by qRT-PCR. (B) Expression of miR-30e in two EC cell lines (Eca109 and TE-1) and normal esophageal epithelial cells detected by qRT-PCR. (C) Pearson correlation analysis of SOX2 expression with miR-30e expression. (D) Expression of SOX2 and miR-30e in Eca109 cells transfected with oe-SOX2 detected by qRT-PCR. Eca109 cells were transfected with oe-SOX2 or in combination with miR-30e inhibitor. (E) Expression of SOX2 and miR-30e in Eca109 cells measured by qRT-PCR. (F) Proliferation of Eca109 cells detected by EdU assay (×200). (G) Quantitation of migration of Eca109 cells detected by scratch test. (H) Quantitation of invasion of Eca109 cells detected by Transwell assay. (I) Quantitative western blot analysis of EMT-related proteins (E-cadherin, N-cadherin, and Vimentin) in Eca109 cells. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. Paired t test was used for comparison between data of EC tissues and adjacent normal tissues, while unpaired data between the other two groups were compared by unpaired t test. Comparisons among multiple groups were performed using one-way ANOVA. Data are shown as mean ± standard deviation of three technical replicates.

Article Snippet: Human esophageal epithelial cell line HEEC and human esophageal cancer cell lines (Eca109, EC9706, KYSE150, and TE-1) were attained from the Cell Bank, China Center for Type Culture Collection (Wuhan, Hubei, P.R.

Techniques: Migration, In Vitro, Expressing, Quantitative RT-PCR, Transfection, EdU Assay, Quantitation Assay, Transwell Assay, Western Blot, Comparison, Standard Deviation

Journal: Molecular Therapy. Nucleic Acids

Article Title: SOX2-Upregulated microRNA-30e Promotes the Progression of Esophageal Cancer via Regulation of the USP4/SMAD4/CK2 Axis

doi: 10.1016/j.omtn.2020.10.027

Figure Lengend Snippet: miR-30e Promotes Cell Proliferation, Migration, Invasion, and EMT in Eca109 Cells by Targeting USP4 In Vitro (A) Putative miR-30e binding sites in the 3′ UTR of USP4 mRNA in the Starbase database. (B) A heatmap of DEGs in the EC gene expression profiling GSE89102, in which each row represents a DEG, and each column represents a sample. (C) Expression of USP4 analyzed in the EC gene expression profiling GSE89102. (D) Expression of USP4 in EC analyzed by the GEPIA website, where red represents cancer tissues and black represents normal control tissues. (E) mRNA expression of USP4 examined by qRT-PCR in EC tissues and adjacent normal tissues (n = 30). (F) Quantitation of USP4 protein in EC tissues and adjacent normal tissues by immunohistochemistry analysis. (G) Expression of USP4 examined by qRT-PCR in Eca109, TE-1, and HEEC cells. (H) Quantitative western blot analysis of USP4 protein in Eca109, TE-1, and HEEC cells. (I) Pearson correlation analysis of USP4 expression with miR-30e expression. (J) miR-30e binding to USP4 confirmed by dual luciferase reporter assay in 293T cells. (K) Silencing efficiency of USP4 expression in Eca109 cells measured by qRT-PCR. (L) Silencing efficiency of USP4 expression in Eca109 cells measured by quantitative western blot analysis. Eca109 cells were transfected with mimic-NC + vector, miR-30e mimic + vector, and miR-30e mimic + USP4. (M) Expression of USP4 and miR-30e determined by qRT-PCR in Eca109 cells. (N) Quantitation of EdU-positive Eca109 cells. (O) Quantitation of migration of Eca109 cells detected by scratch test. (P) Quantitation of invasion of Eca109 cells detected by Transwell assay. (Q) Quantitative western blot analysis of EMT-related proteins (E-cadherin, N-cadherin, and Vimentin) in Eca109 cells. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. Paired t test was used for comparison between data of EC tissues and adjacent normal tissues, while unpaired data between the other two groups were compared by unpaired t test. Comparisons among multiple groups were performed using one-way ANOVA. Data are shown as mean ± standard deviation of three technical replicates.

Article Snippet: Human esophageal epithelial cell line HEEC and human esophageal cancer cell lines (Eca109, EC9706, KYSE150, and TE-1) were attained from the Cell Bank, China Center for Type Culture Collection (Wuhan, Hubei, P.R.

Techniques: Migration, In Vitro, Binding Assay, Gene Expression, Expressing, Control, Quantitative RT-PCR, Quantitation Assay, Immunohistochemistry, Western Blot, Luciferase, Reporter Assay, Transfection, Plasmid Preparation, Transwell Assay, Comparison, Standard Deviation