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: Thoracic Cancer

Article Title: circ‐ TTC17 Promotes Esophagus Squamous Cell Carcinoma Cell Growth, Metastasis, and Inhibits Autophagy‐Mediated Radiosensitivity Through miR ‐145‐5p/ SIRT1 Axis

doi: 10.1111/1759-7714.15494

Figure Lengend Snippet: The expression of circ‐TTC17 and miR‐145‐5p in ESCC. (A–B) QRT‐PCR results indicated that circ‐TTC17 was highly expressed in ESCC tissues (T) and cell lines compared with that in adjacent normal tissues (N) and HEEC cells, respectively. * p < 0.05.

Article Snippet: ESCC cell lines (TE1, KYSE150, KYSE180, and KYSE30) and human esophageal epithelial cells (HEEC) were purchased from Procell (Wuhan, China) and grown in RPMI‐1640 medium containing 10% FBS and 1% penicillin/streptomycin. circ‐TTC17 siRNA, lentiviral shRNA, miR‐145‐5p mimic, anti‐miR‐145‐5p, and negative controls were transfected into KYSE180 and KYSE30 cells by Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA).

Techniques: Expressing, Quantitative RT-PCR

Journal: Thoracic Cancer

Article Title: circ‐ TTC17 Promotes Esophagus Squamous Cell Carcinoma Cell Growth, Metastasis, and Inhibits Autophagy‐Mediated Radiosensitivity Through miR ‐145‐5p/ SIRT1 Axis

doi: 10.1111/1759-7714.15494

Figure Lengend Snippet: miR‐145‐5p targeted SIRT1 in ESCC. (A) The binding sequences between miR‐145‐5p and SIRT1 3′UTR were presented. The interaction was verified using dual‐luciferase reporter assay (B–C) and RIP assay (D–E). (F) QRT‐PCR was used to detect SIRT1 mRNA expression in ESCC tissues (T) and adjacent normal tissues (N). (G) The correlation between SIRT1 and miR‐145‐5p expression was assessed by Pearson correlation coefficient analysis. (H) The protein expression of SIRT1 in T and N was determined by WB analysis. The mRNA and protein levels of SIRT1 in ESCC cell lines and HEEC cells were measured using qRT‐PCR (I) and WB analysis (J), respectively. (K) The transfection efficiency of miR‐145‐5p mimic and inhibitor was evaluated using qRT‐PCR. (L) SIRT1 protein expression in KYSE180 and KYSE30 cells was determined by WB analysis. * p < 0.05.

Article Snippet: ESCC cell lines (TE1, KYSE150, KYSE180, and KYSE30) and human esophageal epithelial cells (HEEC) were purchased from Procell (Wuhan, China) and grown in RPMI‐1640 medium containing 10% FBS and 1% penicillin/streptomycin. circ‐TTC17 siRNA, lentiviral shRNA, miR‐145‐5p mimic, anti‐miR‐145‐5p, and negative controls were transfected into KYSE180 and KYSE30 cells by Lipofectamine 3000 (Invitrogen, Carlsbad, CA, USA).

Techniques: Binding Assay, Luciferase, Reporter Assay, Quantitative RT-PCR, Expressing, Transfection

Journal: Frontiers in Endocrinology

Article Title: Betulinic Acid Inhibits Endometriosis Through Suppression of Estrogen Receptor β Signaling Pathway

doi: 10.3389/fendo.2020.604648

Figure Lengend Snippet: Expression of ERβ and its target genes is significantly increased in endometriotic cells compared to endometrial cells. Human Endometrial Epithelial Cells HEEC, immortalized Human Endometriotic Epithelial Cell Line 12Z, human primary endometrial epithelial cells (EM), and primary endometriotic epithelial cells (EMT) were used for gene expression analysis. (A) mRNA levels for ERβ/ERα and its target genes by qPCR, n = 4. (B) Quantitation of protein levels, n = 5. (C) Representative western blotting pictures for (B) . * P < 0.05, vs CTL group. Data were expressed as mean ± SEM.

Article Snippet: Human Endometrial Epithelial Cells HEEC (#ABC-TC4601) and Immortalized Human Endometriotic Epithelial Cell Line 12Z (#ABI-TC278D) were obtained from ACCEGEN Biotechnology.

Techniques: Expressing, Gene Expression, Quantitation Assay, Western Blot

Journal: Frontiers in Endocrinology

Article Title: Betulinic Acid Inhibits Endometriosis Through Suppression of Estrogen Receptor β Signaling Pathway

doi: 10.3389/fendo.2020.604648

Figure Lengend Snippet: Betulinic acid suppresses the ERβ signaling pathway by modulation of epigenetic changes on the ERβ promoter and subsequent expression. (A–C) The conditional immortalized HEEC or 12Z cells were either treated by 20 µM betulinic acid (BA) for 24 h or infected by empty control (CTL), ERβ expression (↑ERβ), or ERβ knockdown (shERβ) lentivirus. The cells were then harvested for gene expression analysis. (A) The mRNA levels qPCR, n = 4. (B) Quantitation of protein levels, n = 5. (C) Representative western blotting pictures for (B) . * P < 0.05, vs HEEC/CTL group; ¶ P < 0.05, vs 12Z/CTL. (D) 12Z cells were treated by different concentrations of betulinic acid for 24 h, and the cells were harvested for analysis of ERα and ERβ mRNA by qPCR, n = 4. (E) 12Z cells were transiently transfected by either ERβ full length (pERβ-2000) or deletion reporter plasmids. After 24 h, the cells were treated by either control (CTL) or 20 µM BA for 24 h, and BA-induced relative ERβ reporter activities (% control) were calculated, n = 5. * P < 0.05, vs pERβ-2000 group. (F) Treated cells were harvested to measure epigenetic changes on the ERβ promoter by ChIP analysis, n = 4. * P < 0.05, vs HEEC/CTL group. Data were expressed as mean ± SEM.

Article Snippet: Human Endometrial Epithelial Cells HEEC (#ABC-TC4601) and Immortalized Human Endometriotic Epithelial Cell Line 12Z (#ABI-TC278D) were obtained from ACCEGEN Biotechnology.

Techniques: Expressing, Infection, Control, Knockdown, Gene Expression, Quantitation Assay, Western Blot, Transfection

Journal: Frontiers in Endocrinology

Article Title: Betulinic Acid Inhibits Endometriosis Through Suppression of Estrogen Receptor β Signaling Pathway

doi: 10.3389/fendo.2020.604648

Figure Lengend Snippet: Betulinic acid mimics ERβ knockdown-induced oxidative stress in endometriotic cells. The conditional immortalized HEEC or 12Z cells were either treated by 20 µM betulinic acid (BA) for 24 h or infected by either empty control (CTL) or ERβ knockdown (shERβ) lentivirus; the cells were then harvested for analysis of oxidative stress. (A) ROS formation, n = 5. (B) Quantitation of 3-nitrotyrosine formation, n = 5. (C) 8-OHdG formation, n = 5. (D) Quantitation of γH2AX formation. (E) Representative γH2AX western blotting band for (D) , n = 5. (F) SOD2 activity, n = 5. (G) Quantitation of 8-oxo-dG formation, n = 5. (H) Representative pictures of 8-oxo-dG staining for oxidative stress (green) and DAPI staining for nuclei (blue), n = 4. * P < 0.05, vs HEEC/CTL group; ¶ P < 0.05, vs 12Z/CTL group. Data were expressed as mean ± SEM.

Article Snippet: Human Endometrial Epithelial Cells HEEC (#ABC-TC4601) and Immortalized Human Endometriotic Epithelial Cell Line 12Z (#ABI-TC278D) were obtained from ACCEGEN Biotechnology.

Techniques: Knockdown, Infection, Control, Quantitation Assay, Western Blot, Activity Assay, Staining

Journal: Frontiers in Endocrinology

Article Title: Betulinic Acid Inhibits Endometriosis Through Suppression of Estrogen Receptor β Signaling Pathway

doi: 10.3389/fendo.2020.604648

Figure Lengend Snippet: Betulinic acid mimics ERβ knockdown-induced mitochondrial dysfunction and apoptosis in endometriotic cells. The conditional immortalized HEEC or 12Z cells were infected by lentivirus with either empty control (CTL), or ERβ knockdown (shERβ), or treated by 20 µM betulinic acid (BA) for 24 h; the cells were then harvested for analysis of mitochondrial function. (A) Mitochondrial DNA copies, n = 4; (B) Intracellular ATP levels, n = 5. (C) Caspase-3 activity, n = 5. (D) Δψm by TMRE fluorescence, n = 5. (E) Apoptosis rate by TUNEL assay, n = 5. (F) Representative pictures for (E) . * P < 0.05, vs HEEC/CTL group; ¶ P < 0.05, vs 12Z/CTL group. Data were expressed as mean ± SEM.

Article Snippet: Human Endometrial Epithelial Cells HEEC (#ABC-TC4601) and Immortalized Human Endometriotic Epithelial Cell Line 12Z (#ABI-TC278D) were obtained from ACCEGEN Biotechnology.

Techniques: Knockdown, Infection, Control, Activity Assay, Fluorescence, TUNEL Assay

Journal: Frontiers in Endocrinology

Article Title: Betulinic Acid Inhibits Endometriosis Through Suppression of Estrogen Receptor β Signaling Pathway

doi: 10.3389/fendo.2020.604648

Figure Lengend Snippet: ERβ knockdown reduces endometriosis-mediated elevated proinflammatory cytokine secretion, and betulinic acid treatment mimics this effect. The conditionally immortalized HEEC or 12Z cells were either treated with 20µM betulinic acid (BA) for 24 hours or infected by either empty control (CTL) or ERβ knockdown (shERβ) lentivirus; the cells were then harvested for analysis of proinflammtory cytokine secretion. (A) mRNA levels by qPCR, n=4. (B) IL1β secretion, n=5. (C) IL6 secretion, n=5. (D) TNFα secretion, n=5. (E) PGE2 secretion, n=5. * P < 0.05, vs HEEC/CTL group; ¶ P < 0.05, vs 12Z/CTL group. Data were expressed as mean ± SEM.

Article Snippet: Human Endometrial Epithelial Cells HEEC (#ABC-TC4601) and Immortalized Human Endometriotic Epithelial Cell Line 12Z (#ABI-TC278D) were obtained from ACCEGEN Biotechnology.

Techniques: Knockdown, Infection, Control

Journal: Frontiers in Endocrinology

Article Title: Betulinic Acid Inhibits Endometriosis Through Suppression of Estrogen Receptor β Signaling Pathway

doi: 10.3389/fendo.2020.604648

Figure Lengend Snippet: ERβ knockdown reduces endometriosis-mediated cell proliferation, and betulinic acid treatment mimics this effect. The conditional immortalized HEEC or 12Z cells were either treated with 20 µM betulinic acid (BA) for 24 h or infected by either empty control (CTL) or ERβ knockdown (shERβ) lentivirus; the cells were then harvested for analysis of cell proliferation. (A) Cell proliferation analysis by thymidine incorporation, n = 5. (B) Cell migration assay, n = 5. (C) Cell invasion assay, n = 5. (D) Colony formation assay in soft agar, n = 5. (E) Quantitation of Ki-67 positive cells, n = 3. (F) Representative picture for Ki-67 staining. * P < 0.05, vs HEEC/CTL group; ¶ P < 0.05, vs 12Z/CTL group. Data were expressed as mean ± SEM.

Article Snippet: Human Endometrial Epithelial Cells HEEC (#ABC-TC4601) and Immortalized Human Endometriotic Epithelial Cell Line 12Z (#ABI-TC278D) were obtained from ACCEGEN Biotechnology.

Techniques: Knockdown, Infection, Control, Cell Migration Assay, Invasion Assay, Colony Assay, Quantitation Assay, Staining

Journal: Frontiers in Endocrinology

Article Title: Betulinic Acid Inhibits Endometriosis Through Suppression of Estrogen Receptor β Signaling Pathway

doi: 10.3389/fendo.2020.604648

Figure Lengend Snippet: Gain of ERβ in endometrial cells promotes cell proliferation, while BA treatment diminishes this effect. The conditional immortalized HEEC cells were infected by either empty control (CTL) or ERβ expression (↑ERβ) lentivirus, or ERβ expression (↑ERβ) lentivirus plus 20 µM betulinic acid (BA) (↑ERβ/BA) for 24 h, and the cells were harvested for biomedical analysis. (A) mRNA levels by qPCR, n = 4. (B) ROS formation, n = 5. (C) Quantitation of 3-nitrotyrosine formation, n = 5. (D) Apoptosis rate by TUNEL assay, n = 5. (E) Δψm by TMRE fluorescence, n = 5. (F) IL1β secretion, n = 5. (G) IL6 secretion, n = 5. (H) TNFα secretion, n = 5. (I) PGE2 secretion, n = 5. (J) Cell proliferation analysis by thymidine incorporation, n = 5. (K) Quantitation of Ki-67 positive cells, n = 4. * P < 0.05, vs CTL group; ¶ P < 0.05, vs ↑ERβ group. Data were expressed as mean ± SEM.

Article Snippet: Human Endometrial Epithelial Cells HEEC (#ABC-TC4601) and Immortalized Human Endometriotic Epithelial Cell Line 12Z (#ABI-TC278D) were obtained from ACCEGEN Biotechnology.

Techniques: Infection, Control, Expressing, Quantitation Assay, TUNEL Assay, Fluorescence

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: Technology in Cancer Research & Treatment

Article Title: Sirtuin 2 in Endometrial Cancer: A Potential Regulator for Cell Proliferation, Apoptosis and RAS/ERK Pathway

doi: 10.1177/1533033820980781

Figure Lengend Snippet: Comparison of SIRT2 expression between human EC cell lines and HEEC. (A) Comparison of SIRT2 mRNA expression between human EC cell lines and HEEC. (B) Comparison of SIRT2 protein expression between human EC cell lines and HEEC. SIRT2: sirtuin 2; EC: endometrial cancer; HEEC: human endometrial (uterine) epithelial cells; GAPDH: glyceraldehyde-3-phosphate dehydrogenase.

Article Snippet: Normal human endometrial (uterine) epithelial cells (HEEC) were purchased from Lifeline® Cell Technology (Lifeline® Cell Technology, USA).

Techniques: Comparison, Expressing

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