Journal: Frontiers in Oncology

Article Title: Targeting NANOS1 in triple-negative breast cancer: synergistic effects of digoxin and PD-1 inhibitors in modulating the tumor immune microenvironment

doi: 10.3389/fonc.2024.1536406

Figure Lengend Snippet: TOE suppresses malignant phenotype of triple negative breast cancer cells. (A) The effect of TOE on cell proliferation in MDA-MB-231 and 4T1 cells. Cells were treated with varying concentrations of TOE for 24 hours, and cell proliferation was assessed using the CCK-8 assay. Data are expressed as the mean ± SEM (n = 3). Statistical significances were calculated via Student’s t-test. **p < 0.01 and ***p < 0.001 and ****p < 0.0001. (B) Transwell migration and invasion assay of MDA-MB-231 and 4T1 cells after treatment with TOE for 24 hours. Representative images of the migrated and invaded cells from randomly selected fields of Transwell inserts are shown on the left, while quantitative data for cell numbers are presented on the right. Cell numbers were calculated and expressed as the mean ± SEM of three independent experiments. Statistical significance was determined by t-test, with ** p < 0.01 and *** p < 0.001 and **** p < 0.0001 indicating significant differences between TOE-treated and DMSO-treated cells. Scale bar = 100 μm. (C) MA plot of DGEs in MDA-MB-231 treated with TOE. (D) Enrichment and scatter map of KEGG pathway of DGEs.

Article Snippet: MDA-MB-231 or 4T1 cells were seeded in a 96‐well plate at 2.0×10^5 cells/mL and cultured in medium at 37°C with a 5% CO2 atmosphere for 24 h. After that, the cells were pretreated with various concentrations of TOE for 24 h. At the end of the stimulation, the medium was removed, and 20 μL of CCK-8 reagent (Elabscience, China) was added to each well, and the incubation was continued for 1 h. The absorbance of each well was measured at 450 nm.

Techniques: CCK-8 Assay, Migration, Invasion Assay

Journal: Frontiers in Oncology

Article Title: Targeting NANOS1 in triple-negative breast cancer: synergistic effects of digoxin and PD-1 inhibitors in modulating the tumor immune microenvironment

doi: 10.3389/fonc.2024.1536406

Figure Lengend Snippet: Dig and AA inhibited tumor growth in breast cancer mouse models. (A) Inhibition of growth by Dig and AA in MDA-MB-231 and 4T1 cells for 24 h. MDA-MB-231 and 4T1 cells were treated with Dig and AA (at various concentration) for 24 hours, and cell proliferation was assessed using the CCK-8 assay. Data are presented as the mean ± SEM from three independent experiments (n = 3). Statistical significance was determined using unpaired t -tests, with * p < 0.05, ** p < 0.01, *** p < 0.001, and **** p < 0.0001 indicating significant differences compared to the DMSO control. (B) Transwell migration and invasion assay of MDA-MB-231 and 4T1 cells with Dig and AA treatment for 24 h. Representative images from randomly selected fields of transwell inserts, and Scalebar = 100 μm. (C) Quantitative data from the Transwell migration and invasion assays. Cell numbers were calculated and are expressed as the mean ± SEM of three independent experiments. * p < 0.05,** p < 0.01, *** p < 0.001 and **** p < 0.0001, as determined by unpaired t -tests. (D) Diagrammatic representation of tumor volume measurement. The diagram illustrates the measurement method, including caliper-based measurements of length and width used to calculate tumor volume (Volume = 1/2 × length × width^2). (E) Tumor sizes at day 14. (F) The body weight changes of mice in the period of 14 days after different treatments. The body weight of mice was monitored every 2 days after Dig and AA treatment. Data are expressed as the mean ± SEM. No significant changes in body weight were observed, suggesting that the treatments did not cause overt toxicity in mice.

Article Snippet: MDA-MB-231 or 4T1 cells were seeded in a 96‐well plate at 2.0×10^5 cells/mL and cultured in medium at 37°C with a 5% CO2 atmosphere for 24 h. After that, the cells were pretreated with various concentrations of TOE for 24 h. At the end of the stimulation, the medium was removed, and 20 μL of CCK-8 reagent (Elabscience, China) was added to each well, and the incubation was continued for 1 h. The absorbance of each well was measured at 450 nm.

Techniques: Inhibition, Concentration Assay, CCK-8 Assay, Control, Migration, Invasion Assay

Journal: Frontiers in Oncology

Article Title: Targeting NANOS1 in triple-negative breast cancer: synergistic effects of digoxin and PD-1 inhibitors in modulating the tumor immune microenvironment

doi: 10.3389/fonc.2024.1536406

Figure Lengend Snippet: Dig combined with PD-1 immunotherapy can promote immune stimulation of in situ 4T1 breast tumors. (A) HE staining of mouse tumor tissues (scale bar = 100 μm). (B) Representative flow cytometry plots showing tumor-associated macrophages (TAMs) (CD45.2+, CD11b+, F4/80+) were obtained after different treatments. (C) Representative flow cytometry plots showing tumor immune cells after different treatments, including CTLs (CD45+, CD3+, CD8+) and Th cells (CD45+, CD3+, CD4+). (D) Representative flow cytometry plots demonstrating tumor-infiltrating LAG-3+ exhausted T cells (CD3+, CD8+, LAG-3+) after different treatments. (E) Representative flow cytometry plots demonstrating tumor-infiltrating TIM-3+ exhausted T cells (CD3+, CD8+, TIM-3+) after different treatments. (F) Representative flow cytometry plots demonstrating tumor-infiltrating PD-1+ exhausted T cells (CD3+, CD8+, PD-1+) after different treatments. (G) The levels of TAMs were quantified through flow cytometry analysis (n = 5). Statistical analysis was performed using Tukey’s multiple comparison test. The following adjusted p-values were obtained for each comparison: Saline vs PD-1: 0.0033; Saline vs Dig: 0.0106; Saline vs Dig+PD-1: <0.0001; Saline vs AA: 0.2039; Saline vs AA+PD-1: 0.9935; PD-1 vs Dig: 0.9961; PD-1 vs Dig+PD-1: 0.4395; PD-1 vs AA: 0.4330; PD-1 vs AA+PD-1: 0.0009; Dig vs Dig+PD-1: 0.2080; Dig vs AA: 0.7273; Dig vs AA+PD-1: 0.0028; Dig+PD-1 vs AA: 0.0109; Dig+PD-1 vs AA+PD-1: <0.0001; AA vs AA+PD-1: 0.0715. (H) The levels of CTLs were quantified by flow cytometry analysis (n = 5). The following adjusted p-values were obtained for each comparison: Saline vs PD-1: 0.7941; Saline vs Dig: 0.2550; Saline vs Dig+PD-1: <0.0001; Saline vs AA: 0.0161; Saline vs AA+PD-1: <0.0001; PD-1 vs Dig: 0.9237; PD-1 vs Dig+PD-1: 0.0013; PD-1 vs AA: 0.2253; PD-1 vs AA+PD-1: 0.0016; Dig vs Dig+PD-1: 0.0136; Dig vs AA: 0.7542; Dig vs AA+PD-1: 0.0164; Dig+PD-1 vs AA: 0.2253; Dig+PD-1 vs AA+PD-1: >0.9999; AA vs AA+PD-1: 0.2576. (I) Flow cytometry analysis quantified the levels of LAG-3+ exhausted T cells (n = 5). Statistical analysis was performed using Tukey’s multiple comparison test. Flow cytometry analysis quantified the levels of LAG-3+ exhausted T cells (n = 5). Statistical analysis was performed using Tukey’s multiple comparison test. The following adjusted p-values were obtained for each comparison: Saline vs PD-1: 0.0006; Saline vs Dig: 0.0030; Saline vs Dig+PD-1: <0.0001; Saline vs AA: 0.5950; Saline vs AA+PD-1: >0.9999; PD-1 vs Dig: 0.9841; PD-1 vs Dig+PD-1: 0.0094; PD-1 vs AA: 0.0282; PD-1 vs AA+PD-1: 0.0005; Dig vs Dig+PD-1: 0.0019; Dig vs AA: 0.1152; Dig vs AA+PD-1: 0.0027; Dig+PD-1 vs AA: <0.0001; Dig+PD-1 vs AA+PD-1: <0.0001; AA vs AA+PD-1: 0.5676. (J) Flow cytometry analysis quantified the levels of TIM-3+ exhausted T cells (n = 5). Statistical analysis was performed using Tukey’s multiple comparison test. The following adjusted p-values were obtained for each comparison: Saline vs PD-1: 0.0073; Saline vs Dig: 0.3784; Saline vs Dig+PD-1: <0.0001; Saline vs AA: 0.9296; Saline vs AA+PD-1: 0.3206; PD-1 vs Dig: 0.4016; PD-1 vs Dig+PD-1: 0.0959; PD-1 vs AA: 0.0007; PD-1 vs AA+PD-1: <0.0001; Dig vs Dig+PD-1: 0.0011; Dig vs AA: 0.0698; Dig vs AA+PD-1: 0.0050; Dig+PD-1 vs AA: <0.0001; Dig+PD-1 vs AA+PD-1: <0.0001; AA vs AA+PD-1: 0.8546. (K) Flow cytometry analysis quantified the levels of PD-1+ exhausted T cells (n = 5). Statistical analysis was performed using Tukey’s multiple comparison test. The following adjusted p-values were obtained for each comparison: Saline vs PD-1: <0.0001; Saline vs Dig: 0.2205; Saline vs Dig+PD-1: 0.0276; Saline vs AA: 0.9984; Saline vs AA+PD-1: 0.8260; PD-1 vs Dig: 0.0043; PD-1 vs Dig+PD-1: 0.0469; PD-1 vs AA: <0.0001; PD-1 vs AA+PD-1: <0.0001; Dig vs Dig+PD-1: 0.9030; Dig vs AA: 0.1042; Dig vs AA+PD-1: 0.0181; Dig+PD-1 vs AA: 0.0108; Dig+PD-1 vs AA+PD-1: 0.0015; AA vs AA+PD-1: 0.9632. Data are expressed as the mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001 vs. Saline.

Article Snippet: MDA-MB-231 or 4T1 cells were seeded in a 96‐well plate at 2.0×10^5 cells/mL and cultured in medium at 37°C with a 5% CO2 atmosphere for 24 h. After that, the cells were pretreated with various concentrations of TOE for 24 h. At the end of the stimulation, the medium was removed, and 20 μL of CCK-8 reagent (Elabscience, China) was added to each well, and the incubation was continued for 1 h. The absorbance of each well was measured at 450 nm.

Techniques: In Situ, Staining, Flow Cytometry, Comparison, Saline

Journal: Frontiers in Oncology

Article Title: Targeting NANOS1 in triple-negative breast cancer: synergistic effects of digoxin and PD-1 inhibitors in modulating the tumor immune microenvironment

doi: 10.3389/fonc.2024.1536406

Figure Lengend Snippet: NANOS1 Silencing Reduces TNF-α Expression and Cell Invasiveness. (A) Volcano plots of RNA-seq. Differential gene expression analysis was performed using RNA sequencing data from MDA-MB-231 cells with siNANOS1#1 silencing. The volcano plot shows the distribution of genes based on log-fold change versus the negative log-transformed p-value. Significant upregulated and downregulated genes are indicated with colored dots. Genes that meet the threshold of log-fold change (|logFC| > 2) and adjusted p-value < 0.05 are considered differentially expressed. (B) Heatmap of differential gene expression. (C) Chord diagram of GO enrichment results and related genes. (D) Top 15 KEGG pathways. The x-axis represents the gene ratio ( p < 0.05), and the y-axis represents the enriched terms. (E) The knockdown efficiency of siRNA and the expression of NANOS1 and TNF-α following Dig and AA treatment were quantified by qPCR. MDA-MB-231 and 4T1 cells were transfected with siRNA targeting NANOS1, followed by treatment with Dig and AA. Quantitative PCR was performed to assess the expression levels of NANOS1 and TNF-α. The data are expressed as the mean ± SEM, with statistical significance calculated using one-way ANOVA. (F) Dig and AA decreased the expression of NANOS1 protein (n = 3). Data are expressed as the mean ± SEM. Statistical significances were calculated via one-way ANOVA, * p < 0.05, ** p < 0.010, *** p < 0.001 and **** p < 0.0001 vs. DMSO. (G) Perforation migration and invasion assays of MDA-MB-231 and 4T1 cells after 24 h of siRNA treatment, scale bar = 100 mm. Cell numbers were calculated and are expressed as the mean ± SEM of three independent experiments. * p < 0.05, ** p < 0.01, *** p < 0.001 and **** p < 0.0001, as determined by unpaired t-tests, were regarded as significant.

Article Snippet: MDA-MB-231 or 4T1 cells were seeded in a 96‐well plate at 2.0×10^5 cells/mL and cultured in medium at 37°C with a 5% CO2 atmosphere for 24 h. After that, the cells were pretreated with various concentrations of TOE for 24 h. At the end of the stimulation, the medium was removed, and 20 μL of CCK-8 reagent (Elabscience, China) was added to each well, and the incubation was continued for 1 h. The absorbance of each well was measured at 450 nm.

Techniques: Expressing, RNA Sequencing, Gene Expression, Transformation Assay, Knockdown, Transfection, Real-time Polymerase Chain Reaction, Migration

Journal: Toxics

Article Title: Selenium Nanoparticles Attenuate Cobalt Nanoparticle-Induced Skeletal Muscle Injury: A Study Based on Myoblasts and Zebrafish

doi: 10.3390/toxics12020130

Figure Lengend Snippet: Cellular toxicity induced by cobalt and selenium nanoparticles. ( A ) Cells were treated with 5 μg/mL FITC-labeled CoNPs and SeNPs for 2 h. Confocal microscopy revealed fluorescent signals into the cells. Yellow arrows indicating cells that are filled with green fluorescence, while the culture medium contains free FITC-labeled CoNPs and SeNPs. ( B – D ) Viability of C2C12 cells treated with 5–80 μg/mL CoNPs, SeNPs, and mixing of CoNPs and SeNPs were determined by CCK-8. Data are presented as mean ± standard deviation of three identical experiments conducted in triplicate. * Statistically significant difference compared with the controls ( p < 0.05 for each).

Article Snippet: According to the manufacturer’s instructions, the levels of MDA (malondialdehyde), GSH (glutathione), and SOD (superoxide dismutase) in C2C12 cells were measured using a GSH ELISA Kit (Elabscience, Wuhan, China), MDA Content Detection Kit (Beyotime, Nantong, China), and SOD Activity Detection Kit (Beyotime, Nantong, China).

Techniques: Labeling, Confocal Microscopy, Fluorescence, CCK-8 Assay, Standard Deviation

Journal: Toxics

Article Title: Selenium Nanoparticles Attenuate Cobalt Nanoparticle-Induced Skeletal Muscle Injury: A Study Based on Myoblasts and Zebrafish

doi: 10.3390/toxics12020130

Figure Lengend Snippet: Protective effects of low-dose SeNPs on CoNP-induced oxidative stress in muscle cells. ( A ) C2C12 cells were exposed to control, 20 μg/mL CoNPs, 500 µM H2O2, 5 μg/mL SeNPs with or without H2O2, and mixing of Co and Se NPs. Oxidative stress condition induced by H2O2 treatment for 4 h. Intracellular ROS production was analyzed by H2DCFDA staining. ( B – E ) MDA, GSH, and SOD levels were determined in C2C12 cells from control, 20 μg/mL CoNPs, 500 µM H2O2, 5 μg/mL SeNPs with or without H2O2, and mixing of Co and Se NP-treated groups. Data are displayed as mean ± SD; * means p < 0.05 between two indicated groups (n = 10).

Article Snippet: According to the manufacturer’s instructions, the levels of MDA (malondialdehyde), GSH (glutathione), and SOD (superoxide dismutase) in C2C12 cells were measured using a GSH ELISA Kit (Elabscience, Wuhan, China), MDA Content Detection Kit (Beyotime, Nantong, China), and SOD Activity Detection Kit (Beyotime, Nantong, China).

Techniques: Control, Staining

Journal: Toxics

Article Title: Selenium Nanoparticles Attenuate Cobalt Nanoparticle-Induced Skeletal Muscle Injury: A Study Based on Myoblasts and Zebrafish

doi: 10.3390/toxics12020130

Figure Lengend Snippet: CoNP-induced apoptosis was inhibited by SeNPs in muscle cells. ( A , B ) The expression of caspase–3 and cleaved caspase–3 in C2C12 cells treated with 5 μg/mL SeNPs or 20 μg/mL CoNPs with or without SeNPs was analyzed by Western blotting. GAPDH was used as internal reference. Relative expression levels are shown in the graph. ( C , D ) The degree of apoptosis of cells subjected to different treatments was determined by annexin V-FITC/PI double-staining flow cytometry. The percentage of apoptotic cells was counted and compared between the different groups. Data are displayed as mean ± SD; * means p < 0.05 between two indicated groups (n = 3).

Article Snippet: According to the manufacturer’s instructions, the levels of MDA (malondialdehyde), GSH (glutathione), and SOD (superoxide dismutase) in C2C12 cells were measured using a GSH ELISA Kit (Elabscience, Wuhan, China), MDA Content Detection Kit (Beyotime, Nantong, China), and SOD Activity Detection Kit (Beyotime, Nantong, China).

Techniques: Expressing, Western Blot, Double Staining, Flow Cytometry

Journal: Toxics

Article Title: Selenium Nanoparticles Attenuate Cobalt Nanoparticle-Induced Skeletal Muscle Injury: A Study Based on Myoblasts and Zebrafish

doi: 10.3390/toxics12020130

Figure Lengend Snippet: Promotive effects of SeNPs on CoNP-induced inhibition of myogenic differentiation. ( A ) Immunofluorescence staining in SeNPs, CoNPs, and Co Se NP-treated C2C12 cells was analyzed at 4 days of differentiation to detect the effect of different nanoparticles on myogenic differentiation. Nuclei (DAPI, blue), F-actin (Green), and α-SMA (Red) were labeled; scale bar = 25 µm. ( B ) Quantification of α-SMA fluorescence intensity in ( A ) was performed. Data are displayed as mean ± SD. * means p < 0.05 between two indicated groups (n = 6). ( C – E ) mRNA expression of myogenic markers, MyoD, myogenin, and Myf5, in C2C12 cells treated with 5 μg/mL SeNPs, 20 μg/mL CoNPs with or without SeNPs was analyzed by qPCR. The control group was set to 1.0. Data are displayed as mean ± SD. * means p < 0.05 between two indicated groups (n = 6).

Article Snippet: According to the manufacturer’s instructions, the levels of MDA (malondialdehyde), GSH (glutathione), and SOD (superoxide dismutase) in C2C12 cells were measured using a GSH ELISA Kit (Elabscience, Wuhan, China), MDA Content Detection Kit (Beyotime, Nantong, China), and SOD Activity Detection Kit (Beyotime, Nantong, China).

Techniques: Inhibition, Immunofluorescence, Staining, Labeling, Fluorescence, Expressing, Control

Journal: International Journal of Molecular Sciences

Article Title: In Vitro Effects of Papaverine on Cell Migration and Vascular Endothelial Growth Factor in Cancer Cell Lines

doi: 10.3390/ijms23094654

Figure Lengend Snippet: The relative percentage migration compared to MDA-MB-231 ( A ), A549 ( B ), and DU145 ( C ) cells at 0 h exposure when cells were exposed to PPV (10–150 µM). The average of three independent experiments is represented by the graph with error bars indicating standard deviation. The blue bar represents the percentage of the cell migration of the cells after 18 h of exposure, the orange bar represents the percentage of cell migration of cells after 24 h of exposure, and the grey bar represents the percentage of cell migration of cells after 48 h of exposure. The statistical significance of cell migration after 18 h of exposure is indicated with an asterisk (**), the statistical significance of cell migration after 24 h of exposure is indicated with two asterisks (***), and the statistical significance of cell migration after 48 h of exposure indicated with three asterisks (*). The positive control for the migration assay-included cells exposed to 2-ethyl-17-hydroxy-13-methyl-7,8,9,11,12,13,14,15,16,17-decahydro-6-cyclopenta[a]phenanthren-3-yl sulphamate (ESE-ol) (0.4 µM) for 48 h. Statistical significance is represented by an * when using the student t -test with a p value of 0.05 compared to cells propagated in complete growth medium.

Article Snippet: The effects of PPV on the expression of VEGF ligand B and receptors 1 and 2 were investigated in MDA-MB-231, A549, and DU145 cells using a VEGF B ELISA kit (Elabscience biotechnology incorporated (Houston, TX, USA)) a VEGF R1 ELISA kit and a VEGF R2 ELISA kit (Abcam plc. (Cambridge, UK).

Techniques: Migration, Standard Deviation, Positive Control

Journal: International Journal of Molecular Sciences

Article Title: In Vitro Effects of Papaverine on Cell Migration and Vascular Endothelial Growth Factor in Cancer Cell Lines

doi: 10.3390/ijms23094654

Figure Lengend Snippet: Spectrophotometry results of VEGF B ELISA demonstrating the effects of PPV (10–300 µM) on proliferation on MDA-MB-231 cells compared to A549- and DU145 cell lines at 48 h. The average of three independent experiments is represented by the graph with error bars indicating standard deviation. The statistical significance of MDA-MB-231 cells is indicated with an asterisk (*), the statistical significance of A549 cells is indicated with two asterisks (**), and the statistical significance of DU145 cells is indicated with three asterisks (***). Statistical significance is represented by an * when using the student t -test with a p value of 0.05 compared to cells propagated in complete growth medium.

Article Snippet: The effects of PPV on the expression of VEGF ligand B and receptors 1 and 2 were investigated in MDA-MB-231, A549, and DU145 cells using a VEGF B ELISA kit (Elabscience biotechnology incorporated (Houston, TX, USA)) a VEGF R1 ELISA kit and a VEGF R2 ELISA kit (Abcam plc. (Cambridge, UK).

Techniques: Spectrophotometry, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: In Vitro Effects of Papaverine on Cell Migration and Vascular Endothelial Growth Factor in Cancer Cell Lines

doi: 10.3390/ijms23094654

Figure Lengend Snippet: Spectrophotometry results of VEGF B ELISA demonstrating the effects of PPV (10–300 µM) on proliferation on MDA-MB-231 cells compared to A549- and DU145 cell lines at 72 h. The average of three independent experiments is represented by the graph with error bars indicating standard deviation. The statistical significance of MDA-MB-231 cells is indicated with an asterisk (*), the statistical significance of A549 cells is indicated with two asterisks (**), and the statistical significance of DU145 cells is indicated with three asterisks (***). Statistical significance is represented by an * when using the student t -test with a p value of 0.05 compared to cells propagated in complete growth medium.

Article Snippet: The effects of PPV on the expression of VEGF ligand B and receptors 1 and 2 were investigated in MDA-MB-231, A549, and DU145 cells using a VEGF B ELISA kit (Elabscience biotechnology incorporated (Houston, TX, USA)) a VEGF R1 ELISA kit and a VEGF R2 ELISA kit (Abcam plc. (Cambridge, UK).

Techniques: Spectrophotometry, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: In Vitro Effects of Papaverine on Cell Migration and Vascular Endothelial Growth Factor in Cancer Cell Lines

doi: 10.3390/ijms23094654

Figure Lengend Snippet: Spectrophotometry results of VEGF R1 ELISA demonstrating the effects of PPV (10–300 µM) on proliferation on MDA-MB-231 cells compared to A549- and DU145 cell lines at 48 h. The average of three independent experiments is represented by the graph with error bars indicating standard deviation. The statistical significance of MDA-MB-231 cells is indicated with an asterisk (*), the statistical significance of A549 cells is indicated with two asterisks (**), and the statistical significance of DU145 cells is indicated with three asterisks (***). Statistical significance is represented by an * when using the student t -test with a P value of 0.05 compared to cells propagated in complete growth medium.

Article Snippet: The effects of PPV on the expression of VEGF ligand B and receptors 1 and 2 were investigated in MDA-MB-231, A549, and DU145 cells using a VEGF B ELISA kit (Elabscience biotechnology incorporated (Houston, TX, USA)) a VEGF R1 ELISA kit and a VEGF R2 ELISA kit (Abcam plc. (Cambridge, UK).

Techniques: Spectrophotometry, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: In Vitro Effects of Papaverine on Cell Migration and Vascular Endothelial Growth Factor in Cancer Cell Lines

doi: 10.3390/ijms23094654

Figure Lengend Snippet: Spectrophotometry results of VEGF R2 ELISA demonstrating the effects of PPV (10–300 µM) on proliferation on MDA-MB-231 cells compared to A549- and DU145 cell lines at 48 h. The average of three independent experiments is represented by the graph with error bars indicating standard deviation. The statistical significance of MDA-MB-231 cells is indicated with an asterisk (*), the statistical significance of A549 cells is indicated with two asterisks (**), and the statistical significance of DU145 cells is indicated with three asterisks (***). Statistical significance is represented by an * when using the student t -test with a p value of 0.05 compared to cells propagated in complete growth medium.

Article Snippet: The effects of PPV on the expression of VEGF ligand B and receptors 1 and 2 were investigated in MDA-MB-231, A549, and DU145 cells using a VEGF B ELISA kit (Elabscience biotechnology incorporated (Houston, TX, USA)) a VEGF R1 ELISA kit and a VEGF R2 ELISA kit (Abcam plc. (Cambridge, UK).

Techniques: Spectrophotometry, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: In Vitro Effects of Papaverine on Cell Migration and Vascular Endothelial Growth Factor in Cancer Cell Lines

doi: 10.3390/ijms23094654

Figure Lengend Snippet: Spectrophotometry results of FAK ELISA demonstrating the effects of PPV (10–300 µM) on proliferation on MDA-MB-231 cells compared to A549- and DU145 cell lines at 48 h. The average of there independent experiments is represented by the graph with error bars indicating standard deviation. The statistical significance of MDA-MB-231 cells is indicated with an asterisk (*), the statistical significance of A549 cells is indicated with two asterisks (**), and the statistical significance of DU145 cells is indicated with three asterisks (***). Statistical significance is represented by an * when using the student t -test with a p value of 0.05 compared to cells propagated in complete growth medium.

Article Snippet: The effects of PPV on the expression of VEGF ligand B and receptors 1 and 2 were investigated in MDA-MB-231, A549, and DU145 cells using a VEGF B ELISA kit (Elabscience biotechnology incorporated (Houston, TX, USA)) a VEGF R1 ELISA kit and a VEGF R2 ELISA kit (Abcam plc. (Cambridge, UK).

Techniques: Spectrophotometry, Enzyme-linked Immunosorbent Assay, Standard Deviation

Journal: Advanced Science

Article Title: Dipyridamole Acts as Clinical Ferroptosis Inhibitor to Prevent from Tissue Injury

doi: 10.1002/advs.202500566

Figure Lengend Snippet: Dipyridamole inhibits ferroptotic cell death in myocardium cells. a) Cell death measurement of H9c2 cells treated with RSL3 (250 and 500 n m ), dipyridamole (10 µ m ), or Fer‐1 (4 µ m ) for 4 h. Dead cells were labeled with SYTOX Green. b–d) The relative mRNA levels of Ptgs2 (b), Bnp (c), and Myh7 (d) were quantified by qRT‐PCR in H9c2 cells. e) Immunofluorescence staining of BODIPY 581/591 C11 to detect the levels of lipid peroxidation in the H9c2 cells treated with RSL3 (250 n m ), Fer‐1(4 µ m ) and dipyridamole (10 µ m ) for 4 h. f) Dose‐dependent toxicity of RSL3‐induced cell death in AC16 cells after pre‐treatment with dipyridamole (10 µ m ) for 12 h. Cell viability was assessed 12 h post‐treatment after using CCK8. g) Cell death measurement of AC16 cells treated with RSL3 (200 n m ), dipyridamole (10 µ m ), and Fer‐1 (4 µ m ) for 6 h. Dead cells were labeled with SYTOX Green. h,i) The relative mRNA levels of PTGS2 (h) and CHAC1 (i) were quantified by qRT‐PCR in AC16 cells treated with DMSO, RSL3 (200 n m ), dipyridamole (10 µ m ), and Fer‐1 (4 µ m ) for 3 h. j,k) BODIPYTM 581/591 C11 staining of lipid peroxidation in AC16 cells treated with DMSO, RSL3 (200 n m ), Lipro‐1(4 µ m ) or dipyridamole (10 µ m ) for 3 h. l) The relative mRNA level of BNP was quantified by qRT‐PCR in AC16 cells treated with DMSO, RSL3 (400 n m ), dipyridamole (10 µ m ), or Fer‐1 (4 µ m ) for 3 h. m,n) The relative mRNA level of Ptgs2 was quantified by qRT‐PCR in H9c2 cells (m) and NCM (n) pretreated with Lipro‐1 (4 µ m ) or dipyridamole (10 µ m ) for 4 h, followed by hypoxia/reoxygenation (H/R). Data and error bars are mean ± SEM, n = 3 biologically independent experiments in a–d and f‐n. All P values were calculated using a two‐tailed, unpaired Student's t ‐test.

Article Snippet: H9c2 cells were exposed to RSL3, Fer‐1 (4 μ m ) or dipyridamole (10 μ m ) for 6 h. 1 × 10 6 cells per sample were collected and intracellular CK‐MB or BNP levels were determined using the Rat CK‐MB (Creatine Kinase MB Isoenzyme) ELISA Kit (Elabscience, E‐EL‐R1327) and Rat BNP (Brain Natriuretic Peptide) ELISA Kit (E‐EL‐R0126) according to the manufacturer's instructions.

Techniques: Labeling, Quantitative RT-PCR, Immunofluorescence, Staining, Two Tailed Test

Journal: Advanced Science

Article Title: Dipyridamole Acts as Clinical Ferroptosis Inhibitor to Prevent from Tissue Injury

doi: 10.1002/advs.202500566

Figure Lengend Snippet: Dipyridamole‐mediated ferroptosis depends on SLC7A11. a,b) Immunoblot assays of classical ferroptosis‐related genes in HT1080 (a) and H9c2 (b) cells with dipyridamole (20 µ m ) treatment at indicated times. c‐e) Immunoblot assays of SLC7A11 expression in HT1080 (c), OVCAR8 (d), and 786‐O (e) cells with dipyridamole (20 µ m ) treatment at indicated times. f) Immunoblot assays of SLC7A11 expression in HT1080 WT and SLC7A11 KO cells. g) Cell viability assay in HT1080 WT and SLC7A11 KO cells treated with DMSO, dipyridamole (20 µ m ), or RSL3 at the indicated concentration. h) The GSH levels in HT1080 cells treated with RSL3 (250 n m ), Fer‐1(4 µ m ), and dipyridamole (10 µ m ) for 4 h. i) Immunoblot assays of exogenous SLC7A11 expression in HT1080 SLC7A11 KO cells stably expressing with the Mock and SLC7A11‐Flag vector. j) Cell viability assay in HT1080 SLC7A11 KO cells stably expressing with the Mock and SLC7A11‐Flag vector treated with DMSO, RSL3 (5 µ m ), and dipyridamole (10 µ m ) at the indicated concentration. k) The cell death of HT1080 SLC7A11 KO cells stably expressing the Mock and SLC7A11‐Flag vector was measured. These cells were treated with DMSO, RSL3 (5 µ m ), or dipyridamole (10 µ m ) for 6 h. Dead cells were labeled with SYTOX™ Green. Data and error bars are mean ± SEM, n = 3 biologically independent experiments in g, h, j, and k. All P values were calculated using a two‐tailed, unpaired Student's t‐test.

Article Snippet: H9c2 cells were exposed to RSL3, Fer‐1 (4 μ m ) or dipyridamole (10 μ m ) for 6 h. 1 × 10 6 cells per sample were collected and intracellular CK‐MB or BNP levels were determined using the Rat CK‐MB (Creatine Kinase MB Isoenzyme) ELISA Kit (Elabscience, E‐EL‐R1327) and Rat BNP (Brain Natriuretic Peptide) ELISA Kit (E‐EL‐R0126) according to the manufacturer's instructions.

Techniques: Western Blot, Expressing, Viability Assay, Concentration Assay, Stable Transfection, Plasmid Preparation, Labeling, Two Tailed Test

Journal: Discover Oncology

Article Title: RBMS1 interference inhibits malignant progression of glioblastoma cells and promotes ferroptosis

doi: 10.1007/s12672-024-01430-1

Figure Lengend Snippet: RBMS1 silence inhibited the proliferation and promoted the apoptosis of T98G cells. A The transfection efficiency of sh-RBMS1 was examined with RT-qPCR and western blot. B The cell proliferation was detected using CCK-8 assay. C The colony forming ability was detected using colony formation assay. D The cell apoptosis was detected using flow cytometry. E The expression of proliferation- and apoptosis-related proteins were detected using western blot. ** P < 0.01 and *** P < 0.001 vs. sh-NC

Article Snippet: For the detection of lipid peroxidation in T98G cells, TBARS Assay Kits (cat. no. E-BC-K298-M; Elabscience) were performed according to the manufacturer’s instructions.

Techniques: Transfection, Quantitative RT-PCR, Western Blot, CCK-8 Assay, Colony Assay, Flow Cytometry, Expressing

Journal: Discover Oncology

Article Title: RBMS1 interference inhibits malignant progression of glioblastoma cells and promotes ferroptosis

doi: 10.1007/s12672-024-01430-1

Figure Lengend Snippet: RBMS1 silence inhibited the migration, invasion and EMT process in T98G cells. A , B The cell migration and invasion were detected using wound healing and transwell assay. C The activities of MMP2 and MMP9 were detected using western blot. D The expressions of EMT-related proteins were detected using western blot. ** P < 0.01 and *** P < 0.001 vs. sh-NC

Article Snippet: For the detection of lipid peroxidation in T98G cells, TBARS Assay Kits (cat. no. E-BC-K298-M; Elabscience) were performed according to the manufacturer’s instructions.

Techniques: Migration, Transwell Assay, Western Blot

Journal: Discover Oncology

Article Title: RBMS1 interference inhibits malignant progression of glioblastoma cells and promotes ferroptosis

doi: 10.1007/s12672-024-01430-1

Figure Lengend Snippet: RBMS1 silence promoted ferroptosis in T98G cells. A The lipid peroxidation was detected using TBARS Assay Kit. B The lipid ROS was detected using a BODIPY 581/591 C11 kit. C The total iron level was detected using corresponding assay. (D) The expressions of ferroptosis-related proteins were detected using western blot. *** P < 0.001 vs. sh-NC

Article Snippet: For the detection of lipid peroxidation in T98G cells, TBARS Assay Kits (cat. no. E-BC-K298-M; Elabscience) were performed according to the manufacturer’s instructions.

Techniques: TBARS Assay, Western Blot

Journal: Discover Oncology

Article Title: RBMS1 interference inhibits malignant progression of glioblastoma cells and promotes ferroptosis

doi: 10.1007/s12672-024-01430-1

Figure Lengend Snippet: RBMS1 silence inhibited the proliferation and promoted the apoptosis of T98G cells by promoting ferroptosis. A The cell proliferation was detected using CCK-8 assay. B The colony forming ability was detected using colony formation assay. C The cell apoptosis was detected using flow cytometry. D The expression of proliferation- and apoptosis-related proteins were detected using western blot. ** P < 0.01 and *** P < 0.001 vs. sh-NC, ## P < 0.01 and ### P < 0.001 vs. sh-RBMS1

Article Snippet: For the detection of lipid peroxidation in T98G cells, TBARS Assay Kits (cat. no. E-BC-K298-M; Elabscience) were performed according to the manufacturer’s instructions.

Techniques: CCK-8 Assay, Colony Assay, Flow Cytometry, Expressing, Western Blot

Journal: Discover Oncology

Article Title: RBMS1 interference inhibits malignant progression of glioblastoma cells and promotes ferroptosis

doi: 10.1007/s12672-024-01430-1

Figure Lengend Snippet: RBMS1 silence inhibited the migration, invasion and EMT process in T98G cells by promoting ferroptosis. A , B The cell migration and invasion were detected using wound healing and transwell assay. C The activities of MMP2 and MMP9 were detected using western blot. D The expressions of EMT-related proteins were detected using western blot. *** P < 0.001 vs. sh-NC, # P < 0.05, ## P < 0.01 and ### P < 0.001 vs. sh-RBMS1

Article Snippet: For the detection of lipid peroxidation in T98G cells, TBARS Assay Kits (cat. no. E-BC-K298-M; Elabscience) were performed according to the manufacturer’s instructions.

Techniques: Migration, Transwell Assay, Western Blot