Journal: PLoS ONE

Article Title: Bcl-w Enhances Mesenchymal Changes and Invasiveness of Glioblastoma Cells by Inducing Nuclear Accumulation of β-Catenin

doi: 10.1371/journal.pone.0068030

Figure Lengend Snippet: A , premade brain cancer tissue microarrays (AccuMax, A221-iv) from patients were subjected to immunohistochemical staining with anti-Bcl-w antibody (R & D systems, Minneapolis, MN). Non-neoplastic and corresponding glial tumor grade iii/iv tissues from patients (P1-P12). Bar scale, 100 µm. B , U251 cells were transfected with either empty pcDNA vector or that containing Bcl-w cDNA. Bcl-w expression was detected using Western blotting with β-actin as the loading control. Expression levels of mesenchymal proteins were analyzed using Western blot analysis with anti-Twist1, anti-Snail, anti-Slug, anti-vimentin, anti-E-cadherin and anti-Bcl-w in control vector and Bcl-w-overexpressing cells. C , U251 cells were transfected with control or siRNA oligonucleotides targeting Bcl-w (20nM) for 24 hours. Transfected control or si-Bcl-w U251 cells were subjected to Western blot analysis with the indicated antibodies. D , confocal microscopy analysis of vector or Bcl-w-overexpressing cells showing Bcl-w (Red, Alexa 568) and vimentin (Green, Alexa 488) and DAPI (Blue). Scale bar, 50 µm.

Article Snippet: The U251, U373, U87MG (glioma), MDA-MB-231 (breast cancer) and H1299 (lung cancer) were obtained from the Korean Cell Line Bank (KCLB).

Techniques: Immunohistochemical staining, Staining, Transfection, Plasmid Preparation, Expressing, Western Blot, Control, Confocal Microscopy

Journal: PLoS ONE

Article Title: Bcl-w Enhances Mesenchymal Changes and Invasiveness of Glioblastoma Cells by Inducing Nuclear Accumulation of β-Catenin

doi: 10.1371/journal.pone.0068030

Figure Lengend Snippet: Down-regulation of Twist1 and Snail leads to inhibition of glioma invasion and vimentin expression. A , the indicated U251 cell transfectants were incubated in serum-free medium in the presence or absence of PI3K inhibitor (LY294002 (LY); 10 µmol/L) and Akt inhibitor (Akt-I; 10 µmol/L) for 1 hour. Expression levels of Twist1, snail and vimentin proteins were compared using Western blotting. B , control or Bcl-w-expressing U251 cells were transfected with 20nM of Twist1, Snail or vimentin siRNA for 24 hours and were subjected to Western blotting with mesenchymal-related proteins or anti-Bcl-w antibodies. C , cells in incubated in a Matrigel-coated transwell for 20 hours. *, p < 0.05, **, p < 0.005, n = 5.

Article Snippet: The U251, U373, U87MG (glioma), MDA-MB-231 (breast cancer) and H1299 (lung cancer) were obtained from the Korean Cell Line Bank (KCLB).

Techniques: Inhibition, Expressing, Incubation, Western Blot, Control, Transfection

Journal: PLoS ONE

Article Title: Bcl-w Enhances Mesenchymal Changes and Invasiveness of Glioblastoma Cells by Inducing Nuclear Accumulation of β-Catenin

doi: 10.1371/journal.pone.0068030

Figure Lengend Snippet: A , either empty pcDNA vector or that containing Bcl-w cDNA introduced into U251 cells. Bcl-w expression was detected using Western blotting. B , Bcl-w promotes migration and invasiveness of U251 glioblastoma cells. Top, the confluent cells in five fields from the scratched area (200 x 500 µm 2 ) were counted under a light microscope. Transfectants were seeded onto Matrigel-coated polycarbonate filters to analyze their invasive potential. Cells were incubated for 20 hours in modified Boyden chambers, and the number of cells invading through filters stained and counted under a light microscope. Bottom, mean of triplicate experiments significantly different from controls. *, p< 0.05. C , two different siRNA sequences targeting Bcl-w (20nM of si-Bcl-w-1 and si-Bcl-w-2) were introduced into U251 cells for 24 hours, and the invasion assay conducted after 24 hours of incubation. Experiments were repeated five times, and the mean values and standard deviations determined. *, p< 0.05; **, p < 0.005.

Article Snippet: The U251, U373, U87MG (glioma), MDA-MB-231 (breast cancer) and H1299 (lung cancer) were obtained from the Korean Cell Line Bank (KCLB).

Techniques: Plasmid Preparation, Expressing, Western Blot, Migration, Light Microscopy, Incubation, Modification, Staining, Invasion Assay

Journal: PLoS ONE

Article Title: Bcl-w Enhances Mesenchymal Changes and Invasiveness of Glioblastoma Cells by Inducing Nuclear Accumulation of β-Catenin

doi: 10.1371/journal.pone.0068030

Figure Lengend Snippet: A , levels of p-Akt, p-GSK3β, GSK3β, p-β-catenin, β-catenin and TCF-4 in cell lysates were compared by Western blotting using β-actin as a loading control. Conditioned media were prepared by incubating the vector and Bcl-w transfectants in serum-free medium for 24 hours. MMP-2 and MMP-9 activities were compared using zymography. Protein loading volumes were verified with Ponceau S staining. B , levels of β-catenin protein that translocated into the nucleus and Bcl-w protein in vector- or Bcl-w-transfected U251 cells were examined using confocal microscopy. Cells were stained with anti-β-catenin (green) or anti-Bcl-w (red) antibody, followed by nuclear staining with DAPI (blue). Scale bar, 50 µm. C , after separation of cells into cytoplasm and nuclear fractions for the indicated transfectants, each fraction was subjected to Western blotting with anti-β-catenin, anti-Lamin A/C (nucleus marker) and anti-β-actin (cytoplasm marker) antibodies.

Article Snippet: The U251, U373, U87MG (glioma), MDA-MB-231 (breast cancer) and H1299 (lung cancer) were obtained from the Korean Cell Line Bank (KCLB).

Techniques: Western Blot, Control, Plasmid Preparation, Zymography, Staining, Transfection, Confocal Microscopy, Marker

Journal: PLoS ONE

Article Title: Bcl-w Enhances Mesenchymal Changes and Invasiveness of Glioblastoma Cells by Inducing Nuclear Accumulation of β-Catenin

doi: 10.1371/journal.pone.0068030

Figure Lengend Snippet: A , the indicated U251 cell transfectants were incubated in serum-free medium in the presence or absence of PI3K inhibitor (LY294002 (LY); 10 µmol/L) or Akt inhibitor (Akt-I; 10 µmol/L) for 1 hour. Expression levels and activities of p-Akt, p-GSK3β, β-catenin, TCF-4 and MMP-2 proteins were compared using Western blotting. B , cells treated with PI3K inhibitor or Akt inhibitor in the lower compartments of the invasion chambers for 24 hours, respectively. Invasive potential of treated cells was compared. *, p< 0.05 versus untreated control, n = 5. C , β-catenin and TCF-4 siRNAs (20nM) were introduced into vector or Bcl-w overexpressing cells, and cellular levels of β-catenin, TCF-4, MMP-2 and p-FAK compared after 24 hours of incubation using Western blotting with β-actin as a loading control. D , invasive potential of the indicated transfectants was compared. *, p< 0.05, n = 5.

Article Snippet: The U251, U373, U87MG (glioma), MDA-MB-231 (breast cancer) and H1299 (lung cancer) were obtained from the Korean Cell Line Bank (KCLB).

Techniques: Incubation, Expressing, Western Blot, Control, Plasmid Preparation

Journal: PLoS ONE

Article Title: Bcl-w Enhances Mesenchymal Changes and Invasiveness of Glioblastoma Cells by Inducing Nuclear Accumulation of β-Catenin

doi: 10.1371/journal.pone.0068030

Figure Lengend Snippet: A , right image, MMP-2 siRNA (20nM) was introduced into the indicated U251 transfectants, and after 24 hours of incubation, p-FAK (Y397) and MMP-2 protein levels were compared using Western blotting. Left image, invasion assays were performed using small interfering RNA MMP-2-treated and untreated cells. *, p< 0.01 versus untreated control, n = 5. B , top image, FAK siRNA was introduced into the indicated transfectants, and after 24 hours of incubation, cellular levels of FAK, p-FAK and MMP-2 compared using Western blotting. Bottom plots, invasion assays were conducted using the indicated cells. *, p< 0.05, n = 5. C , top images, vector- and Bcl-w-expressing cells were transiently transfected with expression vectors for HA-tagged dominant-negative FAK mutant (FAKY397F). After 24 hours of incubation, expression of the introduced mutants in cells was verified by Western blotting. Bottom plots, invasive potentials of the indicated cells were compared. *, p < 0.05, n = 5.

Article Snippet: The U251, U373, U87MG (glioma), MDA-MB-231 (breast cancer) and H1299 (lung cancer) were obtained from the Korean Cell Line Bank (KCLB).

Techniques: Incubation, Western Blot, Small Interfering RNA, Control, Plasmid Preparation, Expressing, Transfection, Dominant Negative Mutation, Mutagenesis

Journal: American Journal of Translational Research

Article Title: MicroRNA-365 suppressed cell proliferation and migration via targeting PAX6 in glioblastoma

doi:

Figure Lengend Snippet: Serum expression level of miR-365 was downregulated in the glioblastoma. The serum expression of miR-365 in the glioblastoma and healthy controls was detected using qRT-PCR. ***P < 0.001. Statistical analysis was performed using Mann-Whitney U tests.

Article Snippet: Four glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) were collected from the American Tissue Culture Colection (ATCC) and was cultured in DMEM medium. miR-365 mimics and its scramble oligonucleotides were obtained from RiboBio (Guangzhou, China).

Techniques: Expressing, Quantitative RT-PCR, MANN-WHITNEY

Journal: American Journal of Translational Research

Article Title: MicroRNA-365 suppressed cell proliferation and migration via targeting PAX6 in glioblastoma

doi:

Figure Lengend Snippet: Expressionl level of miR-365 was decreased in the glioblastoma tissue. A. The expression of miR-365 was downregulated in the glioblastoma tissue compared to the adjacent normal tissues. Statistically significant difference was determined using Student’s t test. B. The miR-365 expression in 31 cases (31/40; 77%) compared to adjacent tissues. C. The expression of miR-365 in glioblastoma tissue was associated with those in glioblastoma patients’ serum. ***P < 0.001. Association between miR-365 expression in serum and matched glioma tissues was detected by Spearman correlation test.

Article Snippet: Four glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) were collected from the American Tissue Culture Colection (ATCC) and was cultured in DMEM medium. miR-365 mimics and its scramble oligonucleotides were obtained from RiboBio (Guangzhou, China).

Techniques: Expressing

Journal: American Journal of Translational Research

Article Title: MicroRNA-365 suppressed cell proliferation and migration via targeting PAX6 in glioblastoma

doi:

Figure Lengend Snippet: miR-365 suppressed the glioblastoma cell proliferation and migration. A. The expression of miR-365 in the glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) was detecte using qRT-PCR. B. The expression of miR-365 was measured by qRT-PCR in the U87 cell. C. Overexpression of miR-365 suppressed the U87 cell proliferation. D. Overexpression of miR-365 suppressed the U87 cell migration. E. The relative open wound was shown. *P < 0.05, **P < 0.01 and ***P < 0.001. Statistically significant difference was determined using Student’s t test.

Article Snippet: Four glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) were collected from the American Tissue Culture Colection (ATCC) and was cultured in DMEM medium. miR-365 mimics and its scramble oligonucleotides were obtained from RiboBio (Guangzhou, China).

Techniques: Migration, Expressing, Quantitative RT-PCR, Over Expression

Journal: American Journal of Translational Research

Article Title: MicroRNA-365 suppressed cell proliferation and migration via targeting PAX6 in glioblastoma

doi:

Figure Lengend Snippet: miR-365 inhibited the glioblastoma cell epithelial-to-mesenchymal transition. A. The mRNA expression of Ecadherin, N-cadherin and Vimentin was measured by qRT-PCR. B. The protein expression of Ecadherin, N-cadherin and Vimentin was detected by western blot.

Article Snippet: Four glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) were collected from the American Tissue Culture Colection (ATCC) and was cultured in DMEM medium. miR-365 mimics and its scramble oligonucleotides were obtained from RiboBio (Guangzhou, China).

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: American Journal of Translational Research

Article Title: MicroRNA-365 suppressed cell proliferation and migration via targeting PAX6 in glioblastoma

doi:

Figure Lengend Snippet: PAX6 was a direct a target gene of miR-365 in the glioblastoma cell. A. One putative binding sites of the miR-365 in the 3’UTR of PAX6 are shown. B. The expression of PAX6 in the glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) was measured using qRT-PCR. C. The luciferase activity was decreased in the U87 cell co-transfected with miR-365 mimic and PAX6-3’UTR-WT luciferase reporter. D. Ecoptic expression of miR-365 suppressed the PAX6 mRNA expression. E. Overexpression of miR-365 inhibited the protein pression of PAX6.

Article Snippet: Four glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) were collected from the American Tissue Culture Colection (ATCC) and was cultured in DMEM medium. miR-365 mimics and its scramble oligonucleotides were obtained from RiboBio (Guangzhou, China).

Techniques: Binding Assay, Expressing, Quantitative RT-PCR, Luciferase, Activity Assay, Transfection, Over Expression

Journal: American Journal of Translational Research

Article Title: MicroRNA-365 suppressed cell proliferation and migration via targeting PAX6 in glioblastoma

doi:

Figure Lengend Snippet: miR-365 suppressed glioblastoma cell proliferation and migration by inhibiting PAX6. A. The mRNA expression of PAX6 was determined by using qRT-PCR. B. The protein expression of PAX6 was detected by using western blot. C. PAX6 overexpression promoted the miR-365-overexpressing U87 cell proliferation. D. PAX6 Overexpression abrogated the reduction of migration ability caused by ectopic expression of miR-365 in U87 cells. The relative migrative wound was shown. E. The mRNA expression of E-cadherin, N-cadherin and vimentin was determined by qRT-PCR. F. The protein expression of E-cadherin, N-cadherin and vimentin was determined by western blot. *P < 0.05, **P < 0.01 and ***P < 0.001. Statistically significant difference was determined using Student’s t test.

Article Snippet: Four glioblastoma cell lines (U373, U87, A172 and U251) and one normal astrocyte line (NHAs) were collected from the American Tissue Culture Colection (ATCC) and was cultured in DMEM medium. miR-365 mimics and its scramble oligonucleotides were obtained from RiboBio (Guangzhou, China).

Techniques: Migration, Expressing, Quantitative RT-PCR, Western Blot, Over Expression

Journal: Cell Death & Disease

Article Title: The CDK inhibitor AT7519 inhibits human glioblastoma cell growth by inducing apoptosis, pyroptosis and cell cycle arrest

doi: 10.1038/s41419-022-05528-8

Figure Lengend Snippet: A Process for high-throughput drug screening. B , C U87MG, U251, GBM60, and GBM38 cell viability was determined using a CCK-8 assay after treatment with various concentrations of AT7519. * P < 0.05, *** P < 0.01, *** P < 0.001, and **** P < 0.0001 compared with the control using one-way ANOVA followed by Dunnett’s multiple test. D U87MG and U251 cell viability was determined using a CCK-8 assay after treatment with 0.4 µM AT7519 for 6, 12, 24, 48, and 60 h. * P < 0.05, **** P < 0.0001 compared with the control using one-way ANOVA followed by Dunnett’s multiple test. E Colony formation assays to verify the effect of AT7519 on glioblastoma cell proliferation. * P < 0.05, *** P < 0.001, and **** P < 0.0001 based on one-way ANOVA followed by Dunnett’s multiple test. F , G The level of DNA synthesis of U87MG and U251 cells was determined using an EdU assay after treatment with increasing concentrations of AT7519. The nuclei were stained with Hoechst (blue), and the proliferating cells were stained with EdU (yellow). * P < 0.05, *** P < 0.001 and **** P < 0.0001 based on one-way ANOVA followed by Dunnett’s multiple test. The results are presented as the mean ± SD from three independent experiments.

Article Snippet: Human glioblastoma cell lines (U87MG and U251) were purchased from ATCC.

Techniques: High Throughput Screening Assay, Drug discovery, CCK-8 Assay, Control, DNA Synthesis, EdU Assay, Staining

Journal: Cell Death & Disease

Article Title: The CDK inhibitor AT7519 inhibits human glioblastoma cell growth by inducing apoptosis, pyroptosis and cell cycle arrest

doi: 10.1038/s41419-022-05528-8

Figure Lengend Snippet: A U87MG and U251 cells were treated with AT7519 and morphological features of pyroptosis in SEM (red arrows, membrane pore-forming). B After U87MG and U251 cells were treated with AT7519 for 48 h, cytotoxicity was detected by lactate dehydrogenase (LDH) released into the cell culture medium. **** P < 0.0001 by Student’s t -test. C , D Full-length GSDME (GSDME-FL) and N-terminal GSDME (GSDME-N) were detected in glioblastoma cells after treatment with AT7519 for 48 h by western blot analysis. E , F U87MG and U251 cells were pretreated with Z-VAD-FMK for 2 h and then treated with AT7519 for 48 h. Cytotoxicity was detected by LDH release assay. *** P < 0.001 and **** P < 0.0001 as assessed by Student’s t -test. The apoptosis marker cleaved PARP and pyroptosis marker GSDME-N were detected by western blot. G U87MG and U251 cells were treated with Z-DEVD-FMK combined with AT7519 for 48 h, and western blot analysis of cleaved caspase-3, GSDME-FL and GSDME-N proteins was performed.

Article Snippet: Human glioblastoma cell lines (U87MG and U251) were purchased from ATCC.

Techniques: Membrane, Cell Culture, Western Blot, Lactate Dehydrogenase Assay, Marker

Journal: Cell Death & Disease

Article Title: The CDK inhibitor AT7519 inhibits human glioblastoma cell growth by inducing apoptosis, pyroptosis and cell cycle arrest

doi: 10.1038/s41419-022-05528-8

Figure Lengend Snippet: A Image of the subcutaneous xenograft tumors formed in nude mouse models. B Tumor volumes were measured and calculated every week. **** P < 0.0001 as assessed by two-way ANOVA followed by Sidak’s multiple test. C Tumors were excised and weighed at the end of the experiment. *** P < 0.001 by Student’s t -test. D Body weight of nude mice during administration of AT7519. E Representative H&E-stained images of the intracranial xenograft model in the AT7519 treatment group and control group. * p < 0.05 as assessed by Student’s t -test. F Western blot assay of the apoptosis, pyroptosis, and cell cycle-related key protein expression levels in tumor tissue. G Schematic model of the antitumor mechanism of AT7519 in glioblastoma cells.

Article Snippet: Human glioblastoma cell lines (U87MG and U251) were purchased from ATCC.

Techniques: Staining, Control, Western Blot, Expressing

Journal: Scientific Reports

Article Title: A specific dispiropiperazine derivative that arrests cell cycle, induces apoptosis, necrosis and DNA damage

doi: 10.1038/s41598-023-35927-6

Figure Lengend Snippet: IC 50 of SPOPP-3 ( 1 ), SPOPP-5 ( 2 ) and doxorubicin against human cancer cell lines.

Article Snippet: For example, SPOPP-3 has stronger anti-proliferative effect on the faster growing human leukemia cell lines (K562, KG1a, CEM) and glioblastoma cell lines (U251, U87) but has weaker effect on the slower growing HepG2 liver cancer cell line, DU145 prostate cancer cell line, and pancreatic cancer cell lines (MiaPaca2, Panc1) (Table ).

Techniques: