Journal: Cell Death Discovery

Article Title: Recombinant measles virus vaccine rMV-Hu191 exerts an oncolytic effect on esophageal squamous cell carcinoma via caspase-3/GSDME-mediated pyroptosis

doi: 10.1038/s41420-023-01466-2

Figure Lengend Snippet: A 3 × 10 6 KYSE-30 cells were subcutaneously inoculated into BALB/c nude mice to establish the S.C. tumor model, and then 1 × 10 7 PFU of rMV-Hu191 were administrated intratumorally (I.T.) from day 12 to day 17. B Tumors were measured in the long and short dimensions using a vernier caliper, and tumor volumes were estimated using the equation: V = (length × Width 2 )/2. n = 8 tumors for each group. * p < 0.05, **** p < 0.0001, two-tailed Student’s t -test. The solid line represents the average volume ± SEM. C Difference in tumor volume on day 27 post tumor cell inoculation. * p < 0.05, two-tailed Student’s t -test. The solid line represents the average volume ± SEM. D The Kaplan–Meier survival curves of mice from rMV-Hu191 and mock-treated groups. A log-rank (Mantel–Cox) test was used to analyze the significance of differences between groups. *** p < 0.0001. E Body weights of the mock and rMV-Hu191-treated mice were measured on day 0 and 15 post virus injection. The solid line represents the average weight ± SEM ( n = 8). ns, no significant differences, two-tailed Student’s t -test. F – H The levels of ALT, AST and CR in mouse serum were measured. Data are presented as mean ± SEM ( n = 3). ns, no significant differences, two-tailed Student’s t -test. I , J Immunofluorescence staining of the mock and rMV-Hu191-treated tumors with the indicated antibodies. The scale bars represent 50 μm. K Immunoblot analysis revealed elevated GSDME cleavage in rMV-Hu191-treated tumors. L Schematic model for the regulation of pyroptosis in ESCC by rMV-Hu191. rMV-Hu191 exhibits an antitumor effect through BAK/BAX-dependent caspase-3/GSDME-mediated pyroptosis.

Article Snippet: The human ESCC cell lines KYSE-30, KYSE-150 and KYSE-510 were from the JCRB cell bank (Japanese Collection of Research Bioresources cell bank) and cultured in RMPI 1640 (GIBCO) supplemented with 10% fetal bovine serum (FBS, GIBCO) and 1% penicillin/streptomycin (GIBCO) at 37 °C with 5% CO 2 .

Techniques: Two Tailed Test, Virus, Injection, Immunofluorescence, Staining, Western Blot

Journal: Theranostics

Article Title: Significance of serglycin and its binding partners in autocrine promotion of metastasis in esophageal cancer.

doi: 10.7150/thno.49547

Figure Lengend Snippet: Figure 1. SRGN expression in human ESCC samples and the effects of SRGN overexpression on ESCC cells in vitro and in vivo. (A) The gene expression of SRGN in parental and I-3 cells was evaluated by RT-PCR. (B) Western blot analysis of SRGN in CM and cell lysates of I-3 cells compared with corresponding parental cells. (C) Invasive ability of four SRGN-overexpressing ESCC cell lines were compared with corresponding vector controls using transwell invasion assay. Scale bar, 100 µm. (D) Kaplan– Meier survival curves of 94 patients with ESCC from TCGA (left panel) and the subgroup of patients with stage I/II ESCC (right panel) segregated according to high and low expression of SRGN. (E) Representative images of immunohistchemical staining for SRGN in primary ESCC tumor tissue and matched lymph node (LN) metastasis tissue (upper panel; scale bar, 100 µm). Lower panel shows Fisher’s exact test correlation analysis of SRGN expression in the primary tumor and LN metastasis. (F) Experimental metastasis assay in nude mice showing the effect of SRGN overexpression on colonization of KYSE150-Luc cells to the lungs (n = 7/group).

Article Snippet: Cell lines, inhibitors and recombinant human midkine Human ESCC cell lines KYSE30, KYSE140, KYSE150, KYSE410 [14] (DSMZ, Braunschweig, Germany), and T.Tn [15] were used in this study.

Techniques: Expressing, Over Expression, In Vitro, In Vivo, Gene Expression, Reverse Transcription Polymerase Chain Reaction, Western Blot, Plasmid Preparation, Transwell Invasion Assay, Staining

Journal: Theranostics

Article Title: Significance of serglycin and its binding partners in autocrine promotion of metastasis in esophageal cancer.

doi: 10.7150/thno.49547

Figure Lengend Snippet: Figure 2. SRGN-knockdown suppresses malignant potential of ESCC cells in vitro and in vivo. (A) Flow cytometry was used to compare apoptosis in SRGN-knockdown cells and control cells. (B) Inhibitory effect of SRGN-knockdown by siRNA and shRNA on invasion of ESCC cells. (C) Reduced viability of ESCC cells with SRGN-knockdown, compared with vector controls. (D) Effects of SRGN-knockdown on growth of ESCC tumor xenografts. Upper panels show tumor volume at different time points (n = 6/group), and the excised tumors at endpoint of experiment (15 days and 20 days after subcutanous injection of KYSE150 and KYSE410 cells, respectively). Lower panels show representative sections of tumor xenografts with H&E staining and Ki-67-immunostaining, and Ki-67 proliferation index of subcutaneous tumor xenografts at endpoint of experiment. Scale bar, 200 µm.

Article Snippet: Cell lines, inhibitors and recombinant human midkine Human ESCC cell lines KYSE30, KYSE140, KYSE150, KYSE410 [14] (DSMZ, Braunschweig, Germany), and T.Tn [15] were used in this study.

Techniques: Knockdown, In Vitro, In Vivo, Flow Cytometry, Control, shRNA, Plasmid Preparation, Injection, Staining, Immunostaining

Journal: Theranostics

Article Title: Significance of serglycin and its binding partners in autocrine promotion of metastasis in esophageal cancer.

doi: 10.7150/thno.49547

Figure Lengend Snippet: Figure 3. Significance of GAG attachment domain of SRGN in its promotion of ESCC cell invasion, ERK activation and c-Myc upregulation. (A) Effects of forced expression of intact SRGN, SRGN with truncated GAG (∆GAG), and mutated GAG (mGAG) on invasion (left panel) and migration (right panel) of ESCC cells. (B) Transwell invasion assay showing the effects of the CM collected from KYSE150-SRGN cells as chemoattractant on invasion of KYSE150 and KYSE410 cells, compared with CM from ∆GAG, mGAG, and CON cells. (C) Heatmap of reverse phase protein array (RPPA) analysis performed on cell lysates of four ESCC cell lines with SRGN overexpression compared with their corresponding control cells. The red bars on the right side indicate the proteins associated with the MAPK pathway. (D) Western blotting analysis of MEK/ERK pathway cascade and c-Myc in ESCC cells expressing SRGN, ∆GAG, mGAG, and CON. (E) Western blotting of subcellular fractions (cytosol and nuclear) of ESCC cells with SRGN, ∆GAG, mGAG overexpression. (F) Effect of SRGN-knockdown on MEK/ERK pathway cascade and c-Myc expression in ESCC cells. (G) Comparison of c-Myc expression in tumor xenografts established from SRGN-knockdown cells and vector control cells.

Article Snippet: Cell lines, inhibitors and recombinant human midkine Human ESCC cell lines KYSE30, KYSE140, KYSE150, KYSE410 [14] (DSMZ, Braunschweig, Germany), and T.Tn [15] were used in this study.

Techniques: Activation Assay, Expressing, Migration, Transwell Invasion Assay, Protein Array, Over Expression, Control, Western Blot, Knockdown, Comparison, Plasmid Preparation

Journal: Theranostics

Article Title: Significance of serglycin and its binding partners in autocrine promotion of metastasis in esophageal cancer.

doi: 10.7150/thno.49547

Figure Lengend Snippet: Figure 4. Significance of ERK pathway in SRGN-induced cell invasion and c-Myc stabilization. (A) Effects of SRGN-knockdown on mRNA expression of SRGN, CD44, c-Myc, CCND1 in ESCC cells. (B) SRGN-overexpressing ESCC cells were treated with trametinib (at indicated concentrations) for 24 h, and the SRGN mRNA expression level evaluated by RT-PCR. Cells treated with DMSO served as control. (C) SRGN-overexpressing ESCC cells were treated with 100 nM trametinib for 48 h, and then evaluated by invasion assay (upper panel, scale bar, 100 µm). The quantification is shown in the lower panel. (D) SRGN-overexpressing cells were treated with increasing concentrations of trametinib as indicated for 72 h. The expressions of members of the MEK/ERK pathway cascade and c-Myc were evaluated by western blotting. Untreated vector controls were included for comparison. (E) KYSE150-SRGN cells with or without trametinib (100 nM) treatment were incubated in 100 µg/mL cycloheximide (CHX) for indicated duration before western blotting analysis (left panel). The numbers below the c-Myc blots are the band intensities of c-Myc that were normalized against GAPDH and then expressed relative to that at 0 h time point. The relative c-Myc degradation rate is presented in the graph (right panel). (F) KYSE150 cells with SRGN-knockdown were subjected to CHX chase assay and the band intensities of c-Myc were quantified. CHX chase assay results of KYSE410 cells are shown in Figure S6A-B.

Article Snippet: Cell lines, inhibitors and recombinant human midkine Human ESCC cell lines KYSE30, KYSE140, KYSE150, KYSE410 [14] (DSMZ, Braunschweig, Germany), and T.Tn [15] were used in this study.

Techniques: Knockdown, Expressing, Reverse Transcription Polymerase Chain Reaction, Control, Invasion Assay, Western Blot, Plasmid Preparation, Comparison, Incubation

Journal: Theranostics

Article Title: Significance of serglycin and its binding partners in autocrine promotion of metastasis in esophageal cancer.

doi: 10.7150/thno.49547

Figure Lengend Snippet: Figure 5. SRGN-induced MDK mediates the pro-invasive and ERK/c-Myc-upregulating effects of SRGN. (A) Chemokine profiling of CM from KYSE410 cells expressing control vector (CON), SRGN and ∆GAG, respectively. The intensity of the MDK spots was quantified and presented in the right panel. (B) RT-PCR results showing the effect of SRGN overexpression on mRNA expression of MDK in ESCC cells. (C) The expression of MDK in the CM of ESCC cells with SRGN, ∆GAG, mGAG overexpression was evaluated by western blotting. GAPDH in the cell lysates was used as loading control. (D) The expression of MDK in CM and cell lysates of ESCC cells with SRGN knockdown was compared with that of vector control cells. (E) Correlation between serum SRGN and serum MDK in 100 patients with ESCC. (F) Kaplan–Meier curves comparing the survival outcome of patients with high versus low serum SRGN expression. (G) Effect of MDK overexpression on invasion of KYSE30 and T.Tn cells. (H) Effects of MDK overexpression on p-ERK1/2 and p-AKT expression in KYSE150 and KYSE410 cells. Cells were first transfected with siMDK #3, which targeted the 3’UTR of MDK, to knockdown MDK and then MDK was re-expressed by overexpression of MDK. (I) Effect of rhMDK (500 ng/mL) on invasion of KYSE150 and KYSE410 cells. (J) KYSE150 and KYSE410 cells were treated with 500 ng/mL rhMDK for indicated duration before analysis of MEK/ERK pathway cascade by western blots.

Article Snippet: Cell lines, inhibitors and recombinant human midkine Human ESCC cell lines KYSE30, KYSE140, KYSE150, KYSE410 [14] (DSMZ, Braunschweig, Germany), and T.Tn [15] were used in this study.

Techniques: Expressing, Control, Plasmid Preparation, Reverse Transcription Polymerase Chain Reaction, Over Expression, Western Blot, Knockdown, Transfection

Journal: Theranostics

Article Title: Significance of serglycin and its binding partners in autocrine promotion of metastasis in esophageal cancer.

doi: 10.7150/thno.49547

Figure Lengend Snippet: Figure 6. Effects of MDK suppression on SRGN-induced malignancy of ESCC cells in vitro and in vivo. (A) Cell viability of KYSE150 cells treated with MDK inhibitor (iMDK) at indicated concentrations. (B) The effect of iMDK (10, 50 and 100 nM) on invasion of SRGN-overexpressing cells was evaluated using transwell invasion assay. (C) Effect of iMDK treatment (at indicated concentrations for 72 h) on c-Myc and ERK phosphorylation in SRGN-overexpressing cells. (D) Effect of SRGN overexpression together with MDK-knockdown by siRNA on invasion of ESCC cells. (E) Bioluminescence imaging and quantification of lung metastasis in nude mice (n = 6 /group) 4 weeks after intravenous injection of KYSE150-Luc cells with manipulated SRGN and MDK expressions. The SRGN overexpression (SRGN + shCON), SRGN overexpression with MDK-knockdown (SRGN + shMDK #5), and SRGN-knockdown (CON + shSRGN #3) groups were compared with the control group (CON + shCON).

Article Snippet: Cell lines, inhibitors and recombinant human midkine Human ESCC cell lines KYSE30, KYSE140, KYSE150, KYSE410 [14] (DSMZ, Braunschweig, Germany), and T.Tn [15] were used in this study.

Techniques: In Vitro, In Vivo, Transwell Invasion Assay, Phospho-proteomics, Over Expression, Knockdown, Imaging, Injection, Control

Journal: Theranostics

Article Title: Significance of serglycin and its binding partners in autocrine promotion of metastasis in esophageal cancer.

doi: 10.7150/thno.49547

Figure Lengend Snippet: Figure 7. MDK binds to glycosylated SRGN. (A) Typical images of ESCC cells immunostained for SRGN (red) and MDK (green), and counterstained with 4',6-diamidino-2-phenylindole (DAPI) are shown in the left panel (scale bar, 10 µm). Line-scan profiles of immunofluorescence signals along the white arrows showed co-localization of SRGN and MDK. (B) Proximity ligation assay (PLA) for SRGN and MDK was performed on KYSE150 and KYSE410 cells. After ligation and amplification, the nuclei were counterstained with DAPI. Red spots (indicated by white arrows) represent the binding between SRGN and MDK proteins. Negative control was conducted by replacing SRGN antibody with mouse IgG. Scale bar, 10 µm. (C) Whole cell lysates of MDK-SFB transfected ESCC cells were immunoprecipitated with anti-FLAG M2 beads before immunoblotting for cellular SRGN and CD44. Western blotting showed that FLAG-fusion MDK co-precipitated with SRGN at ~250 kDa (red frames), and with CD44 at over 130 kDa. (D) Cell lysates and CM of FLAG-fusion SRGN-overexpressing (F-SRGN) and FLAG-fusion ∆GAG-expressing (F-∆GAG) cells were incubated with anti-FLAG M2 beads. The core protein of SRGN was predominantly precipitated in cell lysate and glycosylated SRGN was predominantly precipitated in CM. Western blots showed that MDK and MMP2 were bound to glycosylated SRGN, but not to SRGN core protein. MMP9 and CD44 were bound to both glycosylated SRGN and core protein. (E) Proteoglycans isolated from CM of KYSE150-SRGN cells were digested with the indicated enzymes before detection of SRGN using western blotting. (F) Fluorophore-assisted carbohydrate electrophoresis (FACE) analysis of disaccharide products in the CM of KYSE150 cells expressing CON, SRGN, ΔGAG and mGAG, with or without predigestion with chondroitinase ABC. Lanes Std1, Std2, and Std3 contained the standard markers. ∆di-0S, ∆di-6S, ∆di-4S and ∆di-SE in the CM of SRGN-expressing cells are indicated by red arrows.

Article Snippet: Cell lines, inhibitors and recombinant human midkine Human ESCC cell lines KYSE30, KYSE140, KYSE150, KYSE410 [14] (DSMZ, Braunschweig, Germany), and T.Tn [15] were used in this study.

Techniques: Immunofluorescence, Proximity Ligation Assay, Ligation, Amplification, Binding Assay, Negative Control, Transfection, Immunoprecipitation, Western Blot, Expressing, Incubation, Isolation, Electrophoresis

Journal: Cancers

Article Title: Drug Repurposing Applications to Overcome Male Predominance via Targeting G2/M Checkpoint in Human Esophageal Squamous Cell Carcinoma

doi: 10.3390/cancers14235854

Figure Lengend Snippet: Clinical information of ESCC patients.

Article Snippet: The human ESCC cell lines KYSE30, KYSE150, KYSE450, and KYSE510 were generously provided by Dr. Yutaka Shimada.

Techniques:

Journal: Cancers

Article Title: Drug Repurposing Applications to Overcome Male Predominance via Targeting G2/M Checkpoint in Human Esophageal Squamous Cell Carcinoma

doi: 10.3390/cancers14235854

Figure Lengend Snippet: Male ESCC cell lines were more sensitive to decitabine and MK1775 than females. ( a ) Workflow of drug repurposing by using the gene expression data of patients over 60 years old. ( b ) Ten drugs were selected from the drug repurposing results. ( c , d ), Growth curves of KYSE150, KYSE510, KYSE30, and KYSE450 cells were measured by IncuCyte S3 for 72 h. KYSE30 and KYSE450 cells were derived from male patients, while KYSE150 and KYSE510 cells were derived from female patients. Cells were treated with decitabine (10 μM) or MK1775 (200 nM). ( e – g ), Representative image, tumor weights, and tumor volumes of xenografts derived from KYSE30 cells (male), KYSE150 cells (female), and KYSE450 (male) that were treated with decitabine (1.0 mg/kg, i.p.) or MK1775 (60 mg/kg, p.o.). The data shown are the mean ± SD; n = 6 mice per group in KYSE30 and KYSE150; n = 5 mice per group in 450. For tumor weights, data were analyzed using two-tailed t-tests; for tumor volumes, data were analyzed using two-way ANOVA with Bonferroni correction. (* p < 0.05, ** p < 0.01, and *** p < 0.001; ns = not significant).

Article Snippet: The human ESCC cell lines KYSE30, KYSE150, KYSE450, and KYSE510 were generously provided by Dr. Yutaka Shimada.

Techniques: Gene Expression, Derivative Assay, Two Tailed Test

Journal: Cancers

Article Title: Drug Repurposing Applications to Overcome Male Predominance via Targeting G2/M Checkpoint in Human Esophageal Squamous Cell Carcinoma

doi: 10.3390/cancers14235854

Figure Lengend Snippet: Validation RNA-seq proves that MK1775 and decitabine showed a sex-biased treatment response by targeting the G2/M checkpoint. ( a ) Drug-gene network between decitabine and MK1775 and their target genes was assessed in STICH. Gene functional ontology analysis in the ( b ) decitabine KYSE30 cell line, ( c ) MK1775 KYSE30 cell line, and ( d ) MK1775 KYSE150 cell line.

Article Snippet: The human ESCC cell lines KYSE30, KYSE150, KYSE450, and KYSE510 were generously provided by Dr. Yutaka Shimada.

Techniques: Biomarker Discovery, RNA Sequencing, Functional Assay

Journal: Frontiers in Oncology

Article Title: LINC00673 Represses CDKN2C and Promotes the Proliferation of Esophageal Squamous Cell Carcinoma Cells by EZH2-Mediated H3K27 Trimethylation

doi: 10.3389/fonc.2020.01546

Figure Lengend Snippet: Up-regulated LINC00673 is associated with poor prognosis in esophageal squamous cell carcinoma (ESCC) patients. (A) LINC00673 expression levels were evaluated using quantitative PCR (qPCR) in 39 pairs of ESCC tissues and corresponding non-tumor tissues. *** P < 0.001 (unpaired Student’s t test). (B) Receiver operating characteristic (ROC) curve of LINC00673 in 39 ESCC patients. (C–E) Relationships between LINC00673 expression and the differentiation grade, tumor size, and lymph node metastasis, respectively. (F) Kaplan–Meier curves show the survival of ESCC patients, as grouped by the LINC00673 expression levels. * P < 0.05.

Article Snippet: The human ESCC cell lines (KYSE30, KYSE510, and EC9706) were purchased from Guangdong Hybribio Biotech Ltd. (Guangdong, China), and an immortalized normal esophageal epithelial cell line (HET-1A) was purchased from Cinoasia Institute (Shanghai, China).

Techniques: Expressing, Real-time Polymerase Chain Reaction

Journal: Frontiers in Oncology

Article Title: LINC00673 Represses CDKN2C and Promotes the Proliferation of Esophageal Squamous Cell Carcinoma Cells by EZH2-Mediated H3K27 Trimethylation

doi: 10.3389/fonc.2020.01546

Figure Lengend Snippet: LINC00673 knockdown suppresses the proliferation of esophageal squamous cell carcinoma (ESCC) cells in vitro . (A) Cell proliferation was assessed by the CCK8 method in KYSE30 and KYSE510 cells. Bars show the mean ± SD of OD 450 from triplicate samples. (B) Cell proliferation was assessed using a colony forming assay. Bar plots show the average number of colonies (±SD) from triplicate samples. * P < 0.05.

Article Snippet: The human ESCC cell lines (KYSE30, KYSE510, and EC9706) were purchased from Guangdong Hybribio Biotech Ltd. (Guangdong, China), and an immortalized normal esophageal epithelial cell line (HET-1A) was purchased from Cinoasia Institute (Shanghai, China).

Techniques: Knockdown, In Vitro

Journal: Frontiers in Oncology

Article Title: LINC00673 Represses CDKN2C and Promotes the Proliferation of Esophageal Squamous Cell Carcinoma Cells by EZH2-Mediated H3K27 Trimethylation

doi: 10.3389/fonc.2020.01546

Figure Lengend Snippet: LINC00673 knockdown inhibits esophageal squamous cell carcinoma (ESCC) cell proliferation in vivo . (A,B) Stable KYSE30-shLINC00673 and KYSE510-shLINC00673 cells were injected subcutaneously into nude mice. After 4 weeks, the tumors were resected. (C) Tumor growth was measured by Vernier calipers every 3 days starting on the fourth day after injection. Line chart show the tumor growth curve. (D) Box plot showing the tumor weights of the control and LINC00673 knockdown groups. * P < 0.05, *** P < 0.001.

Article Snippet: The human ESCC cell lines (KYSE30, KYSE510, and EC9706) were purchased from Guangdong Hybribio Biotech Ltd. (Guangdong, China), and an immortalized normal esophageal epithelial cell line (HET-1A) was purchased from Cinoasia Institute (Shanghai, China).

Techniques: Knockdown, In Vivo, Injection, Control

Journal: Frontiers in Oncology

Article Title: LINC00673 Represses CDKN2C and Promotes the Proliferation of Esophageal Squamous Cell Carcinoma Cells by EZH2-Mediated H3K27 Trimethylation

doi: 10.3389/fonc.2020.01546

Figure Lengend Snippet: LINC00673 knockdown arrests the cell cycle at the G1/S checkpoint in esophageal squamous cell carcinoma (ESCC) cells. (A,B) Cell cycle examined by a flow cytometry assay. Bar plots show the proportion of cells in the G1 phase, S phase, and G2 phase. * P < 0.05, ** P < 0.01, and *** P < 0.001.

Article Snippet: The human ESCC cell lines (KYSE30, KYSE510, and EC9706) were purchased from Guangdong Hybribio Biotech Ltd. (Guangdong, China), and an immortalized normal esophageal epithelial cell line (HET-1A) was purchased from Cinoasia Institute (Shanghai, China).

Techniques: Knockdown, Flow Cytometry

Journal: Frontiers in Oncology

Article Title: LINC00673 Represses CDKN2C and Promotes the Proliferation of Esophageal Squamous Cell Carcinoma Cells by EZH2-Mediated H3K27 Trimethylation

doi: 10.3389/fonc.2020.01546

Figure Lengend Snippet: LINC00673 knockdown improved CDKN2C levels in esophageal squamous cell carcinoma (ESCC) cells. (A,C) The levels of the G1/S checkpoint regulator expression were detected by Western blotting analysis in stable KYSE30-shLINC00673, KYSE510-shLINC00673, and corresponding control cells. (B,D) Bar plots showing the relative expression levels of the G1/S checkpoint associated regulators. The image was processed by ImageJ software. * P < 0.05, ** P < 0.01.

Article Snippet: The human ESCC cell lines (KYSE30, KYSE510, and EC9706) were purchased from Guangdong Hybribio Biotech Ltd. (Guangdong, China), and an immortalized normal esophageal epithelial cell line (HET-1A) was purchased from Cinoasia Institute (Shanghai, China).

Techniques: Knockdown, Expressing, Western Blot, Control, Software

Journal: Frontiers in Oncology

Article Title: LINC00673 Represses CDKN2C and Promotes the Proliferation of Esophageal Squamous Cell Carcinoma Cells by EZH2-Mediated H3K27 Trimethylation

doi: 10.3389/fonc.2020.01546

Figure Lengend Snippet: Schematic of the regulatory relationships among LINC00673, EZH2, and CDKN2C in ESCC cells. (A,B) Illustrated the effect of up-regulated or down-regulated LINC00673 on ESCC cell proliferation, respectively.

Article Snippet: The human ESCC cell lines (KYSE30, KYSE510, and EC9706) were purchased from Guangdong Hybribio Biotech Ltd. (Guangdong, China), and an immortalized normal esophageal epithelial cell line (HET-1A) was purchased from Cinoasia Institute (Shanghai, China).

Techniques:

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: Dihydrotanshinone I inhibits proliferation of esophageal squamous cell carcinoma cells while showing no proliferation inhibition in normal esophageal epithelial cells. A-E: Cell viability of esophageal squamous cell carcinoma cells (KYSE30, Eca109, KYSE-450, and KYSE-510) and normal esophageal epithelial cells (Het-1A) after treatment with dihydrotanshinone I for 12 h, 24 h, and 36 h. DHTS: Dihydrotanshinone I.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Inhibition

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: Dihydrotanshinone I induces cell cycle arrest in the G0/1 phase in esophageal squamous cell carcinoma cells. A and B: Cell cycle distribution after treatment with DHTS for 24 h in KYSE30 cells and Eca109 cells; C and D: Western blot analysis of proteins associated with the cell cycle in KYSE30 cells and Eca109 cells. a P < 0.05 vs control. DHTS: Dihydrotanshinone I.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Western Blot, Control

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: Dihydrotanshinone I induces apoptosis in esophageal squamous cell carcinoma cells. A and B: Cell apoptosis analyzed using flow cytometry with annexin V-PE/7-ADD double staining in KYSE30 cells and Eca109 cells; C and D: Apoptosis-related morphologic changes in the cells detected using Hoechst 33258 staining in KYSE30 cells and Eca109 cells. a P < 0.05 vs control.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Flow Cytometry, Double Staining, Staining, Control

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: Dihydrotanshinone I activates the mitochondrial pathway in esophageal squamous cell carcinoma cells. A and B: Bax and Bcl2 levels detected using Western blot and the ratio of Bax and Bcl2 calculated in KYSE30 cells and Eca109 cells; C and D: Proteins associated with the mitochondrial pathway detected using Western blot in KYSE30 cells and Eca109 cells. a P < 0.05 vs control.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Western Blot, Control

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: Dihydrotanshinone I inhibits the expression of phosphorylated STAT3 in esophageal squamous cell carcinoma cells. A and B: Levels of STAT3 and pSTAT3 detected using Western blot in KYSE30 cells and Eca109 cells; C and D: Expression and location of phosphorylated STAT3 detected by immunofluorescence in KYSE30 cells and Eca109 cells. pSTAT3: Phosphorylated STAT3.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Expressing, Western Blot, Immunofluorescence

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: STAT3 knockdown promotes dihydrotanshinone I-induced apoptosis in esophageal squamous cell carcinoma cells. A: Knockdown efficiency of STAT3 and phosphorylated STAT3 identified using Western blot in esophageal squamous cell carcinoma cells; B and C: Knockdown efficiency of STAT3 identified using RT-qPCR in KYSE30 cells and Eca109 cells; D: Apoptosis-related morphological changes in STAT3 knockdown cells detected using Hoechst 33258 staining in KYSE30/sh-vector and KYSE30/shSTAT3 cells treated with 0.5 μmol/L DHTS; E: Apoptosis-related morphologic changes detected using Hoechst 33258 staining in Eca109/sh-vector and Eca109/shSTAT3 cells treated with 5 μmol/L DHTS; F: Cell apoptosis analyzed using flow cytometry with annexin V-PE/7-ADD double staining in KYSE30/sh-vector and KYSE30/shSTAT3 cells treated with 0.5 μmol/L DHTS; G: Cell apoptosis analyzed using flow cytometry with annexin V-PE/7-ADD double staining in Eca109/sh-vector and Eca109/shSTAT3 cells treated with 5 μmol/L. a P < 0.05.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Knockdown, Western Blot, Quantitative RT-PCR, Staining, Plasmid Preparation, Flow Cytometry, Double Staining

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: STAT3 knockdown synergizes with dihydrotanshinone I in the activation of the mitochondrial pathway in esophageal squamous cell carcinoma cells. A: KYSE30/sh-vector and KYSE30/shSTAT3 cells were treated with 0.5 μmol/L DHTS, and Western blot was performed to detect the levels of related proteins; B: Eca109/sh-vector and Eca109/shSTAT3 cells were treated with 5 μmol/L dihydrotanshinone I, and Western blot was performed to detect the levels of related proteins.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Knockdown, Activation Assay, Plasmid Preparation, Western Blot

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: STAT3 overexpression blocks the mitochondrial pathway activated by dihydrotanshinone I in esophageal squamous cell carcinoma cells. A: Overexpression efficiency of STAT3 identified using Western blot in esophageal squamous cell carcinoma cells; B and C: Overexpression efficiency of STAT3 identified using RT-qPCR in KYSE30 cells and Eca109 cells; D: KYSE30/OE-vector and KYSE30/OE-STAT3 cells were treated with 0.5 μmol/L DHTS, and Western blot was performed to detect the levels of related proteins; E: Eca109/OE-vector and Eca109/OE-STAT3 cells were treated with 5 μmol/L DHTS, and Western blot was performed to detect the levels of related proteins. OE: Overexpression. a P < 0.05 vs control.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: Over Expression, Western Blot, Quantitative RT-PCR, Plasmid Preparation, Control

Journal: World Journal of Gastrointestinal Oncology

Article Title: STAT3-mediated activation of mitochondrial pathway contributes to antitumor effect of dihydrotanshinone I in esophageal squamous cell carcinoma cells

doi: 10.4251/wjgo.v13.i8.893

Figure Lengend Snippet: Dihydrotanshinone I inhibits proliferation and induces apoptosis in esophageal squamous cell carcinoma in vivo . A: Morphology of the subcutaneous implanted tumors; B: Mean tumor volume at each time point; C: Tumor weight obtained at the end of the experiment; D: Recorded body weight of the mice; E: Immunohistochemistry detection of the expression of Ki67 in the tumor tissue; F and G : A TUNEL assay was performed to detect apoptotic cells in the tumor tissue. a P < 0.05 vs control.

Article Snippet: The human ESCC cell lines KYSE30, Eca109, KYSE450, and KYSE510 and human normal esophageal epithelial cell line Het-1A were donated by the China Center for Type Culture Collection.

Techniques: In Vivo, Immunohistochemistry, Expressing, TUNEL Assay, Control

Journal: International journal of biological sciences

Article Title: M6PR- and EphB4-Rich Exosomes Secreted by Serglycin-Overexpressing Esophageal Cancer Cells Promote Cancer Progression.

doi: 10.7150/ijbs.79875

Figure Lengend Snippet: Figure 1. Inhibition of exosome secretion suppresses pro-invasive effect of SRGN CM on ESCC cells. (A) Effect of DMA treatment on the pro-invasive property of SRGN CM. Left panel, a schematic diagram of the invasion assay; middle panel, representative images of invaded cells; right panel, statistical analysis of the invasion assay. Scale bar, 200 μm. (B) Validation of RAB27A-knockdown efficiency in ESCC cells with SRGN overexpression. Data are presented as mean ± SD. n =3. *, P < 0.05; **, P < 0.01; ***, P < 0.001. (C) Effect of RAB27A-knockdown on the pro-invasive property of SRGN CM. Left panel, a schematic diagram of the invasion assay; middle panel, representative images of the invasion assay; right panel, statistical analysis of the invasion assay. Scale bar, 200 μm. Data are presented as mean ± SD. n = 4. *, P < 0.05; **, P < 0.01; ***, P < 0.001.

Article Snippet: Human ESCC cell lines KYSE30, KYSE150, KYSE410 [15] from DSMZ (Braunschweig, Germany) and T.Tn [16] were used in this study.

Techniques: Inhibition, Invasion Assay, Biomarker Discovery, Knockdown, Over Expression

Journal: International journal of biological sciences

Article Title: M6PR- and EphB4-Rich Exosomes Secreted by Serglycin-Overexpressing Esophageal Cancer Cells Promote Cancer Progression.

doi: 10.7150/ijbs.79875

Figure Lengend Snippet: Figure 2. Exosomes isolated from SRGN-overexpressing ESCC cells enhance the invasion and metastasis of parental ESCC cells. (A) Western blot analysis of density gradient fractions of crude exosomes from ESCC cells. Crude exosomes isolated by differential centrifugation were purified by density gradient ultracentrifugation and equal volumes of each fraction were applied for Western blot analysis. (B-G) Samples obtained from DUGC F6 were used for further analysis. (B) Equal amounts of proteins from cell lysates and exosomes were loaded for comparison by Western blot. (C) Representative whole-mount TEM images of exosomes derived from ESCC cells. (D) Nanoparticle tracking analysis of exosomes isolated from ESCC cells. (E) Quantitative comparison of exosomes secreted by ESCC cells overexpressing SRGN and empty vector by ZetaView® PMX-220 TWIN Laser. Data are presented as mean ± SD. n = 3 and 4 for KYSE410 and KYSE150, respectively. ns, not significant. (F) Effect of exosomes isolated from Con- and SRGN-overexpressing cells on invasion of parental ESCC cells. Left panel, a schematic diagram of the experiment; middle panel, representative images of the invasion assay; right panel, statistical analysis of the invasion assay. Scale bar, 200 μm. Data are presented as mean ± SD. n = 4 and 3 for KYSE410 and KYSE150, respectively. **, P < 0.01; ***, P < 0.001. (G) Effect of SRGN Exo on the colonization of KYSE150-luc cells to lungs of nude mice. Data are presented as mean ± SD. n = 4. ns, not significant; *, P < 0.05.

Article Snippet: Human ESCC cell lines KYSE30, KYSE150, KYSE410 [15] from DSMZ (Braunschweig, Germany) and T.Tn [16] were used in this study.

Techniques: Isolation, Western Blot, Centrifugation, Purification, Comparison, Derivative Assay, Plasmid Preparation, Invasion Assay

Journal: International journal of biological sciences

Article Title: M6PR- and EphB4-Rich Exosomes Secreted by Serglycin-Overexpressing Esophageal Cancer Cells Promote Cancer Progression.

doi: 10.7150/ijbs.79875

Figure Lengend Snippet: Figure 3. Exosomes from SRGN-overexpressing ESCC cells facilitate angiogenesis in vitro. (A) Representative images of uptake of PKH26-labelled exosomes derived from ESCC cells by HUVECs. (B) Representative images of HUVECs treated with Con Exo and SRGN Exo from ESCC cells and (C) corresponding quantifications of number of nodes, junction, segments and meshes. Scale bar, 200 μm. Data are presented as mean ± SD. n = 3 and 4 for KYSE410 and KYSE150, respectively. *, P < 0.05; ***, P < 0.001.

Article Snippet: Human ESCC cell lines KYSE30, KYSE150, KYSE410 [15] from DSMZ (Braunschweig, Germany) and T.Tn [16] were used in this study.

Techniques: In Vitro, Derivative Assay

Journal: International journal of biological sciences

Article Title: M6PR- and EphB4-Rich Exosomes Secreted by Serglycin-Overexpressing Esophageal Cancer Cells Promote Cancer Progression.

doi: 10.7150/ijbs.79875

Figure Lengend Snippet: Figure 4. M6PR and EphB4 are enriched in exosomes from SRGN-overexpressing ESCC cells. (A) GO analysis of differentially expressed proteins in exosomes from KYSE150-SRGN. (B) PANTHER™ Pathway enrichment analysis for differentially expressed proteins in exosomes from KYSE150-SRGN. All enriched pathways were shown. (C) Western blot validation of the upregulated proteins identified by LC-MS/MS in SRGN Exo compared with Con Exo. The concentration of exosomes was measured by Nanosight NS500 and equal numbers of exosomes were used for Western blot analysis. (D) Western blot analysis of M6PR, EphB4, ALIX and CD63 expressions in density gradient fractions of exosomes from ESCC cells. After flotation of crude exosomes in iodixanol gradients, equal volumes of each fraction were used for Western blot analysis.

Article Snippet: Human ESCC cell lines KYSE30, KYSE150, KYSE410 [15] from DSMZ (Braunschweig, Germany) and T.Tn [16] were used in this study.

Techniques: Western Blot, Biomarker Discovery, Liquid Chromatography with Mass Spectroscopy, Concentration Assay

Journal: International journal of biological sciences

Article Title: M6PR- and EphB4-Rich Exosomes Secreted by Serglycin-Overexpressing Esophageal Cancer Cells Promote Cancer Progression.

doi: 10.7150/ijbs.79875

Figure Lengend Snippet: Figure 5. M6PR has prognostic significance in ESCC patients. (A) Correlation analysis between serum SRGN and M6PR in 98 patients with ESCC. (B) Kaplan-Meier curves comparing the survival outcome of ESCC patients with high versus low serum M6PR expression.

Article Snippet: Human ESCC cell lines KYSE30, KYSE150, KYSE410 [15] from DSMZ (Braunschweig, Germany) and T.Tn [16] were used in this study.

Techniques: Expressing

Journal: International journal of biological sciences

Article Title: M6PR- and EphB4-Rich Exosomes Secreted by Serglycin-Overexpressing Esophageal Cancer Cells Promote Cancer Progression.

doi: 10.7150/ijbs.79875

Figure Lengend Snippet: Figure 6. Exosomal M6PR mediates the effect of SRGN on angiogenesis in vitro and in vivo. (A) Effect of M6PR Exo on tube formation ability of HUVECs. Scale bar, 200 μm. Data are presented as mean ± SD. n = 7. ***, P < 0.001. (B) Western blot analysis of M6PR in cell lysates, CM and exosomes of ESCC cells with manipulated SRGN and M6PR expression. (C) Representative images of HUVECs treated with indicated CMs from ESCC cells. Scale bar, 200 μm. (D) Quantification of the numbers of nodes, junctions, segments and meshes in (C). Data are presented as mean ± SD. n = 8 and 3 for KYSE150 and KYSE410, respectively. *, P < 0.05; ***, P < 0.001. (E) Representative images of HUVECs treated with indicated Exo from ESCC cells. Scale bar, 200 μm. (F) Quantification of the numbers of nodes, junctions, segments and meshes in (E). Data are presented as mean ± SD. n = 6. *, P < 0.05; **, P < 0.01; ***, P < 0.001. (G) Effects of exosomes from ESCC cells with SRGN overexpression and M6PR-knockdown and rhM6PR on in vivo angiogenesis. Data are presented as mean ± SD. n = 4. *, P < 0.05.

Article Snippet: Human ESCC cell lines KYSE30, KYSE150, KYSE410 [15] from DSMZ (Braunschweig, Germany) and T.Tn [16] were used in this study.

Techniques: In Vitro, In Vivo, Western Blot, Expressing, Over Expression, Knockdown

Journal: International journal of biological sciences

Article Title: M6PR- and EphB4-Rich Exosomes Secreted by Serglycin-Overexpressing Esophageal Cancer Cells Promote Cancer Progression.

doi: 10.7150/ijbs.79875

Figure Lengend Snippet: Figure 7. Exosomal EphB4 partially mediates the effect of SRGN on invasion of ESCC cells. (A) Validation of EPHB4-knockdown efficiency in ESCC cells with SRGN overexpression. (B) Effect of EPHB4-knockdown on pro-invasive ability of exosomes derived from SRGN-overexpressing ESCC cells. Scale bar, 200 μm. Data are presented as mean ± SD. n = 4. *, P < 0.05; ***, P < 0.001.

Article Snippet: Human ESCC cell lines KYSE30, KYSE150, KYSE410 [15] from DSMZ (Braunschweig, Germany) and T.Tn [16] were used in this study.

Techniques: Biomarker Discovery, Knockdown, Over Expression, Derivative Assay

Journal: Biomedical Reports

Article Title: Peroxiredoxin 4 suppresses ferroptosis in esophageal squamous cell carcinoma by activating the phosphoinositide 3-kinase  signaling pathway

doi: 10.3892/br.2026.2133

Figure Lengend Snippet: PRDX4 exhibits high expression in ESCC tissues and cells. (A) Sangerbox 3.0 online software assay for PRDX4 expression in pan-cancer. (B) UALCAN database investigation for PRDX4 expression in ESCA samples and normal esophageal epithelial tissues. (C) GEO dataset GSE111011 was used to investigate the expression of PRDX4 in ESCC samples and paired normal samples. (D) RT-qPCR assay was used to assess the expression of PRDX4 in 65 ESCC samples and paired normal samples. (E) Western blot analysis of the protein expression of PRDX4 in eight ESCC samples and paired normal samples. (F) The relative protein levels of PRDX4 in eight ESCC samples and paired normal samples. (G) IHC detection of PRDX4 expression in normal tissues and ESCC tissues. Scale bar, 20 µm. (H) Western blot analysis of PRDX4 protein expression in ESCC cell lines (KYSE70, KYSE450, KYSE520, KYSE30 and KYSE270) and normal esophageal epithelial cell line Het-1A. (I) The relative protein levels of PRDX4 in ESCC cell lines and Het-1A cells. (J) RT-qPCR assay of PRDX4 mRNA expression in the aforementioned ESCC cell lines and Het-1A cells. ** P<0.01, *** P<0.001 and **** P<0.0001, indicate statistical significance. PRDX4, peroxiredoxin 4; ESCC, esophageal squamous cell carcinoma; ESCA, esophageal carcinoma; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; GEO, Gene Expression Omnibus; IHC, immunohistochemistry; ns, not significant.

Article Snippet: The human ESCC cell line KYSE30 was purchased from Procell Life Science &Technology Co., Ltd. Normal esophageal epithelial cell line Het-1A and human ESCC cell lines KYSE520, KYSE70, KYSE450, and KYSE270, which were authenticated by STR profiling, were purchased from Qingqi Shanghai Biotechnology Development Co., Ltd. ESCC cells were cultured in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) supplemented with 10% fetal bovine serum (FBS; Suzhou ShuangRu Biotech Co., Ltd.), as well as penicillin (100 U/ml) and streptomycin (0.1 mg/ml) solution (Solarbio Life Sciences), and were incubated at 37 ̊C with 5% CO 2 .

Techniques: Expressing, Software, Quantitative RT-PCR, Western Blot, Reverse Transcription, Real-time Polymerase Chain Reaction, Gene Expression, Immunohistochemistry

Journal: Biomedical Reports

Article Title: Peroxiredoxin 4 suppresses ferroptosis in esophageal squamous cell carcinoma by activating the phosphoinositide 3-kinase  signaling pathway

doi: 10.3892/br.2026.2133

Figure Lengend Snippet: High expression of PRDX4 predicts a poor prognosis in patients with ESCC (A) UALCAN assay of the effects of PRDX4 expression on the survival of patients with ESCA. (B) GEPIA online software assay of the effects of PRDX4 expression on the survival of patients with ESCA. (C) Sangerbox 3.0 online software assay identifying high PRDX4 expression as a poor prognostic factor in patients with ESCA. (D) RT-qPCR assay for PRDX4 expression in patients with ESCC with different TNM stages. (E) RT-qPCR assay of PRDX4 expression in patients with ESCC without lymph node metastasis and with lymph node metastasis. (F) Log-rank test determination of the prognostic value of PRDX4 in patients with ESCC. ** P<0.01 and *** P<0.001, indicate statistical significance. PRDX4, peroxiredoxin 4; ESCC, esophageal squamous cell carcinoma; ESCA, esophageal carcinoma; GEPIA, Gene Expression Profiling Interactive Analysis; RT-qPCR, reverse transcription quantitative polymerase chain reaction; TNM, tumor-node-metastasis.

Article Snippet: The human ESCC cell line KYSE30 was purchased from Procell Life Science &Technology Co., Ltd. Normal esophageal epithelial cell line Het-1A and human ESCC cell lines KYSE520, KYSE70, KYSE450, and KYSE270, which were authenticated by STR profiling, were purchased from Qingqi Shanghai Biotechnology Development Co., Ltd. ESCC cells were cultured in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) supplemented with 10% fetal bovine serum (FBS; Suzhou ShuangRu Biotech Co., Ltd.), as well as penicillin (100 U/ml) and streptomycin (0.1 mg/ml) solution (Solarbio Life Sciences), and were incubated at 37 ̊C with 5% CO 2 .

Techniques: Expressing, Software, Quantitative RT-PCR, Gene Expression, Reverse Transcription, Real-time Polymerase Chain Reaction

Journal: Biomedical Reports

Article Title: Peroxiredoxin 4 suppresses ferroptosis in esophageal squamous cell carcinoma by activating the phosphoinositide 3-kinase  signaling pathway

doi: 10.3892/br.2026.2133

Figure Lengend Snippet: PRDX4 knockdown suppresses cell proliferation in ESCC cells. (A) Western blot analysis of the protein expression of PRDX4 in KYSE270 cells transfected with PRDX4 siRNA and KYSE30 cells transfected with pcDNA3.1-PRDX4. (B) The relative protein levels of PRDX4 in KYSE270 and KYSE30 cells with different transfections. (C) RT-qPCR assay of the mRNA expression of PRDX4 in KYSE270 cells transfected with PRDX4 siRNA and KYSE30 cells transfected with pcDNA3.1-PRDX4. (D) CCK-8 assay of cell proliferation in KYSE270 cells transfected with PRDX4 siRNA. (E) Colony formation assay of the colony-forming ability of KYSE270 cells transfected with PRDX4 siRNA. (F) Statistical analysis of the number of colonies formed in KYSE270 cells transfected with PRDX4 siRNA. (G) CCK-8 assay of cell proliferation in KYSE30 cells transfected with pcDNA3.1-PRDX4. (H) Colony formation assay of the colony-forming ability of KYSE30 cells transfected with pcDNA3.1-PRDX4. (I) Statistical analysis of the number of colonies formed in KYSE30 cells transfected with pcDNA3.1-PRDX4. (J) EdU staining assay of EdU-positive cells in KYSE270 cells transfected with PRDX4 siRNA. Scale bar, 100 µm. (K) EdU staining assay of EdU-positive cells in KYSE30 cells transfected with pcDNA3.1-PRDX4. Scale bar, 100 µm. (L) Statistical analysis of the number of EdU-positive cells in KYSE270 cells transfected with PRDX4 siRNA. (M) Statistical analysis of the number of EdU-positive cells in KYSE30 cells transfected with pcDNA3.1-PRDX4. *** P<0.001 and **** P<0.0001, indicate statistical significance. PRDX4, peroxiredoxin 4; ESCC, esophageal squamous cell carcinoma; siRNA, small interfering RNA; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; CCK-8, Cell Counting Kit-8; EdU, 5-ethynyl-2'-deoxyuridine.

Article Snippet: The human ESCC cell line KYSE30 was purchased from Procell Life Science &Technology Co., Ltd. Normal esophageal epithelial cell line Het-1A and human ESCC cell lines KYSE520, KYSE70, KYSE450, and KYSE270, which were authenticated by STR profiling, were purchased from Qingqi Shanghai Biotechnology Development Co., Ltd. ESCC cells were cultured in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) supplemented with 10% fetal bovine serum (FBS; Suzhou ShuangRu Biotech Co., Ltd.), as well as penicillin (100 U/ml) and streptomycin (0.1 mg/ml) solution (Solarbio Life Sciences), and were incubated at 37 ̊C with 5% CO 2 .

Techniques: Knockdown, Western Blot, Expressing, Transfection, Quantitative RT-PCR, CCK-8 Assay, Colony Assay, Staining, Small Interfering RNA, Reverse Transcription, Real-time Polymerase Chain Reaction, Cell Counting

Journal: Biomedical Reports

Article Title: Peroxiredoxin 4 suppresses ferroptosis in esophageal squamous cell carcinoma by activating the phosphoinositide 3-kinase  signaling pathway

doi: 10.3892/br.2026.2133

Figure Lengend Snippet: PRDX4 downregulation suppresses cell migration and invasion in ESCC cells. (A) PRDX4 knockdown suppresses cell migration and invasion in KYSE270 cells after transfection with PRDX4 siRNA. Scale bar, 100 µm. (B) Statistical analysis of the number of migratory cells in KYSE270 cells transfected with PRDX4 siRNA. (C) Statistical analysis of the number of invasive cells in KYSE270 cells transfected with PRDX4 siRNA. (D) Western blot analysis of the expression levels of E-cadherin, N-cadherin and vimentin in KYSE270 cells transfected with PRDX4 siRNA. (E) The relative protein levels of E-cadherin, N-cadherin and vimentin in KYSE270 cells transfected with PRDX4 siRNA. (F) PRDX4 overexpression suppresses cell migration and invasion in KYSE30 cells after transfection with pcDNA3.1-PRDX4. Scale bar, 100 µm. (G) Statistical analysis of the number of migratory cells in KYSE30 cells transfected with pcDNA3.1-PRDX4. (H) Statistical analysis of the number of invasive cells in KYSE30 cells transfected with pcDNA3.1-PRDX4. (I) Western blot analysis of the expression levels of E-cadherin, N-cadherin and vimentin in KYSE30 cells transfected with pcDNA3.1-PRDX4. (J) The relative protein levels of E-cadherin, N-cadherin and vimentin in KYSE30 cells transfected with pcDNA3.1-PRDX4. *** P<0.001 and **** P<0.0001, indicate statistical significance. PRDX4, peroxiredoxin 4; ESCC, esophageal squamous cell carcinoma; siRNA, small interfering RNA.

Article Snippet: The human ESCC cell line KYSE30 was purchased from Procell Life Science &Technology Co., Ltd. Normal esophageal epithelial cell line Het-1A and human ESCC cell lines KYSE520, KYSE70, KYSE450, and KYSE270, which were authenticated by STR profiling, were purchased from Qingqi Shanghai Biotechnology Development Co., Ltd. ESCC cells were cultured in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) supplemented with 10% fetal bovine serum (FBS; Suzhou ShuangRu Biotech Co., Ltd.), as well as penicillin (100 U/ml) and streptomycin (0.1 mg/ml) solution (Solarbio Life Sciences), and were incubated at 37 ̊C with 5% CO 2 .

Techniques: Migration, Knockdown, Transfection, Western Blot, Expressing, Over Expression, Small Interfering RNA

Journal: Biomedical Reports

Article Title: Peroxiredoxin 4 suppresses ferroptosis in esophageal squamous cell carcinoma by activating the phosphoinositide 3-kinase  signaling pathway

doi: 10.3892/br.2026.2133

Figure Lengend Snippet: PRDX4 is an important regulator of ferroptosis in ESCC cells. (A) Determination of MDA, LPO and GSH contents in KYSE270 cells after transfection with PRDX4 siRNA. (B) Western blot analysis of the protein levels of GPX4, SLC7A11 and PTGS2 in KYSE270 cells transfected with PRDX4 siRNA. (C) The relative protein levels of GPX4, SLC7A11 and PTGS2 in KYSE270 cells transfected with PRDX4 siRNA. (D) Determination of MDA, LPO and GSH contents in KYSE30 cells after transfection with pcDNA3.1-PRDX4. (E) Western blot analysis of the protein levels of GPX4, SLC7A11 and PTGS2 in KYSE30 cells transfected with pcDNA3.1-PRDX4. (F) The relative protein levels of GPX4, SLC7A11 and PTGS2 in KYSE30 cells transfected with pcDNA3.1-PRDX4. (G) Detection of the levels of MDA, LPO and GSH in the control group, PRDX4 siRNA group and PRDX4 siRNA plus Fer-1 group in KYSE270 cells. (H) Western blot analysis of the protein expression levels of GPX4, SLC7A11 and PTGS2 in the control group, PRDX4 siRNA group and PRDX4 siRNA plus Fer-1 group in KYSE270 cells. (I) The relative protein levels of GPX4, SLC7A11 and PTGS2 in the control group, PRDX4 siRNA group and PRDX4 siRNA plus Fer-1 group in KYSE270 cells. (J) Detection of the levels of MDA, LPO and GSH in the pcDNA3.1 group, pcDNA3.1-PRDX4 group and pcDNA3.1-PRDX4 plus erastin group in KYSE30 cells. (K) Western blot analysis of the protein expression levels of GPX4, SLC7A11 and PTGS2 in the pcDNA3.1 group, pcDNA3.1-PRDX4 group and pcDNA3.1-PRDX4 plus erastin group in KYSE30 cells. (L) The relative protein levels of GPX4, SLC7A11 and PTGS2 in the pcDNA3.1 group, pcDNA3.1-PRDX4 group and pcDNA3.1-PRDX4 plus erastin group in KYSE30 cells. ** P<0.01, *** P<0.001 and **** P<0.0001, indicate statistical significance. PRDX4, peroxiredoxin 4; ESCC, esophageal squamous cell carcinoma; siRNA, small interfering RNA; MDA, malondialdehyde; LPO, lipid peroxidation; GSH, glutathione; GPX4, glutathione peroxidase 4; SLC7A11, solute carrier family 7 member 11; PTGS2, prostaglandin-endoperoxide synthase 2; Fer-1, ferrostatin-1; ns, not significant.

Article Snippet: The human ESCC cell line KYSE30 was purchased from Procell Life Science &Technology Co., Ltd. Normal esophageal epithelial cell line Het-1A and human ESCC cell lines KYSE520, KYSE70, KYSE450, and KYSE270, which were authenticated by STR profiling, were purchased from Qingqi Shanghai Biotechnology Development Co., Ltd. ESCC cells were cultured in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) supplemented with 10% fetal bovine serum (FBS; Suzhou ShuangRu Biotech Co., Ltd.), as well as penicillin (100 U/ml) and streptomycin (0.1 mg/ml) solution (Solarbio Life Sciences), and were incubated at 37 ̊C with 5% CO 2 .

Techniques: Transfection, Western Blot, Control, Expressing, Small Interfering RNA

Journal: Biomedical Reports

Article Title: Peroxiredoxin 4 suppresses ferroptosis in esophageal squamous cell carcinoma by activating the phosphoinositide 3-kinase  signaling pathway

doi: 10.3892/br.2026.2133

Figure Lengend Snippet: PRDX4 suppresses ferroptosis of ESCC cells by activating the PI3K/AKT signaling pathway. (A) Determination of MDA, LPO and GSH contents in the absence or presence of the PI3K activator 740 Y-P after PRDX4 knockdown in KYSE270 cells. (B) Western blot analysis of the protein expression levels of GPX4, p-PI3K, PI3K, p-AKT and AKT in the absence or presence of the PI3K activator 740 Y-P after PRDX4 knockdown in KYSE270 cells. (C) The relative protein levels of GPX4, p-PI3K, PI3K, p-AKT and AKT in the absence or presence of the PI3K activator 740 Y-P after PRDX4 knockdown in KYSE270 cells. (D) Detection of MDA, LPO and GSH contents in the absence or presence of the PI3K inhibitor LY294002 after PRDX4 overexpression in KYSE30 cells. (E) Western blot analysis of the protein expression levels of GPX4, p-PI3K, PI3K, p-AKT and AKT in the absence or presence of the PI3K inhibitor LY294002 after PRDX4 overexpression in KYSE30 cells. (F) The relative protein levels of GPX4, p-PI3K, PI3K, p-AKT and AKT in the absence or presence of the PI3K inhibitor LY294002 after PRDX4 overexpression in KYSE30 cells. * P<0.05, ** P<0.01, and **** P<0.0001, indicate statistical significance. PRDX4, peroxiredoxin 4; ESCC, esophageal squamous cell carcinoma; PI3K, phosphoinositide 3-kinase; AKT, protein kinase B; MDA, malondialdehyde; LPO, lipid peroxidation; GSH, glutathione; GPX4, glutathione peroxidase 4; p-PI3K, phosphorylated PI3K; p-AKT, phosphorylated AKT; ns, not significant.

Article Snippet: The human ESCC cell line KYSE30 was purchased from Procell Life Science &Technology Co., Ltd. Normal esophageal epithelial cell line Het-1A and human ESCC cell lines KYSE520, KYSE70, KYSE450, and KYSE270, which were authenticated by STR profiling, were purchased from Qingqi Shanghai Biotechnology Development Co., Ltd. ESCC cells were cultured in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) supplemented with 10% fetal bovine serum (FBS; Suzhou ShuangRu Biotech Co., Ltd.), as well as penicillin (100 U/ml) and streptomycin (0.1 mg/ml) solution (Solarbio Life Sciences), and were incubated at 37 ̊C with 5% CO 2 .

Techniques: Knockdown, Western Blot, Expressing, Over Expression

Journal: Biomedical Reports

Article Title: Peroxiredoxin 4 suppresses ferroptosis in esophageal squamous cell carcinoma by activating the phosphoinositide 3-kinase  signaling pathway

doi: 10.3892/br.2026.2133

Figure Lengend Snippet: Proposed model of PRDX4-mediated suppression of ferroptosis through regulation of the PI3K/AKT pathway in ESCC. PRDX4 is highly expressed in ESCC samples and cells. High PRDX4 expression is strongly associated with TNM staging and lymph node metastasis in patients with ESCC and may serve as an indicator of prognosis for patients with ESCA. PRDX4 knockdown suppresses cell proliferation and invasion of ESCC cells by inactivating the PI3K/AKT signaling pathway, thereby triggering ferroptosis in these cells. PRDX4, peroxiredoxin 4; PI3K, phosphoinositide 3-kinase; AKT, protein kinase B; ESCC, esophageal squamous cell carcinoma; TNM, tumor-node-metastasis; ESCA, esophageal carcinoma.

Article Snippet: The human ESCC cell line KYSE30 was purchased from Procell Life Science &Technology Co., Ltd. Normal esophageal epithelial cell line Het-1A and human ESCC cell lines KYSE520, KYSE70, KYSE450, and KYSE270, which were authenticated by STR profiling, were purchased from Qingqi Shanghai Biotechnology Development Co., Ltd. ESCC cells were cultured in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) supplemented with 10% fetal bovine serum (FBS; Suzhou ShuangRu Biotech Co., Ltd.), as well as penicillin (100 U/ml) and streptomycin (0.1 mg/ml) solution (Solarbio Life Sciences), and were incubated at 37 ̊C with 5% CO 2 .

Techniques: Expressing, Knockdown