Journal: Oncology Letters

Article Title: Inhibition of δ-opioid receptors induces brain glioma cell apoptosis through the mitochondrial and protein kinase C pathways

doi: 10.3892/ol.2013.1546

Figure Lengend Snippet: Naltrindole inhibits the growth of brain glioma cells. Cell reproductivity was tested using the MTT method following the administration of various concentrations of naltrindole (0, 0.01, 0.1, 1.0 and 10 μM), which acted on the U87 cells for 24, 48 and 72 h. The results are representative of three independent experiments. OD, optical density.

Article Snippet: Human brain glioma U87 cells were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques:

Journal: Oncology Letters

Article Title: Inhibition of δ-opioid receptors induces brain glioma cell apoptosis through the mitochondrial and protein kinase C pathways

doi: 10.3892/ol.2013.1546

Figure Lengend Snippet: Naltrindole induces brain glioma cell apoptosis. U87 cell apoptosis induced by various concentrations of naltrindole for 48 h was identified using (A) Hoechst 33342 nuclear staining and analyzed by (B) annexin V-FITC/PI double-stained flow cytometry. (C) The histogram shows the U87 apoptosis rate. * P<0.05 vs. 0 μM. The data are representative of three independent experiments. FITC, fluorescein isothiocyanate; PI, propidium iodide.

Article Snippet: Human brain glioma U87 cells were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Staining, Flow Cytometry

Journal: Oncology Letters

Article Title: Inhibition of δ-opioid receptors induces brain glioma cell apoptosis through the mitochondrial and protein kinase C pathways

doi: 10.3892/ol.2013.1546

Figure Lengend Snippet: Naltrindole induction of brain glioma cell apoptosis through the mitochondrial pathway. The U87 cells were treated with various doses of naltrindole (0, 0.1 and 1.0 μM) for 48 h. (A) Flow cytometry was used to analyze the change in the mitochondrial membrane potential. (B) A western blot analysis was used to analyze the protein expression levels of Bax and cytochrome c . (C) A western blot analysis was used to analyze the protein expression levels of Bcl-2, Bcl-xL and Bak. (D and E) The histogram shows the results from B and C (%). * P<0.05. Results are representative of three independent experiments.

Article Snippet: Human brain glioma U87 cells were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Flow Cytometry, Membrane, Western Blot, Expressing

Journal: Oncology Letters

Article Title: Inhibition of δ-opioid receptors induces brain glioma cell apoptosis through the mitochondrial and protein kinase C pathways

doi: 10.3892/ol.2013.1546

Figure Lengend Snippet: Impact of naltrindole on the brain glioma cell cycle. Following the administration of various doses of naltrindole (0, 0.1 and 1.0 μM) to the U87 cells for 48 h, flow cytometry was used to analyze the cell cycle. * P<0.05 vs. 0 μM. The data are representative of three independent experiments.

Article Snippet: Human brain glioma U87 cells were purchased from the American Type Culture Collection (ATCC; Manassas, VA, USA).

Techniques: Flow Cytometry

Journal: RSC Advances

Article Title: Cell chirality exhibition of brain microvascular endothelial cells is dependent on micropattern width †

doi: 10.1039/d2ra05434e

Figure Lengend Snippet: Cell chirality varies depending on cell type and pathological state

Article Snippet: Mouse brain microvascular endothelial cells (bEnd.3 ATCC) were cultured in Dubelcco’s modified eagle medium high glucose (Gibco) supplemented with 10% foetal bovine serum (Cytiva) and 1% penicillin streptomycin (Lonza) and maintained at 37 °C and 5% CO 2 .

Techniques:

Journal: RSC Advances

Article Title: Cell chirality exhibition of brain microvascular endothelial cells is dependent on micropattern width †

doi: 10.1039/d2ra05434e

Figure Lengend Snippet: Cell and nucleus chirality of mouse brain microvascular endothelial cells on different line widths. (A) Left: bar plots with the percentage of cells and nuclei exhibiting a chirality bias (negative chirality bias in light blue and positive chirality bias in dark blue). Statistical analysis of cell and nucleus orientation of brain microvascular cells on different line widths compared to cells on a non-patterned surface was assessed using a chi-square test. Significance is symbolized by non-significant or N.S. ( p > 0.05), * ( p ≤ 0.05), ** ( p value ≤ 0.01), *** ( p value ≤ 0.001). Right: schematic illustration depicting a positively oriented cell and a negatively oriented nucleus compared to the green line. (B) Left: bar plots of the mean orientation of the cells and nuclei on the different line widths. Statistical analysis of cell and nucleus chirality of brain microvascular endothelial cells on different line widths and cells seeded on non-patterned hydrogels was determined using a one way analysis of variance (ANOVA) with a Games-Howell post hoc for multiple comparisons at a 95% confidence level. Significance is symbolized by non-significant or N.S. ( p > 0.05), * ( p ≤ 0.05), ** ( p value ≤ 0.01), *** ( p value ≤ 0.001). Right: confocal maximum intensity projection of cells on the 100 μm wide line (where the nuclei are shown in blue) illustrating the angular orientation of the different nuclei, where one negatively and one positively oriented nucleus is enlarged. To obtain the data, three hydrogel samples were utilized per line width, where four images were acquired and analysed per hydrogel sample, which equates to 12 images per line width in total. The number of cells analysed for each line width varied between 400 for the 10 μm wide lines and 1400 for the 400 μm wide lines. Scale bars = 100 μm. Created with https://BioRender.com .

Article Snippet: Mouse brain microvascular endothelial cells (bEnd.3 ATCC) were cultured in Dubelcco’s modified eagle medium high glucose (Gibco) supplemented with 10% foetal bovine serum (Cytiva) and 1% penicillin streptomycin (Lonza) and maintained at 37 °C and 5% CO 2 .

Techniques:

Journal: RSC Advances

Article Title: Cell chirality exhibition of brain microvascular endothelial cells is dependent on micropattern width †

doi: 10.1039/d2ra05434e

Figure Lengend Snippet: Brain microvascular endothelial cell chirality propagation from the boundary. (A) Representative image of cells, stained for ZO-1 (yellow) and nucleus (blue) (left), and F-actin (red) (right), on the 100 μm wide line patterns (green), divided into five equal regions (1–5). The white dashed lines indicate the five resulting regions, each 20 μm wide. These regions (1–5) in the schematic corresponds to the regions in B–D. (B) Bar plots with the percentage of cells, nuclei, and F-actin fibres exhibiting a chirality bias (negative chirality bias in light blue and positive chirality bias in dark blue) within each of the five regions (1–5) depicted in (A). Statistical analysis of cell, nucleus, and actin orientation of brain microvascular cells in the different regions were assessed using a chi-square test. Significance is symbolized by non-significant or N.S. ( p > 0.05), * ( p ≤ 0.05), ** ( p value ≤ 0.01), *** ( p value ≤ 0.001). (C) Polar plots indicating the orientation of the cells, nuclei, and F-actin fibres of cells in the five regions (1–5) as depicted in (A), and the non-patterned (NP) area, included as a comparison. (D) Bar plots showing the mean orientation of the cells, nuclei, and F-actin fibres of the different regions (1–5) depicted in (A). Statistical analysis of cell, nucleus, and actin chirality of brain microvascular endothelial cells in the different regions (1–5) were determined using a one way analysis of variance (ANOVA) with a Games-Howell post hoc for multiple comparisons at a 95% confidence level. Significance is symbolized by non-significant or N.S. ( p > 0.05), * ( p ≤ 0.05), ** ( p value ≤ 0.01), *** ( p value ≤ 0.001). To obtain the data, three hydrogel samples were utilized per line width, where four images were acquired and analysed per hydrogel sample, which equates to 12 images in total. The total number of cells analysed was around 900. Scale bars = 100 μm. Created with https://BioRender.com .

Article Snippet: Mouse brain microvascular endothelial cells (bEnd.3 ATCC) were cultured in Dubelcco’s modified eagle medium high glucose (Gibco) supplemented with 10% foetal bovine serum (Cytiva) and 1% penicillin streptomycin (Lonza) and maintained at 37 °C and 5% CO 2 .

Techniques: Staining, Comparison

Journal: RSC Advances

Article Title: Cell chirality exhibition of brain microvascular endothelial cells is dependent on micropattern width †

doi: 10.1039/d2ra05434e

Figure Lengend Snippet: ZO-1 expression depending on brain microvascular endothelial cell location. (A) Representative image of cells, stained for ZO-1 (yellow) and nucleus (blue), on the 100 μm wide line patterns, divided into five equal regions (1–5). The white dashed lines indicate the five resulting regions, each 20 μm wide. These regions (1–5) in the schematic corresponds to the regions in (B). (B) Bar plot of the mean intensity units of ZO-1 per cell within each of the five regions (1–5) depicted in (A). The statistical significance of the ZO-1 mean intensity in the different regions was determined by performing a one way analysis of variance (ANOVA) with a Games-Howell post hoc for multiple comparisons at a 95% confidence level. The statistical significance is symbolized by * ( p ≤ 0.05), ** ( p value ≤ 0.01), *** ( p value ≤ 0.001). To obtain the data, three hydrogel samples were utilized per line width, where four images were acquired and analysed per hydrogel sample, which equates to 12 images per line width in total. The total number of cells analysed was around 900. Scale bars = 100 μm.

Article Snippet: Mouse brain microvascular endothelial cells (bEnd.3 ATCC) were cultured in Dubelcco’s modified eagle medium high glucose (Gibco) supplemented with 10% foetal bovine serum (Cytiva) and 1% penicillin streptomycin (Lonza) and maintained at 37 °C and 5% CO 2 .

Techniques: Expressing, Staining

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: Response of glioblastoma cell lines to chemotherapy drug temozolomide (TMZ). We performed 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays on T98G, U138MG, A172, GBM8401 and U87MG cell lines after 72 h of nontreatment or 200 or 400 µM of TMZ treatment. Values are expressed as mean ± standard deviation (SD; n = 4). * p < 0.05 relative to nontreatment.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Standard Deviation

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: Glutathione S-transferase (GST) activity and GST subfamily member protein expression in glioblastoma cell lines. ( A ) The reaction mixture contained 100 µg of cytosolic protein of each glioblastoma multiforme (GBM) cell line. Values are expressed as mean ± SD ( n = 3). * p < 0.05 relative to T98G cells. ( B ) Western blot analysis of GSTM1, GSTM2, GSTM3, GSTM4, GSTM5, GSTP1, and GSTK1 of human fetal glial cell line SVGp12 and human glioblastoma cell lines U87MG, T98G, U138MG, A172, and GBM8401. β-Actin was used as the loading control.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Activity Assay, Expressing, Western Blot, Control

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: Bioenergetic profiles of five well-known GBM cell lines. ( a ) Basal extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) levels provide a snapshot of the bioenergetics profile of well-known GBM cell lines U87MG, T98G, U138MG, A172, and GBM8401. Values were measured and normalized to cell numbers by using the Seahorse Extracellular Flux (XF) Analyzer. ( b ) Real-time measurement of the ECAR as the glycolysis rate. Glucose (10 mM), oligomycin (1 µM) and 2-DG (50 mM) were injected at the indicated times. Values are expressed as mean ± SD ( n = 3). ( c ) The glycolytic rate, glycolytic capacity, and glycolytic reserve were analyzed using standard methods. Values are expressed as the mean ± SD ( n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 relative to T98G cells.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Injection

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: Knockdown of GSTM3 T98G cells. ( a ) Protein expression of GSTM3 and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in siNC and siGSTM3 T98G cells. β-Actin was used as the loading control. ( b ) Morphology of small interfering RNA (siRNA) negative control (siNC) and GSTM3 (siGSTM3) T98G glioblastoma. Cells were subjected to microscopy at 40× magnification. ( c ) GST activity of siNC and GSTM3 T98G cells. The reaction mixture contained 100 µg of cytosolic protein in 0.1 M KPO4 buffer, pH 7.5, containing 1 mM GSH and 1 mM CDNB at 25 °C. Values are expressed as mean ± SD ( n = 3). ** p < 0.001 relative to siNC. ( d ) MTT assays were performed on Mock, siNC, and GSTM3 T98G cells. The MTT assay of T98G cells after 72 h of TMZ treatment. Values are expressed as mean ± SD ( n = 6). * p < 0.05 relative to siNC. # p < 0.05 relative to mock cells.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Knockdown, Expressing, Control, Small Interfering RNA, Negative Control, Microscopy, Activity Assay, MTT Assay

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: OCR and ECAR values from a glycolysis stress test in GSTM3-knockdown T98G cells. T98G cells treated with siRNA knockdown were depleted 72 h after transfection. The ECAR was measured and normalized to cell number by using the Seahorse XF Analyzer in cells. ( a ) Real-time measurement of the OCR as mitochondrial respiration. Oligomycin (1 µM), FCCP (0.5 µM), and rotenone/antimycin A (0.5 µM) were injected at the indicated times. Values are expressed as mean ± SD ( n = 3). ( b ) Real-time measurement of the ECAR as the glycolysis rate. Glucose (10 mM), oligomycin (1 µM), and 2-DG (50 mM) were injected at the indicated times. Values are expressed as mean ± SD ( n = 3). ( c ) The glycolytic rate, glycolytic capacity, and glycolytic reserve were analyzed using standard methods. Values are expressed as the mean ± SD ( n = 3). *** p < 0.001 relative to siNC.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Knockdown, Transfection, Injection

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: Knockdown of siGSTM3 affects LDH activity in T98G glioblastoma cells. T98G cells treated with siRNA knockdown were depleted 72 h after transfection. ( a ) The lactate dehydrogenase A (LDHA) mRNA expression level was examined using reverse-transcription polymerase chain reaction. GAPDH was used as an internal control. Values are expressed as the mean ± SD ( n = 3). * p < 0.05 relative to siNC. ( b ) The LDH activities of siNC and siGSTM3 T98G cells were measured using a Pierce LDH Cytotoxicity Assay Kit (Thermo Scientific). Endogenous proteins were extracted from each cell population. Each measurement was obtained for 1 × 10 5 cells. Values are expressed as the mean ± SD ( n = 3). * p < 0.05 relative to siNC. ( c ) The L-lactate content of siNC and siGSTM3 T98G cells was measured using an L-Lactate Assay Kit. Each measurement was obtained for 100 µg of lysate. Values are expressed as the mean ± SD ( n = 3). ** p < 0.01 relative to siNC.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Knockdown, Activity Assay, Transfection, Expressing, Reverse Transcription, Polymerase Chain Reaction, Control, LDH Cytotoxicity Assay, Lactate Assay

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: GSTM3 downregulation decreases the invasion ability of T98G glioblastoma cells. ( a ) Cell migration was measured using a collagen-I-coated Transwell chamber (8-µm pore) for evaluating the migration of siNC and GSTM3 siRNA-treated cells. Migrated cells were stained with Giemsa solution (magnification 200×). ( b ) Cells on the underside of the Transwell insert were counted per file. Data are presented as mean ± SD ( n = 3). *** p < 0.001 relative to siNC.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Migration, Staining

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: Differential gene expression patterns between GSTM3 inhibited or negative control of T98G glioblastoma cells. The volcano plot of log2 (fold change) versus −log10 ( q -value) reveals differentially upregulated (right upper quadrant) and downregulated (left upper quadrant) genes expressed in GSTM3-inhibited cells versus negative control cells. Genes with q-values >0.05 and >2-fold changes are plotted in blue (downregulated) and red (upregulated), respectively.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Gene Expression, Negative Control

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: The upregulated mRNAs revealed in GSTM3 knockdown T98G.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Knockdown

Journal: International Journal of Molecular Sciences

Article Title: Glutathione S-Transferase M3 Is Associated with Glycolysis in Intrinsic Temozolomide-Resistant Glioblastoma Multiforme Cells

doi: 10.3390/ijms22137080

Figure Lengend Snippet: The downregulated mRNAs revealed in GSTM3 knockdown T98G.

Article Snippet: The human fetal astroglia cell line SVGp12 and human brain malignant glioma cell lines T98G, A172, and U138MG were purchased from the American Type Culture Collection (Manassas, VA, USA).

Techniques: Knockdown

Journal: Oncogenesis

Article Title: Chondroitin sulfate synthase 1 enhances proliferation of glioblastoma by modulating PDGFRA stability

doi: 10.1038/s41389-020-0197-0

Figure Lengend Snippet: a Comparison of CHSY1 gene expression in glioma subtypes and normal brain tissue in the REMBRANDT glioma microarray database. ** P < 0.01, **** P < 0.0001. b High expression of CHSY1 was associated with worse overall survival in glioma patients. The high and low expression groups were divided by median expression level of CHSY1 in 329 cases. These data were from the REMBRANDT database ( http://www.betastasis.com/glioma/rembrandt/ ). c Immunohistochemistry of CHSY1 (upper panel) and CS56 (lower panel) on tissue array contains 85 primary glioma cases. The staining was visualized in brown color with a 3,3-diaminobenzidine liquid substrate system. All sections were counterstained with hematoxylin. Representative images of four glioma cases with different staining intensities are shown. Amplified images are shown at the bottom right of each image. Scale bars, 50 μm. Arrows indicate positive stained glioma cells. d Representative images of CHSY1 staining on normal brain tissue ( n = 5). e Statistical analysis of immunohistochemistry in glioma tissue array. Mann–Whitney U -test was used. P -values are shown at top. f Expression of CHSY1 in glioma cell lines and normal human brain tissue. The protein expression was analyzed by western blotting. Total loading protein is shown at bottom. Relative expression levels to total brain tissue form three independent blots are shown at the right.

Article Snippet: Adult normal human brain tissue lysates were purchased from Novus Biologicals.

Techniques: Comparison, Gene Expression, Microarray, Expressing, Immunohistochemistry, Staining, Amplification, MANN-WHITNEY, Western Blot

Journal: bioRxiv

Article Title: Monoclonal antibody to Filarial serpin Wb123 impedes the urokinase plasminogen activator receptor mediated Alternative activation of macrophages

doi: 10.1101/2025.02.05.636671

Figure Lengend Snippet: A) CD163 and IL6 expression in PBMCs of MF+ filarial patients. The graph represents fold change in gene expression of filarial patient compared with endemic normal. Data represent the mean ± SEM (n = 3). * Represents significance values compared to control, *p<0.0214, **p=0.0012, ***p=0.0006 and 0.0001, ****p<0.0001. Statistical significance was calculated using two-way ANOVA; B) SDS-PAGE stained with Coomassie brilliant blue dye and western blot analysis of purified rWb123 probed with His tag antibody; C) Immunofluorescence images showing Wb123 expression in microfilaria by confocal microscopy after incubation with monoclonal antibody; D) Study of activation marker expression after incubation with rWb123. The graph represents fold change in gene expression. Data represent the mean ± SEM (n = 3). * Represents significance values compared to control, *p<0.0194, ***p=0.0002 and 0.0005. Statistical significance was calculated using two-way ANOVA; E) Blots showing CD163 expression at protein level in rWb123 treated cells compared to control cells; F) Bar graph showing CD163 expression in different Wb123 concentrations represented as +(125ng), ++(250ng), +++(500ng), ++++(1µg). * Represents significance values compared to control, ***p=0.0008, ****p<0.0001. Statistical significance was calculated using two-way ANOVA; G) Blot showing arginase-1 expression in Wb123 treated cells and untreated cells; H) Immunofluorescence images showing CD86 expression in Wb123 treated cells and untreated cells; I) Intensity was determined from cells (Violin plot, n=70, * represents significance values compared to control, *P<0.0107, ****p<0.0001). Statistical significance was calculated using unpaired t test; J) Immunoprecipitation of Wb123 was performed from Wb123 treated HMC3 cells. Western blot analysis was performed with immunoprecipitated samples with uPA and uPAR as indicated in figure; K) Strategy for antibody blocking experiment. L) Blot showing CD163 expression in uPAR blocked cells and control cells; M) The bar graph represents quantification of CD163 expression in uPAR pre-treated and untreated cells. normalized to GAPDH (Mean± SEM, n=3, AU depicts arbitrary unit, * represents significance values compared to control, *p<0.00307).

Article Snippet: Human brain macrophage cells HMC3 (Human microglia clone 3) was ordered from ATCC (CRL-3304).

Techniques: Expressing, Gene Expression, Control, SDS Page, Staining, Western Blot, Purification, Immunofluorescence, Confocal Microscopy, Incubation, Activation Assay, Marker, Immunoprecipitation, Blocking Assay

Journal:

Article Title: Combination of vitamin E and selenium causes an induction of apoptosis of human prostate cancer cells by enhancing Bax/Bcl-2 ratio

doi: 10.1002/pros.20824

Figure Lengend Snippet: The chemical structures of methylselenic acid (MSA) (A) and vitamin E (+α-tocopherol) succinate (VES) (B) are depicted. Following treatment of LNCaP, DU145, and PC-3 cells with VES and/or MSA for 48 hours, the effects on cell viability (C) and cell growth (D) were measured using Trypan Blue Exclusion analysis. Similarly, following treatment of PrEC with VES and/or MSA for 48 hours, the effects on cell viability (E) and cell growth (F) were also measured. Cell viability data is expressed as the percent viable cells out of the total number of cells. The data is expressed as the mean ± SE of three experiments (* p < 0.01 versus untreated control) (# p< 0.01 for combination versus either single agent). Details of the experiments are given in ‘Materials and Methods’.

Article Snippet: Human prostate carcinoma cell lines LNCaP (lymph node-derived androgen-sensitive cell line; normal for cell cycle-related tumor suppressor genes p53 and retinoblastoma Rb), PC-3 (bone marrow-derived androgen-insensitive cell line; defective for both p53 alleles but normal for both Rb alleles), and DU145 (brain-derived androgen-insensitive cell line; defective for both p53 and both Rb alleles) were purchased from ATCC (Manassas, VA) and maintained in RPMI-1640, MEM, and F12K media, respectively, with 10% Fetal Bovine Serum and 1% Penicillin-Streptomycin (ATCC; Manassas, VA) at standard cell culture conditions (37°C, 5% CO 2 in a humidified incubator).

Techniques: Control

Journal:

Article Title: Combination of vitamin E and selenium causes an induction of apoptosis of human prostate cancer cells by enhancing Bax/Bcl-2 ratio

doi: 10.1002/pros.20824

Figure Lengend Snippet: Following treatment of LNCaP, DU145, and PC-3 cells with VES and/or MSA for 48 hours, the cell cycle distribution was assessed using the APO-BrdU TUNEL Assay kit. The FAC profiles indicating the positions of G0/G1, S, and G2/M are shown, with the sub- G0 content excluded as much as possible (A). The effect of treatments on clonogenic survival of PCa cells was determined using colony formation assay (B). Following treatments, the cells were replated in triplicate on a 6-well tissue culture plate with 3000 cells/well. The cells were cultured for 12-14 days with growth media being replaced every 3 days. The cells were then stained with 0.5% crystal violet (in methanol:H2O, 1:1) and pictures were taken using a digital camera. The pictures were enhances using Adobe Photoshop for brightness, contrast, and sharpening for uniformity of appearance. Details of the experiments are given in ‘Materials and Methods’.

Article Snippet: Human prostate carcinoma cell lines LNCaP (lymph node-derived androgen-sensitive cell line; normal for cell cycle-related tumor suppressor genes p53 and retinoblastoma Rb), PC-3 (bone marrow-derived androgen-insensitive cell line; defective for both p53 alleles but normal for both Rb alleles), and DU145 (brain-derived androgen-insensitive cell line; defective for both p53 and both Rb alleles) were purchased from ATCC (Manassas, VA) and maintained in RPMI-1640, MEM, and F12K media, respectively, with 10% Fetal Bovine Serum and 1% Penicillin-Streptomycin (ATCC; Manassas, VA) at standard cell culture conditions (37°C, 5% CO 2 in a humidified incubator).

Techniques: TUNEL Assay, Colony Assay, Cell Culture, Staining

Journal:

Article Title: Combination of vitamin E and selenium causes an induction of apoptosis of human prostate cancer cells by enhancing Bax/Bcl-2 ratio

doi: 10.1002/pros.20824

Figure Lengend Snippet: For determining the effect of treatments on PCNA, following treatment of LNCaP, DU145, and PC-3 cells with VES and/or MSA for 48 hours, the protein level of PCNA was assessed by Western blot analysis (A) and quantified by densitometric analysis of protein bands (B). For determining the effect on PSA protein levels, following treatment of LNCaP cells with VES and/or MSA for 48 hours, the protein level of PSA was assessed by Western blot analysis (C) and quantified by densitometric analysis of protein bands (D). Equal loading was confirmed by stripping the blot and reprobing it for β-actin. The data is expressed as mean ± SE of three experiments (* p < 0.01 versus untreated control) (# p< 0.01 for combination versus either single agent). Details of the experiments are given in ‘Materials and Methods’.

Article Snippet: Human prostate carcinoma cell lines LNCaP (lymph node-derived androgen-sensitive cell line; normal for cell cycle-related tumor suppressor genes p53 and retinoblastoma Rb), PC-3 (bone marrow-derived androgen-insensitive cell line; defective for both p53 alleles but normal for both Rb alleles), and DU145 (brain-derived androgen-insensitive cell line; defective for both p53 and both Rb alleles) were purchased from ATCC (Manassas, VA) and maintained in RPMI-1640, MEM, and F12K media, respectively, with 10% Fetal Bovine Serum and 1% Penicillin-Streptomycin (ATCC; Manassas, VA) at standard cell culture conditions (37°C, 5% CO 2 in a humidified incubator).

Techniques: Western Blot, Stripping Membranes, Control

Journal:

Article Title: Combination of vitamin E and selenium causes an induction of apoptosis of human prostate cancer cells by enhancing Bax/Bcl-2 ratio

doi: 10.1002/pros.20824

Figure Lengend Snippet: Following treatment of LNCaP, DU145, and PC-3 cells with VES and/or MSA for 48 hours, the extent of apoptosis was assessed with the APO-BrdU TUNEL Assay kit. BrdU incorporation was analyzed with a flow cytometer (A), followed by a computational analysis (B), of cells staining positive for BrdU. The data is expressed as mean ± SE of three experiments (* p < 0.01 versus untreated control) (# p< 0.01 for combination versus either single agent). Details of the experiments are given in ‘Materials and Methods’.

Article Snippet: Human prostate carcinoma cell lines LNCaP (lymph node-derived androgen-sensitive cell line; normal for cell cycle-related tumor suppressor genes p53 and retinoblastoma Rb), PC-3 (bone marrow-derived androgen-insensitive cell line; defective for both p53 alleles but normal for both Rb alleles), and DU145 (brain-derived androgen-insensitive cell line; defective for both p53 and both Rb alleles) were purchased from ATCC (Manassas, VA) and maintained in RPMI-1640, MEM, and F12K media, respectively, with 10% Fetal Bovine Serum and 1% Penicillin-Streptomycin (ATCC; Manassas, VA) at standard cell culture conditions (37°C, 5% CO 2 in a humidified incubator).

Techniques: TUNEL Assay, BrdU Incorporation Assay, Flow Cytometry, Staining, Control

Journal:

Article Title: Combination of vitamin E and selenium causes an induction of apoptosis of human prostate cancer cells by enhancing Bax/Bcl-2 ratio

doi: 10.1002/pros.20824

Figure Lengend Snippet: Following treatment of LNCaP, DU145, and PC-3 cells with VES and/or MSA for 48 hours, the protein levels of Bcl-2, Bax, Bak, and Bid were assessed by Western blot analysis (A). Equal loading was confirmed by stripping the blot and reprobing it for β-actin. The effect of treatment on the Bax/Bcl-2 ratio at the protein level (B) was determined by densitometric analysis of the immunoblots relative to β-actin. Quantitative Real-Time PCR analysis was used to calculate the Bax/Bcl-2 ratio at the mRNA level (C) relative to GAPDH. The data is expressed as mean ± SE of three experiments (* p < 0.01 versus untreated control) (# p< 0.01 for combination versus either single agent). Details of the experiments are given in ‘Materials and Methods’.

Article Snippet: Human prostate carcinoma cell lines LNCaP (lymph node-derived androgen-sensitive cell line; normal for cell cycle-related tumor suppressor genes p53 and retinoblastoma Rb), PC-3 (bone marrow-derived androgen-insensitive cell line; defective for both p53 alleles but normal for both Rb alleles), and DU145 (brain-derived androgen-insensitive cell line; defective for both p53 and both Rb alleles) were purchased from ATCC (Manassas, VA) and maintained in RPMI-1640, MEM, and F12K media, respectively, with 10% Fetal Bovine Serum and 1% Penicillin-Streptomycin (ATCC; Manassas, VA) at standard cell culture conditions (37°C, 5% CO 2 in a humidified incubator).

Techniques: Western Blot, Stripping Membranes, Real-time Polymerase Chain Reaction, Control