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  • 95
    Vector Laboratories propidium iodide
    MEI-S332–GFP localizes to centromeric regions of female meiotic chromosomes until anaphase II. MEI-S332–GFP is shown in green and chromatin in red. ( A ) Unactivated stage 14 oocytes are arrested in metaphase I, with MEI-S332–GFP localized to two discrete sites on the opposite ends of the condensed meiotic chromosomes. ( B ) At the onset of anaphase I, eight dots of MEI-S332–GFP are visible at the leading edges of the separating anaphase chromosomes, one per pair of sister chromatids, with the fourth chromosomes closest to the poles. Chromosome 4 in Drosophila is very small and sometimes difficult to visualize. ( C ) In late anaphase I, MEI-S332–GFP is still detected at the leading edges of the chromosomes, which become shorter and rounder as they approach the poles. ( D ) Between the first and second meiotic divisions, nuclear decondensation does not occur. Rather, two clusters of 3–4 chromatin balls are observed. Each ball most likely represents a pair of sister chromatids and is associated with a dot of MEI-S332– GFP. ( E ) In metaphase II, the chromatin balls move together to form metaphase plates, and MEI-S322–GFP localizes to the middle of the chromatin. ( F ) When sister-chromatid cohesion is released at anaphase II, the sister chromatids separate, and MEI-S332–GFP is no longer detectable on the meiotic chromosomes. ( G ) During the postmeiotic interphase, MEI-S332–GFP is not visible on the decondensed chromosomes. Oocytes were isolated from females carrying four copies of the mei - S332 + ::GFP transgene, activated in vitro, fixed, and stained with <t>propidium</t> iodide. Images were collected using confocal microscopy. Bar, ∼5 μm.
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    Millipore propidium iodide
    MEI-S332–GFP localizes to centromeric regions of female meiotic chromosomes until anaphase II. MEI-S332–GFP is shown in green and chromatin in red. ( A ) Unactivated stage 14 oocytes are arrested in metaphase I, with MEI-S332–GFP localized to two discrete sites on the opposite ends of the condensed meiotic chromosomes. ( B ) At the onset of anaphase I, eight dots of MEI-S332–GFP are visible at the leading edges of the separating anaphase chromosomes, one per pair of sister chromatids, with the fourth chromosomes closest to the poles. Chromosome 4 in Drosophila is very small and sometimes difficult to visualize. ( C ) In late anaphase I, MEI-S332–GFP is still detected at the leading edges of the chromosomes, which become shorter and rounder as they approach the poles. ( D ) Between the first and second meiotic divisions, nuclear decondensation does not occur. Rather, two clusters of 3–4 chromatin balls are observed. Each ball most likely represents a pair of sister chromatids and is associated with a dot of MEI-S332– GFP. ( E ) In metaphase II, the chromatin balls move together to form metaphase plates, and MEI-S322–GFP localizes to the middle of the chromatin. ( F ) When sister-chromatid cohesion is released at anaphase II, the sister chromatids separate, and MEI-S332–GFP is no longer detectable on the meiotic chromosomes. ( G ) During the postmeiotic interphase, MEI-S332–GFP is not visible on the decondensed chromosomes. Oocytes were isolated from females carrying four copies of the mei - S332 + ::GFP transgene, activated in vitro, fixed, and stained with <t>propidium</t> iodide. Images were collected using confocal microscopy. Bar, ∼5 μm.
    Propidium Iodide, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 70224 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher propidium iodide
    Knock-down of endogenous Ginir affects proliferation and tumourigenic potential of fibroblasts and melanoma cells. (A) Representative RT-PCR for analyses of Ginir knock-down in NIH/3T3 cells expressing Ginir-shRNA (1, 2, and scrambled control) using G2F-G2R primers. Gapdh served as an internal control. (B) Cell proliferation analysis by MTT assay for the indicated cell lines performed over a period of 4 days. Data presented are mean ± SEM. *** P ≤ 0.0001, one tailed, by two-way ANOVA test. (C) Quantification of Ki67 antigen–expressing cells shown as percentage of positively stained cells from the total number of cells present per field. Cells from at least 10 representative fields were counted in NIH/3T3 control and Ginir shRNA1 and 2 cells. Data presented are mean ± SEM.** P ≤ 0.001 by one-way ANOVA test ( n = 3). (D) Quantitative analyses of cell cycle parameters of PI-stained cells in flow cytometry. Stable Ginir knock-down (NIHshGinir1, NIHshGinir2) cells were compared to NIH/3T3 control cells having endogenous Ginir RNA expression for percentage of cells in various phases of the cell cycle. Bars: means ± SEM; * P ≤ 0.05, ** P ≤ 0.001; two tailed; by two-way ANOVA test. (E) Representative RT-PCR data demonstrating Ginir RNA expression levels in B16F10 melanoma cells shControl and the knock-down cells B16F10-Ginir-shRNA1 and B16F10-GinirshRNA2. PCR was done using G2F-G2R primers. Gapdh RNA expression was used for normalisation. (F) Phase contrast images of B16F10 control cells and cells stably transfected with shGinir1 or shGinir2. Arrowheads point towards cells showing altered phenotypes. Magnification 10×. (G) Cell proliferation analysis of indicated cells performed by MTT assay over a period of 4 days. Data shown are mean ± SEM. * P ≤ 0.05, one tailed, Student’s paired t test ( n = 3). (H) Quantification of Ki67 antigen expression shown as percentage of positively stained cells as compared to the total number of cells per field. At least 10 fields were counted. Data shown are mean ± SEM. ** P ≤ 0.001 by one-way ANOVA test. ( n = 3). (I) Cell migration assay in the indicated cell lines. The gaps were measured after 6 hours using ImageJ software; version 1.41. Experiment was repeated at least thrice. (J) Quantitative analysis of relative wound recovery of each B16F10 Ginir knock-down cell induced by shRNA1 and shRNA2 as compared to B16F10-shGinir scrambled expressing control cells. Data represent mean ± SEM ( n = 3). *** P ≤ 0.0001 by two-way ANOVA test. (K) Representative xenograft tumours of B16F10-shControl and B16F10-shGinir1 and B16F10shGinir2 (1 and 2) cells introduced into NOD/SCID mice; tumour growth was monitored for 15 days. These experiments were performed thrice, with 3 mice in each group. (L) Tumour growth kinetics were determined by measuring tumour volume each day for a period of 15 days for the indicated cell lines. Data shown represent mean ± SEM. *** P . Gapdh, glyceride 3-phosphate dehydrogenase; Ginir, Genomic Instability Inducing RNA; MTT, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide; PCR, polymerase chain reaction; PI, <t>propidium</t> iodide; RT-PCR, reverse transcription PCR; shRNA, short hairpin RNA.
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    Becton Dickinson propidium iodide
    Ectopic expression of chimeric Myc-ER protein prevents TGF-β1–mediated inhibition of FasL mRNA and subsequent AICD. (A) Functional Myc-ER interferes with the inhibitory effect of TGF-β1 on activation-induced FasL expression in A1.1 cells. A1.1 or A1.1 Myc-ER cells exposed to 4-OHT (50 nM) were first preincubated for 4 h with the drug, which was then also present during subsequent culture. Cells were then incubated in medium alone (lane 1) or activated for 4 h with anti-CD3 antibodies (lanes 2, 3, and 4). 1 ng/ml TGF-β1 (lane 3) or 100 ng/ ml CsA (lane 4) was added to some cultures. FasL expression was then assessed by RT-PCR. (B) Ectopic expression of a chimeric Myc-ER protein prevents AICD after TGF-β1 treatment. A1.1 or A1.1 Myc-ER T hybridoma cells were first preincubated for 4 h in the presence or absence of 4-OHT (50 nM) and then activated for 16 h with anti-CD3 antibodies with the indicated concentrations of TGF-β1 (1 ng/ml) or CsA (100 ng/ml). Cell death was assessed by <t>propidium</t> iodide uptake using a FACScan ® .
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    Becton Dickinson propidium iodide pi
    Phosphatidylinositol 3-kinase/Akt acts downstream of FAK signaling to regulate apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were treated with LY294002. (A and B) The expression of Akt, pAkt, caspase-3 and c-caspase-3 was examined using western blotting. (C and D) Cell apoptosis was examined using (C) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (D) Annexin <t>V/propidium</t> iodide. (E and F) The expression of FAK and pFAK was examined using western blotting. Scale bar, 200 µm. FAK, focal adhesion kinase; pFAK, phosphorylated FAK; c-caspase-3, cleaved caspase-3; TGFβ, transforming growth factor-β.
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    Millipore propidium iodide solution
    Neutralization of the PM-MSC complex . (a, b) Viability of rat fetal membrane-derived MSCs at 10 min after mixture of PM (final dose 0.8%) with or without neutralization using NaOH in-vitro was assessed by <t>propidium</t> iodide staining. Representative pictures in each group are presented (a). Scale bar = 50 μm. The bar graph presents the averaged data (b). n=4 in each point ; * p
    Propidium Iodide Solution, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2085 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Beyotime propidium iodide
    Neutralization of the PM-MSC complex . (a, b) Viability of rat fetal membrane-derived MSCs at 10 min after mixture of PM (final dose 0.8%) with or without neutralization using NaOH in-vitro was assessed by <t>propidium</t> iodide staining. Representative pictures in each group are presented (a). Scale bar = 50 μm. The bar graph presents the averaged data (b). n=4 in each point ; * p
    Propidium Iodide, supplied by Beyotime, used in various techniques. Bioz Stars score: 97/100, based on 1778 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    BioLegend propidium iodide
    Baricitinib suppresses CD80/CD86 expression on dendritic cells (DCs). Immature monocyte-derived dendritic cells were cultured with or without baricitinib and tofacitinib during lipopolysaccharide stimulation for 48 h. The DC phenotype was evaluated using flow cytometry. (A) Expression of HLA-DR, CD80, and CD86. (B) Representative histogram data of HLA-DR, CD80, and CD86 expression. (C) Rate of viable cells (annexin V neg <t>/propidium</t> iodide neg ). (D) Expression of CD80 and CD86 in the presence of baricitinib and tofacitinib. Data are mean ± SD of three different donors per group. * p
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    Thermo Fisher propidium iodide 1 0 solution in water
    Baricitinib suppresses CD80/CD86 expression on dendritic cells (DCs). Immature monocyte-derived dendritic cells were cultured with or without baricitinib and tofacitinib during lipopolysaccharide stimulation for 48 h. The DC phenotype was evaluated using flow cytometry. (A) Expression of HLA-DR, CD80, and CD86. (B) Representative histogram data of HLA-DR, CD80, and CD86 expression. (C) Rate of viable cells (annexin V neg <t>/propidium</t> iodide neg ). (D) Expression of CD80 and CD86 in the presence of baricitinib and tofacitinib. Data are mean ± SD of three different donors per group. * p
    Propidium Iodide 1 0 Solution In Water, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 1368 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Keygen Biotech propidium iodide
    Radioresistant TE-1 and Eca-109 cells showed more aggressive malignancies. (A) Wound-healing assay of TE-1 and TE-1/R cells. (B) Wound-healing assay of Eca-109 and Eca-109/R cells. Wound healing was observed 24 h after the treatment. (C) Induction of apoptosis by radiation in TE-1 and TE-1/R cells. (D) Induction of apoptosis by radiation in Eca-109 and Eca-109/R cells. Cells were treated with 6 Gy irradiation, and the apoptosis was measured using <t>propidium</t> iodide (PI)/Annexin-V double staining. Data are normalized to the control cells and presented as the mean ± SEM of three independent experiments, * P
    Propidium Iodide, supplied by Keygen Biotech, used in various techniques. Bioz Stars score: 92/100, based on 1347 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Becton Dickinson propidium iodide staining
    PCB-induced arrest is reversible and cells regain sensitivity to VCR after reentering the cell cycle. A . ALL-5 and T98G cells were stained with <t>propidium</t> iodide and subjected to two-parameter flow cytometry to create a gate distinguishing EdU-negative and EdU-positive cells (left panels). ALL-5 and T98G cells were then treated with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G), washed thrice with PBS, and incubated in growth media containing 10 µM EdU for the times indicated. Cells were harvested, fixed, and stained for EdU incorporation and with propidium iodide and analyzed by two-parameter flow cytometry. Data shown are representative of three independent experiments. The percentage of cells positive for EdU is indicated at the top of each panel. B . ALL-5 and T98G cells were treated with 0.1% DMSO or 100 nM (ALL-5) or 1 µM (T98G) VCR for 48 h with or without pretreatment with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G). A third set of cells were pretreated with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G), after which cells were washed thrice with PBS and treated with 0.1% DMSO or 100 nM (ALL-5) or 1 µM (T98G) VCR for 48 h. Cells were stained with propidium iodide and subjected to flow cytometry to determine the percentage of cells with
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    Keygen Biotech propidium iodide pi
    The effect of oridonin nanosuspension and free oridonin on PANC-1 cell apoptosis was measured by annexin V-fluorescein <t>isothiocyanate/propidium</t> iodide staining. The early apoptotic cells stained by annexin-V-fluorescein isothiocyanate are located in the lower right quadrant. Abbreviation: ORI, oridonin.
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    Abcam propidium iodide
    The effect of oridonin nanosuspension and free oridonin on PANC-1 cell apoptosis was measured by annexin V-fluorescein <t>isothiocyanate/propidium</t> iodide staining. The early apoptotic cells stained by annexin-V-fluorescein isothiocyanate are located in the lower right quadrant. Abbreviation: ORI, oridonin.
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    BioVision propidium iodide pi
    Effects of CD54 Knockdown on Cell Proliferation and Apoptosis. ( a ) BrdU analysis of cell cycle progression in prostate cancer patient cells transfected with shCtrl or shCD54-1. ( b ) Ki67 analysis of cell cycle progression in prostate cancer patient cells transfected with shCtrl or shCD54-1. ( c ) FACS analysis of <t>propidium</t> iodide- and annexin V-stained prostate cancer patient cells (from two different patients) transfected with shCtrl or shCD54-1. ( d ) Representative light micrograph fields and comparative quantification of the migratory capacity of prostate cancer patient cells (from two different patients) transfected with shCtrl or shCD54-1 via Transwell assay. ( e ) Representative microscope fields and comparative quantification of apoptosis in vehicle-treated or cisplatin (DDP)-treated PC3 cells transfected with shCtrl or shCD54-1. Arrows indicate apoptotic cells. ( f ) The cisplatin (DDP) IC 50 values of 5 prostate cancer patient cell lines transfected with shCtrl or shCD54-1. * P
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    N/A
    Propidium iodide PI is membrane impermeant and generally excluded from viable cells However it can easily penetrate dead or damaged cells and as such is commonly used for identifying cell
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    Propidium Iodide PI binds to double stranded DNA and RNA after cells have been permeabilized Once bound to nucleic acids the nuclei appear red in color as observed by fluorescence
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    Image Search Results


    MEI-S332–GFP localizes to centromeric regions of female meiotic chromosomes until anaphase II. MEI-S332–GFP is shown in green and chromatin in red. ( A ) Unactivated stage 14 oocytes are arrested in metaphase I, with MEI-S332–GFP localized to two discrete sites on the opposite ends of the condensed meiotic chromosomes. ( B ) At the onset of anaphase I, eight dots of MEI-S332–GFP are visible at the leading edges of the separating anaphase chromosomes, one per pair of sister chromatids, with the fourth chromosomes closest to the poles. Chromosome 4 in Drosophila is very small and sometimes difficult to visualize. ( C ) In late anaphase I, MEI-S332–GFP is still detected at the leading edges of the chromosomes, which become shorter and rounder as they approach the poles. ( D ) Between the first and second meiotic divisions, nuclear decondensation does not occur. Rather, two clusters of 3–4 chromatin balls are observed. Each ball most likely represents a pair of sister chromatids and is associated with a dot of MEI-S332– GFP. ( E ) In metaphase II, the chromatin balls move together to form metaphase plates, and MEI-S322–GFP localizes to the middle of the chromatin. ( F ) When sister-chromatid cohesion is released at anaphase II, the sister chromatids separate, and MEI-S332–GFP is no longer detectable on the meiotic chromosomes. ( G ) During the postmeiotic interphase, MEI-S332–GFP is not visible on the decondensed chromosomes. Oocytes were isolated from females carrying four copies of the mei - S332 + ::GFP transgene, activated in vitro, fixed, and stained with propidium iodide. Images were collected using confocal microscopy. Bar, ∼5 μm.

    Journal: The Journal of Cell Biology

    Article Title: The Cohesion Protein MEI-S332 Localizes to Condensed Meiotic and Mitotic Centromeres until Sister Chromatids Separate

    doi:

    Figure Lengend Snippet: MEI-S332–GFP localizes to centromeric regions of female meiotic chromosomes until anaphase II. MEI-S332–GFP is shown in green and chromatin in red. ( A ) Unactivated stage 14 oocytes are arrested in metaphase I, with MEI-S332–GFP localized to two discrete sites on the opposite ends of the condensed meiotic chromosomes. ( B ) At the onset of anaphase I, eight dots of MEI-S332–GFP are visible at the leading edges of the separating anaphase chromosomes, one per pair of sister chromatids, with the fourth chromosomes closest to the poles. Chromosome 4 in Drosophila is very small and sometimes difficult to visualize. ( C ) In late anaphase I, MEI-S332–GFP is still detected at the leading edges of the chromosomes, which become shorter and rounder as they approach the poles. ( D ) Between the first and second meiotic divisions, nuclear decondensation does not occur. Rather, two clusters of 3–4 chromatin balls are observed. Each ball most likely represents a pair of sister chromatids and is associated with a dot of MEI-S332– GFP. ( E ) In metaphase II, the chromatin balls move together to form metaphase plates, and MEI-S322–GFP localizes to the middle of the chromatin. ( F ) When sister-chromatid cohesion is released at anaphase II, the sister chromatids separate, and MEI-S332–GFP is no longer detectable on the meiotic chromosomes. ( G ) During the postmeiotic interphase, MEI-S332–GFP is not visible on the decondensed chromosomes. Oocytes were isolated from females carrying four copies of the mei - S332 + ::GFP transgene, activated in vitro, fixed, and stained with propidium iodide. Images were collected using confocal microscopy. Bar, ∼5 μm.

    Article Snippet: Embryos in Figs. A , C , and D were treated with 1 mg/ml RNase A for 30 min, stained with 1 μg/ml propidium iodide for 30 min, and mounted in Vectashield with propidium iodide (Vector Labs Inc.).

    Techniques: Isolation, In Vitro, Staining, Confocal Microscopy

    MEI-S332–GFP localizes to condensed chromosomes in embryos. MEI-S332– GFP is shown in green and DNA in red. ( A ) MEI-S332–GFP is present on the polar body rosettes. ( B ) A close-up image of a polar body rosette shows punctate MEI-S332–GFP localization on the inside ring of the rosette where centromeres are believed to be pulled to the center. 22 dots of MEI-S332–GFP can be counted in the single rosette found in this embryo. ( C ) MEI-S332–GFP localizes to discrete dots on a mitotic metaphase plate, resembling those on meiotic metaphase II chromosomes. In addition, a cloud of diffuse MEI-S332–GFP is observed around each mitotic nucleus. ( D ) MEI-S332–GFP is detected in clouds surrounding the interphase nuclei. The nuclei are not centered within the clouds. ( E ) A close-up image of the interphase nucleus demonstrates the absence of MEI-S332–GFP localization on the decondensed interphase chromatin. Embryos were collected from females carrying four copies of the mei - S332 + ::GFP transgene, fixed, and stained with either propidium iodide or DAPI. Images in A , C , and D were collected using confocal microscopy, and images in B and E were collected using a CCD camera. Bars: A–C and E ∼5 μm; ( D ) ∼30 μm.

    Journal: The Journal of Cell Biology

    Article Title: The Cohesion Protein MEI-S332 Localizes to Condensed Meiotic and Mitotic Centromeres until Sister Chromatids Separate

    doi:

    Figure Lengend Snippet: MEI-S332–GFP localizes to condensed chromosomes in embryos. MEI-S332– GFP is shown in green and DNA in red. ( A ) MEI-S332–GFP is present on the polar body rosettes. ( B ) A close-up image of a polar body rosette shows punctate MEI-S332–GFP localization on the inside ring of the rosette where centromeres are believed to be pulled to the center. 22 dots of MEI-S332–GFP can be counted in the single rosette found in this embryo. ( C ) MEI-S332–GFP localizes to discrete dots on a mitotic metaphase plate, resembling those on meiotic metaphase II chromosomes. In addition, a cloud of diffuse MEI-S332–GFP is observed around each mitotic nucleus. ( D ) MEI-S332–GFP is detected in clouds surrounding the interphase nuclei. The nuclei are not centered within the clouds. ( E ) A close-up image of the interphase nucleus demonstrates the absence of MEI-S332–GFP localization on the decondensed interphase chromatin. Embryos were collected from females carrying four copies of the mei - S332 + ::GFP transgene, fixed, and stained with either propidium iodide or DAPI. Images in A , C , and D were collected using confocal microscopy, and images in B and E were collected using a CCD camera. Bars: A–C and E ∼5 μm; ( D ) ∼30 μm.

    Article Snippet: Embryos in Figs. A , C , and D were treated with 1 mg/ml RNase A for 30 min, stained with 1 μg/ml propidium iodide for 30 min, and mounted in Vectashield with propidium iodide (Vector Labs Inc.).

    Techniques: Staining, Confocal Microscopy

    Knock-down of endogenous Ginir affects proliferation and tumourigenic potential of fibroblasts and melanoma cells. (A) Representative RT-PCR for analyses of Ginir knock-down in NIH/3T3 cells expressing Ginir-shRNA (1, 2, and scrambled control) using G2F-G2R primers. Gapdh served as an internal control. (B) Cell proliferation analysis by MTT assay for the indicated cell lines performed over a period of 4 days. Data presented are mean ± SEM. *** P ≤ 0.0001, one tailed, by two-way ANOVA test. (C) Quantification of Ki67 antigen–expressing cells shown as percentage of positively stained cells from the total number of cells present per field. Cells from at least 10 representative fields were counted in NIH/3T3 control and Ginir shRNA1 and 2 cells. Data presented are mean ± SEM.** P ≤ 0.001 by one-way ANOVA test ( n = 3). (D) Quantitative analyses of cell cycle parameters of PI-stained cells in flow cytometry. Stable Ginir knock-down (NIHshGinir1, NIHshGinir2) cells were compared to NIH/3T3 control cells having endogenous Ginir RNA expression for percentage of cells in various phases of the cell cycle. Bars: means ± SEM; * P ≤ 0.05, ** P ≤ 0.001; two tailed; by two-way ANOVA test. (E) Representative RT-PCR data demonstrating Ginir RNA expression levels in B16F10 melanoma cells shControl and the knock-down cells B16F10-Ginir-shRNA1 and B16F10-GinirshRNA2. PCR was done using G2F-G2R primers. Gapdh RNA expression was used for normalisation. (F) Phase contrast images of B16F10 control cells and cells stably transfected with shGinir1 or shGinir2. Arrowheads point towards cells showing altered phenotypes. Magnification 10×. (G) Cell proliferation analysis of indicated cells performed by MTT assay over a period of 4 days. Data shown are mean ± SEM. * P ≤ 0.05, one tailed, Student’s paired t test ( n = 3). (H) Quantification of Ki67 antigen expression shown as percentage of positively stained cells as compared to the total number of cells per field. At least 10 fields were counted. Data shown are mean ± SEM. ** P ≤ 0.001 by one-way ANOVA test. ( n = 3). (I) Cell migration assay in the indicated cell lines. The gaps were measured after 6 hours using ImageJ software; version 1.41. Experiment was repeated at least thrice. (J) Quantitative analysis of relative wound recovery of each B16F10 Ginir knock-down cell induced by shRNA1 and shRNA2 as compared to B16F10-shGinir scrambled expressing control cells. Data represent mean ± SEM ( n = 3). *** P ≤ 0.0001 by two-way ANOVA test. (K) Representative xenograft tumours of B16F10-shControl and B16F10-shGinir1 and B16F10shGinir2 (1 and 2) cells introduced into NOD/SCID mice; tumour growth was monitored for 15 days. These experiments were performed thrice, with 3 mice in each group. (L) Tumour growth kinetics were determined by measuring tumour volume each day for a period of 15 days for the indicated cell lines. Data shown represent mean ± SEM. *** P . Gapdh, glyceride 3-phosphate dehydrogenase; Ginir, Genomic Instability Inducing RNA; MTT, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide; PCR, polymerase chain reaction; PI, propidium iodide; RT-PCR, reverse transcription PCR; shRNA, short hairpin RNA.

    Journal: PLoS Biology

    Article Title: Noncoding RNA Ginir functions as an oncogene by associating with centrosomal proteins

    doi: 10.1371/journal.pbio.2004204

    Figure Lengend Snippet: Knock-down of endogenous Ginir affects proliferation and tumourigenic potential of fibroblasts and melanoma cells. (A) Representative RT-PCR for analyses of Ginir knock-down in NIH/3T3 cells expressing Ginir-shRNA (1, 2, and scrambled control) using G2F-G2R primers. Gapdh served as an internal control. (B) Cell proliferation analysis by MTT assay for the indicated cell lines performed over a period of 4 days. Data presented are mean ± SEM. *** P ≤ 0.0001, one tailed, by two-way ANOVA test. (C) Quantification of Ki67 antigen–expressing cells shown as percentage of positively stained cells from the total number of cells present per field. Cells from at least 10 representative fields were counted in NIH/3T3 control and Ginir shRNA1 and 2 cells. Data presented are mean ± SEM.** P ≤ 0.001 by one-way ANOVA test ( n = 3). (D) Quantitative analyses of cell cycle parameters of PI-stained cells in flow cytometry. Stable Ginir knock-down (NIHshGinir1, NIHshGinir2) cells were compared to NIH/3T3 control cells having endogenous Ginir RNA expression for percentage of cells in various phases of the cell cycle. Bars: means ± SEM; * P ≤ 0.05, ** P ≤ 0.001; two tailed; by two-way ANOVA test. (E) Representative RT-PCR data demonstrating Ginir RNA expression levels in B16F10 melanoma cells shControl and the knock-down cells B16F10-Ginir-shRNA1 and B16F10-GinirshRNA2. PCR was done using G2F-G2R primers. Gapdh RNA expression was used for normalisation. (F) Phase contrast images of B16F10 control cells and cells stably transfected with shGinir1 or shGinir2. Arrowheads point towards cells showing altered phenotypes. Magnification 10×. (G) Cell proliferation analysis of indicated cells performed by MTT assay over a period of 4 days. Data shown are mean ± SEM. * P ≤ 0.05, one tailed, Student’s paired t test ( n = 3). (H) Quantification of Ki67 antigen expression shown as percentage of positively stained cells as compared to the total number of cells per field. At least 10 fields were counted. Data shown are mean ± SEM. ** P ≤ 0.001 by one-way ANOVA test. ( n = 3). (I) Cell migration assay in the indicated cell lines. The gaps were measured after 6 hours using ImageJ software; version 1.41. Experiment was repeated at least thrice. (J) Quantitative analysis of relative wound recovery of each B16F10 Ginir knock-down cell induced by shRNA1 and shRNA2 as compared to B16F10-shGinir scrambled expressing control cells. Data represent mean ± SEM ( n = 3). *** P ≤ 0.0001 by two-way ANOVA test. (K) Representative xenograft tumours of B16F10-shControl and B16F10-shGinir1 and B16F10shGinir2 (1 and 2) cells introduced into NOD/SCID mice; tumour growth was monitored for 15 days. These experiments were performed thrice, with 3 mice in each group. (L) Tumour growth kinetics were determined by measuring tumour volume each day for a period of 15 days for the indicated cell lines. Data shown represent mean ± SEM. *** P . Gapdh, glyceride 3-phosphate dehydrogenase; Ginir, Genomic Instability Inducing RNA; MTT, 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide; PCR, polymerase chain reaction; PI, propidium iodide; RT-PCR, reverse transcription PCR; shRNA, short hairpin RNA.

    Article Snippet: Nuclei were stained with PI (Invitrogen, # P1304MP) for 30 minutes.

    Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, shRNA, MTT Assay, One-tailed Test, Staining, Flow Cytometry, Cytometry, RNA Expression, Two Tailed Test, Polymerase Chain Reaction, Stable Transfection, Transfection, Cell Migration Assay, Software, Mouse Assay

    Cell proliferation and toxicity. BM-MSCs were incubated in growth medium for 24 h in absence (vehicle) or in presence of 1 μ M exogenous sphingosine-1-phosphate (exoS1P), 2 μ M S1PR1 receptor antagonist, W146, or 2 μ M S1PR1 receptor agonist, SEW2871. (a) Representative confocal immunofluorescence images of Ki67 expression. BM-MSCs were immunostained with the specific antibody Ki67 (green), a nuclear proliferation marker, and counterstained with propidium iodide (PI; red). Yellow colour indicates colocalization of red and green fluorescence signals. Scale bar 50 μ . Data are mean ± S.E.M. of four independent experiments performed in quadruplicate. (c) Western blotting analysis of apoptotic (Bax) and autophagic (Beclin) markers. Cell lysates (10–25 μ g) obtained from BM-MSCs were loaded onto SDS-PAGE and proteins immunodetected by specific antibodies. β -Actin was used as loading control. Blot shown is representative of at least three independent experiments with similar results. Data resulting from densitometric analysis of at least three independent experiments are shown in the graph (mean ± S.E.M.). Significance of differences in (a) and (b) (one-way ANOVA and Newman-Keuls multiple comparison test): ∗ p

    Journal: Stem Cells International

    Article Title: Sphingosine 1-Phosphate Receptor 1 Is Required for MMP-2 Function in Bone Marrow Mesenchymal Stromal Cells: Implications for Cytoskeleton Assembly and Proliferation

    doi: 10.1155/2018/5034679

    Figure Lengend Snippet: Cell proliferation and toxicity. BM-MSCs were incubated in growth medium for 24 h in absence (vehicle) or in presence of 1 μ M exogenous sphingosine-1-phosphate (exoS1P), 2 μ M S1PR1 receptor antagonist, W146, or 2 μ M S1PR1 receptor agonist, SEW2871. (a) Representative confocal immunofluorescence images of Ki67 expression. BM-MSCs were immunostained with the specific antibody Ki67 (green), a nuclear proliferation marker, and counterstained with propidium iodide (PI; red). Yellow colour indicates colocalization of red and green fluorescence signals. Scale bar 50 μ . Data are mean ± S.E.M. of four independent experiments performed in quadruplicate. (c) Western blotting analysis of apoptotic (Bax) and autophagic (Beclin) markers. Cell lysates (10–25 μ g) obtained from BM-MSCs were loaded onto SDS-PAGE and proteins immunodetected by specific antibodies. β -Actin was used as loading control. Blot shown is representative of at least three independent experiments with similar results. Data resulting from densitometric analysis of at least three independent experiments are shown in the graph (mean ± S.E.M.). Significance of differences in (a) and (b) (one-way ANOVA and Newman-Keuls multiple comparison test): ∗ p

    Article Snippet: Cells were counted after fixation and propidium iodide staining by TALI® cytometry (Life Technologies).

    Techniques: Incubation, Immunofluorescence, Expressing, Marker, Fluorescence, Western Blot, SDS Page

    Ectopic expression of chimeric Myc-ER protein prevents TGF-β1–mediated inhibition of FasL mRNA and subsequent AICD. (A) Functional Myc-ER interferes with the inhibitory effect of TGF-β1 on activation-induced FasL expression in A1.1 cells. A1.1 or A1.1 Myc-ER cells exposed to 4-OHT (50 nM) were first preincubated for 4 h with the drug, which was then also present during subsequent culture. Cells were then incubated in medium alone (lane 1) or activated for 4 h with anti-CD3 antibodies (lanes 2, 3, and 4). 1 ng/ml TGF-β1 (lane 3) or 100 ng/ ml CsA (lane 4) was added to some cultures. FasL expression was then assessed by RT-PCR. (B) Ectopic expression of a chimeric Myc-ER protein prevents AICD after TGF-β1 treatment. A1.1 or A1.1 Myc-ER T hybridoma cells were first preincubated for 4 h in the presence or absence of 4-OHT (50 nM) and then activated for 16 h with anti-CD3 antibodies with the indicated concentrations of TGF-β1 (1 ng/ml) or CsA (100 ng/ml). Cell death was assessed by propidium iodide uptake using a FACScan ® .

    Journal: The Journal of Experimental Medicine

    Article Title: Transforming Growth Factor ?1 Inhibits Fas Ligand Expression and Subsequent Activation-induced Cell Death in T Cells via Downregulation of c-Myc

    doi:

    Figure Lengend Snippet: Ectopic expression of chimeric Myc-ER protein prevents TGF-β1–mediated inhibition of FasL mRNA and subsequent AICD. (A) Functional Myc-ER interferes with the inhibitory effect of TGF-β1 on activation-induced FasL expression in A1.1 cells. A1.1 or A1.1 Myc-ER cells exposed to 4-OHT (50 nM) were first preincubated for 4 h with the drug, which was then also present during subsequent culture. Cells were then incubated in medium alone (lane 1) or activated for 4 h with anti-CD3 antibodies (lanes 2, 3, and 4). 1 ng/ml TGF-β1 (lane 3) or 100 ng/ ml CsA (lane 4) was added to some cultures. FasL expression was then assessed by RT-PCR. (B) Ectopic expression of a chimeric Myc-ER protein prevents AICD after TGF-β1 treatment. A1.1 or A1.1 Myc-ER T hybridoma cells were first preincubated for 4 h in the presence or absence of 4-OHT (50 nM) and then activated for 16 h with anti-CD3 antibodies with the indicated concentrations of TGF-β1 (1 ng/ml) or CsA (100 ng/ml). Cell death was assessed by propidium iodide uptake using a FACScan ® .

    Article Snippet: PBMCs (106 /ml) were activated for 6 d with 100 ng/ml OKT3, and after elimination of dead cells, were restimulated with PMA (50 ng/ml) and ionomycin (1 μg/ml) for 16 h. Viability was assessed by addition of 5 μg/ml propidium iodide and immediate analysis using a FACScan® ( Becton Dickinson ).

    Techniques: Expressing, Inhibition, Functional Assay, Activation Assay, Incubation, Reverse Transcription Polymerase Chain Reaction

    TGF-β1 as well as AS c-myc oligonucleotides do not induce perturbation of the cell cycle in A1.1 T cell hybridomas. 10 6 A1.1 cells were untreated or activated with anti-CD3 in the presence of 5 μM AS c-myc or NS c-myc oligonucleotides added 4 h before activation or with TGF-β1 (10 ng/ml) added 1 h before activation. Cells were harvested at the indicated times, and cell cycle analysis was performed by propidium iodide staining after permeabilization with Triton X-100. The percentage of cells in G0/G1 (A), S (B), and G2/M (C) phase of the cell cycle was determined for each condition (data shown are means ± SD, n = 3).

    Journal: The Journal of Experimental Medicine

    Article Title: Transforming Growth Factor ?1 Inhibits Fas Ligand Expression and Subsequent Activation-induced Cell Death in T Cells via Downregulation of c-Myc

    doi:

    Figure Lengend Snippet: TGF-β1 as well as AS c-myc oligonucleotides do not induce perturbation of the cell cycle in A1.1 T cell hybridomas. 10 6 A1.1 cells were untreated or activated with anti-CD3 in the presence of 5 μM AS c-myc or NS c-myc oligonucleotides added 4 h before activation or with TGF-β1 (10 ng/ml) added 1 h before activation. Cells were harvested at the indicated times, and cell cycle analysis was performed by propidium iodide staining after permeabilization with Triton X-100. The percentage of cells in G0/G1 (A), S (B), and G2/M (C) phase of the cell cycle was determined for each condition (data shown are means ± SD, n = 3).

    Article Snippet: PBMCs (106 /ml) were activated for 6 d with 100 ng/ml OKT3, and after elimination of dead cells, were restimulated with PMA (50 ng/ml) and ionomycin (1 μg/ml) for 16 h. Viability was assessed by addition of 5 μg/ml propidium iodide and immediate analysis using a FACScan® ( Becton Dickinson ).

    Techniques: Activation Assay, Cell Cycle Assay, Staining

    TGF-β1 does not block recombinant soluble FasL– induced apoptosis. A1.1 T cell hybridomas were preincubated for 1 h in the presence of TGF-β1 (10 ng/ml) and then treated with different doses of soluble recombinant human FasL (sFasL). 15 min after addition of soluble FasL, anti-Flag M2 antibody (1 μg/ml) was added to cross-link soluble FasL. After 12 h incubation, percentage of apoptosis was assessed by propidium iodide uptake and analyzed using a FACScan ® .

    Journal: The Journal of Experimental Medicine

    Article Title: Transforming Growth Factor ?1 Inhibits Fas Ligand Expression and Subsequent Activation-induced Cell Death in T Cells via Downregulation of c-Myc

    doi:

    Figure Lengend Snippet: TGF-β1 does not block recombinant soluble FasL– induced apoptosis. A1.1 T cell hybridomas were preincubated for 1 h in the presence of TGF-β1 (10 ng/ml) and then treated with different doses of soluble recombinant human FasL (sFasL). 15 min after addition of soluble FasL, anti-Flag M2 antibody (1 μg/ml) was added to cross-link soluble FasL. After 12 h incubation, percentage of apoptosis was assessed by propidium iodide uptake and analyzed using a FACScan ® .

    Article Snippet: PBMCs (106 /ml) were activated for 6 d with 100 ng/ml OKT3, and after elimination of dead cells, were restimulated with PMA (50 ng/ml) and ionomycin (1 μg/ml) for 16 h. Viability was assessed by addition of 5 μg/ml propidium iodide and immediate analysis using a FACScan® ( Becton Dickinson ).

    Techniques: Blocking Assay, Recombinant, Incubation

    Ectopic expression of chimeric Myc-ER protein prevents TGF-β1–mediated inhibition of AICD in A1.1 T cell hybridomas. (A) Expression of Myc-ER fusion protein in A1.1 Myc-ER clones. Total cell extracts were prepared from three different A1.1 Myc-ER clones. Samples were analyzed by immunoblot analysis using anti–human c-Myc antibody. (B) Ectopic expression of a chimeric Myc-ER protein prevents AICD after TGF-β1 treatment. A1.1 or A1.1 Myc-ER clones were activated for 16 h with anti-CD3 antibodies with the indicated concentrations of TGF-β1 (1 ng/ml) in the presence or absence of 4-OHT (50 nM) added at time 0 of the activation. Cell death was assessed by propidium iodide uptake using a FACScan ® .

    Journal: The Journal of Experimental Medicine

    Article Title: Transforming Growth Factor ?1 Inhibits Fas Ligand Expression and Subsequent Activation-induced Cell Death in T Cells via Downregulation of c-Myc

    doi:

    Figure Lengend Snippet: Ectopic expression of chimeric Myc-ER protein prevents TGF-β1–mediated inhibition of AICD in A1.1 T cell hybridomas. (A) Expression of Myc-ER fusion protein in A1.1 Myc-ER clones. Total cell extracts were prepared from three different A1.1 Myc-ER clones. Samples were analyzed by immunoblot analysis using anti–human c-Myc antibody. (B) Ectopic expression of a chimeric Myc-ER protein prevents AICD after TGF-β1 treatment. A1.1 or A1.1 Myc-ER clones were activated for 16 h with anti-CD3 antibodies with the indicated concentrations of TGF-β1 (1 ng/ml) in the presence or absence of 4-OHT (50 nM) added at time 0 of the activation. Cell death was assessed by propidium iodide uptake using a FACScan ® .

    Article Snippet: PBMCs (106 /ml) were activated for 6 d with 100 ng/ml OKT3, and after elimination of dead cells, were restimulated with PMA (50 ng/ml) and ionomycin (1 μg/ml) for 16 h. Viability was assessed by addition of 5 μg/ml propidium iodide and immediate analysis using a FACScan® ( Becton Dickinson ).

    Techniques: Expressing, Inhibition, Clone Assay, Activation Assay

    TGF-β1 decreases AICD in human peripheral blood T cells and murine T cell hybridomas. (A) PBMCs were activated for 6 d with OKT3 (100 ng/ml) and then restimulated with PMA (50 ng/ml) and ionomycin (1 μg/ml) (P+I) for 16 h with the indicated concentration of TGF-β1 or CsA (100 ng/ml). Viability was assessed by propidium iodide uptake and analyzed using a FACScan ® . Apoptosis was confirmed by morphological assessment after staining with Hoechst 33342 at 10 μg/ml (not shown). (B) A1.1 or 2B4.11 T hybridoma cells were left unactivated (open symbols) or were activated (filled symbols) for 16 h with anti-CD3 antibody (2C11) in the presence of the indicated concentration of TGF-β1. Viability was assessed as in A. (C) A1.1 T hybridoma cells were cultured with medium alone or activated for 12 h with anti-CD3 antibody in the presence of the indicated concentrations of TGF-β1 (ng/ml) or CsA (100 ng/ml). DNA fragmentation associated with apoptosis was assessed by agarose gel electrophoresis.

    Journal: The Journal of Experimental Medicine

    Article Title: Transforming Growth Factor ?1 Inhibits Fas Ligand Expression and Subsequent Activation-induced Cell Death in T Cells via Downregulation of c-Myc

    doi:

    Figure Lengend Snippet: TGF-β1 decreases AICD in human peripheral blood T cells and murine T cell hybridomas. (A) PBMCs were activated for 6 d with OKT3 (100 ng/ml) and then restimulated with PMA (50 ng/ml) and ionomycin (1 μg/ml) (P+I) for 16 h with the indicated concentration of TGF-β1 or CsA (100 ng/ml). Viability was assessed by propidium iodide uptake and analyzed using a FACScan ® . Apoptosis was confirmed by morphological assessment after staining with Hoechst 33342 at 10 μg/ml (not shown). (B) A1.1 or 2B4.11 T hybridoma cells were left unactivated (open symbols) or were activated (filled symbols) for 16 h with anti-CD3 antibody (2C11) in the presence of the indicated concentration of TGF-β1. Viability was assessed as in A. (C) A1.1 T hybridoma cells were cultured with medium alone or activated for 12 h with anti-CD3 antibody in the presence of the indicated concentrations of TGF-β1 (ng/ml) or CsA (100 ng/ml). DNA fragmentation associated with apoptosis was assessed by agarose gel electrophoresis.

    Article Snippet: PBMCs (106 /ml) were activated for 6 d with 100 ng/ml OKT3, and after elimination of dead cells, were restimulated with PMA (50 ng/ml) and ionomycin (1 μg/ml) for 16 h. Viability was assessed by addition of 5 μg/ml propidium iodide and immediate analysis using a FACScan® ( Becton Dickinson ).

    Techniques: Concentration Assay, Staining, Cell Culture, Agarose Gel Electrophoresis

    Phosphatidylinositol 3-kinase/Akt acts downstream of FAK signaling to regulate apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were treated with LY294002. (A and B) The expression of Akt, pAkt, caspase-3 and c-caspase-3 was examined using western blotting. (C and D) Cell apoptosis was examined using (C) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (D) Annexin V/propidium iodide. (E and F) The expression of FAK and pFAK was examined using western blotting. Scale bar, 200 µm. FAK, focal adhesion kinase; pFAK, phosphorylated FAK; c-caspase-3, cleaved caspase-3; TGFβ, transforming growth factor-β.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Inhibition of focal adhesion kinase induces apoptosis in bladder cancer cells via Src and the phosphatidylinositol 3-kinase/Akt pathway

    doi: 10.3892/etm.2015.2745

    Figure Lengend Snippet: Phosphatidylinositol 3-kinase/Akt acts downstream of FAK signaling to regulate apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were treated with LY294002. (A and B) The expression of Akt, pAkt, caspase-3 and c-caspase-3 was examined using western blotting. (C and D) Cell apoptosis was examined using (C) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (D) Annexin V/propidium iodide. (E and F) The expression of FAK and pFAK was examined using western blotting. Scale bar, 200 µm. FAK, focal adhesion kinase; pFAK, phosphorylated FAK; c-caspase-3, cleaved caspase-3; TGFβ, transforming growth factor-β.

    Article Snippet: The treated cells were stained with FITC, Annexin V and propidium iodide (PI), and the stained cells were analyzed using a FACSort™ flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA) and evaluated with the CellQuest™ software system (BD Biosciences).

    Techniques: Expressing, Western Blot, End Labeling

    Knockdown of FAK induces apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were transfected with small interfering RNA against FAK (siFAK) or control (conRNA). (A and B) The expression of FAK was examined using western blotting. (C and D) Cell apoptosis was examined using (C) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (D) Annexin V/propidium iodide. (E) Cell survival was examined using an MTT assay. Scale bar, 200 µm. *P

    Journal: Experimental and Therapeutic Medicine

    Article Title: Inhibition of focal adhesion kinase induces apoptosis in bladder cancer cells via Src and the phosphatidylinositol 3-kinase/Akt pathway

    doi: 10.3892/etm.2015.2745

    Figure Lengend Snippet: Knockdown of FAK induces apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were transfected with small interfering RNA against FAK (siFAK) or control (conRNA). (A and B) The expression of FAK was examined using western blotting. (C and D) Cell apoptosis was examined using (C) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (D) Annexin V/propidium iodide. (E) Cell survival was examined using an MTT assay. Scale bar, 200 µm. *P

    Article Snippet: The treated cells were stained with FITC, Annexin V and propidium iodide (PI), and the stained cells were analyzed using a FACSort™ flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA) and evaluated with the CellQuest™ software system (BD Biosciences).

    Techniques: Transfection, Small Interfering RNA, Expressing, Western Blot, End Labeling, MTT Assay

    Src is an important mediator of FAK-regulated apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were treated with PP2. (A and B) Cell apoptosis was examined using (A) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (B) Annexin V/propidium iodide. (C and D) The expression of FAK, pFAK, Src and pSrc was examined using western blotting. Scale bar, 200 µm. FAK, focal adhesion kinase; pFAK, phosphorylated FAK; TGFβ, transforming growth factor-β.

    Journal: Experimental and Therapeutic Medicine

    Article Title: Inhibition of focal adhesion kinase induces apoptosis in bladder cancer cells via Src and the phosphatidylinositol 3-kinase/Akt pathway

    doi: 10.3892/etm.2015.2745

    Figure Lengend Snippet: Src is an important mediator of FAK-regulated apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were treated with PP2. (A and B) Cell apoptosis was examined using (A) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (B) Annexin V/propidium iodide. (C and D) The expression of FAK, pFAK, Src and pSrc was examined using western blotting. Scale bar, 200 µm. FAK, focal adhesion kinase; pFAK, phosphorylated FAK; TGFβ, transforming growth factor-β.

    Article Snippet: The treated cells were stained with FITC, Annexin V and propidium iodide (PI), and the stained cells were analyzed using a FACSort™ flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA) and evaluated with the CellQuest™ software system (BD Biosciences).

    Techniques: End Labeling, Expressing, Western Blot

    Suppression of FAK phosphorylation induces apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were treated with PF-228 and 5 ng/ml TGFβ. (A and B) Cell apoptosis was examined using (A) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (B) Annexin V/propidium iodide. (C and D) The expression of FAK, pFAK, Src, pSrc, Akt, pAkt, caspase-3 and c-caspase-3 was examined using western blotting. (E) Cell survival was examined using an MTT assay. Scale bar, 200 µm. *P

    Journal: Experimental and Therapeutic Medicine

    Article Title: Inhibition of focal adhesion kinase induces apoptosis in bladder cancer cells via Src and the phosphatidylinositol 3-kinase/Akt pathway

    doi: 10.3892/etm.2015.2745

    Figure Lengend Snippet: Suppression of FAK phosphorylation induces apoptosis in T24 bladder cancer cells. T24 bladder cancer cells were treated with PF-228 and 5 ng/ml TGFβ. (A and B) Cell apoptosis was examined using (A) deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay and (B) Annexin V/propidium iodide. (C and D) The expression of FAK, pFAK, Src, pSrc, Akt, pAkt, caspase-3 and c-caspase-3 was examined using western blotting. (E) Cell survival was examined using an MTT assay. Scale bar, 200 µm. *P

    Article Snippet: The treated cells were stained with FITC, Annexin V and propidium iodide (PI), and the stained cells were analyzed using a FACSort™ flow cytometer (Becton-Dickinson, Franklin Lakes, NJ, USA) and evaluated with the CellQuest™ software system (BD Biosciences).

    Techniques: End Labeling, Expressing, Western Blot, MTT Assay

    Neutralization of the PM-MSC complex . (a, b) Viability of rat fetal membrane-derived MSCs at 10 min after mixture of PM (final dose 0.8%) with or without neutralization using NaOH in-vitro was assessed by propidium iodide staining. Representative pictures in each group are presented (a). Scale bar = 50 μm. The bar graph presents the averaged data (b). n=4 in each point ; * p

    Journal: Biomaterials

    Article Title: Self-assembling peptide hydrogel enables instant epicardial coating of the heart with mesenchymal stromal cells for the treatment of heart failure

    doi: 10.1016/j.biomaterials.2017.10.050

    Figure Lengend Snippet: Neutralization of the PM-MSC complex . (a, b) Viability of rat fetal membrane-derived MSCs at 10 min after mixture of PM (final dose 0.8%) with or without neutralization using NaOH in-vitro was assessed by propidium iodide staining. Representative pictures in each group are presented (a). Scale bar = 50 μm. The bar graph presents the averaged data (b). n=4 in each point ; * p

    Article Snippet: 5 μl of propidium iodide (PI) solution (Calbiochem, UK) was added to the PM-MSC complex.

    Techniques: Neutralization, Derivative Assay, In Vitro, Staining

    Production and characterization of modified PM . (a) . Appearance and microimage of produced homogenous, neutral PM are shown. Scale bar = 50 μm. (b, c) . Viability of rat fetal membrane-derived MSCs at 10 min after mixture with modified PM (mod-PM; final dose 0.8%) in-vitro was assessed by propidium iodide staining (b). n=4 in each point. Representative pictures of propidium iodide in each group are presented in (c). Scale bar = 50 μm. (d) . Retention of the modified PM-MSC complex was assessed by a tilting test on the plastic plate in-vitro. n=4 in each point. (e) . Retention of the modified PM-MSC complex on the beating rat heart in-vivo. Retained modified PM-MSC complex (white) at 0 min dissociated by 60 min after epicardial placement. Representative pictures from n = 4 are presented.

    Journal: Biomaterials

    Article Title: Self-assembling peptide hydrogel enables instant epicardial coating of the heart with mesenchymal stromal cells for the treatment of heart failure

    doi: 10.1016/j.biomaterials.2017.10.050

    Figure Lengend Snippet: Production and characterization of modified PM . (a) . Appearance and microimage of produced homogenous, neutral PM are shown. Scale bar = 50 μm. (b, c) . Viability of rat fetal membrane-derived MSCs at 10 min after mixture with modified PM (mod-PM; final dose 0.8%) in-vitro was assessed by propidium iodide staining (b). n=4 in each point. Representative pictures of propidium iodide in each group are presented in (c). Scale bar = 50 μm. (d) . Retention of the modified PM-MSC complex was assessed by a tilting test on the plastic plate in-vitro. n=4 in each point. (e) . Retention of the modified PM-MSC complex on the beating rat heart in-vivo. Retained modified PM-MSC complex (white) at 0 min dissociated by 60 min after epicardial placement. Representative pictures from n = 4 are presented.

    Article Snippet: 5 μl of propidium iodide (PI) solution (Calbiochem, UK) was added to the PM-MSC complex.

    Techniques: Modification, Produced, Derivative Assay, In Vitro, Staining, In Vivo

    Baricitinib suppresses CD80/CD86 expression on dendritic cells (DCs). Immature monocyte-derived dendritic cells were cultured with or without baricitinib and tofacitinib during lipopolysaccharide stimulation for 48 h. The DC phenotype was evaluated using flow cytometry. (A) Expression of HLA-DR, CD80, and CD86. (B) Representative histogram data of HLA-DR, CD80, and CD86 expression. (C) Rate of viable cells (annexin V neg /propidium iodide neg ). (D) Expression of CD80 and CD86 in the presence of baricitinib and tofacitinib. Data are mean ± SD of three different donors per group. * p

    Journal: Frontiers in Immunology

    Article Title: Janus Kinase Inhibitor Baricitinib Modulates Human Innate and Adaptive Immune System

    doi: 10.3389/fimmu.2018.01510

    Figure Lengend Snippet: Baricitinib suppresses CD80/CD86 expression on dendritic cells (DCs). Immature monocyte-derived dendritic cells were cultured with or without baricitinib and tofacitinib during lipopolysaccharide stimulation for 48 h. The DC phenotype was evaluated using flow cytometry. (A) Expression of HLA-DR, CD80, and CD86. (B) Representative histogram data of HLA-DR, CD80, and CD86 expression. (C) Rate of viable cells (annexin V neg /propidium iodide neg ). (D) Expression of CD80 and CD86 in the presence of baricitinib and tofacitinib. Data are mean ± SD of three different donors per group. * p

    Article Snippet: Cell viability was evaluated by Annexin V and Propidium Iodide (BioLegend, San Diego, CA, USA).

    Techniques: Expressing, Derivative Assay, Cell Culture, Flow Cytometry, Cytometry

    Baricitinib suppresses type-I interferon (IFN) production from plasmacytoid dendritic cells (pDCs). Peripheral blood mononuclear cells were stimulated with toll-like receptor (TLR) 9 agonist with or without baricitinib and other inhibitors. TLR9-mediated cytokine production by pDCs (Lin − HLA-DR + CD11c − CD123 + cells) was measured by intracellular staining or ELISA. (A) Percentage of TNF-α- and IFN-α-producing pDCs. (B) Representative flow cytometry plots showing IFN-α production by pDCs. (C) IFN-α concentration in the supernatants. (D) Percentage of TNF-α- and IFN-α-producing pDCs in the presence of baricitinib and other inhibitors. (E) Representative histogram data of annexin V/propidium iodide staining from three independent experiments. (F) Expression of CD80 and CD86 on pDCs. Data are mean ± SD of three different donors per group. * p

    Journal: Frontiers in Immunology

    Article Title: Janus Kinase Inhibitor Baricitinib Modulates Human Innate and Adaptive Immune System

    doi: 10.3389/fimmu.2018.01510

    Figure Lengend Snippet: Baricitinib suppresses type-I interferon (IFN) production from plasmacytoid dendritic cells (pDCs). Peripheral blood mononuclear cells were stimulated with toll-like receptor (TLR) 9 agonist with or without baricitinib and other inhibitors. TLR9-mediated cytokine production by pDCs (Lin − HLA-DR + CD11c − CD123 + cells) was measured by intracellular staining or ELISA. (A) Percentage of TNF-α- and IFN-α-producing pDCs. (B) Representative flow cytometry plots showing IFN-α production by pDCs. (C) IFN-α concentration in the supernatants. (D) Percentage of TNF-α- and IFN-α-producing pDCs in the presence of baricitinib and other inhibitors. (E) Representative histogram data of annexin V/propidium iodide staining from three independent experiments. (F) Expression of CD80 and CD86 on pDCs. Data are mean ± SD of three different donors per group. * p

    Article Snippet: Cell viability was evaluated by Annexin V and Propidium Iodide (BioLegend, San Diego, CA, USA).

    Techniques: Staining, Enzyme-linked Immunosorbent Assay, Flow Cytometry, Cytometry, Concentration Assay, Expressing

    Baricitinib inhibits B cell differentiation. Human B cells were purified and cultured for 5 days in the presence of anti-human IgM Ab F(ab′)2 fragments and under interferon (IFN)-α stimulation. Plasmablast (CD3 − CD19 + CD20 − CD27 hi CD38 hi ) differentiation was assessed by flow cytometry, and IgG Ab titers and interleukin (IL)-6 concentrations in culture supernatants were measured by ELISA Cytometric Bead Array. (A) Percentage of plasmablasts. (B) Representative flow cytometry plots showing plasmablasts. (C) Percentage of plasmablast, IL-6, and IgG concentrations in the presence of baricitinib and other inhibitors. (D) Representative histogram data of annexin V/propidium iodide staining. (E) Rate of viable cells (annexin V neg /propidium iodide neg ). Data are mean ± SD of three different donors per group. * p

    Journal: Frontiers in Immunology

    Article Title: Janus Kinase Inhibitor Baricitinib Modulates Human Innate and Adaptive Immune System

    doi: 10.3389/fimmu.2018.01510

    Figure Lengend Snippet: Baricitinib inhibits B cell differentiation. Human B cells were purified and cultured for 5 days in the presence of anti-human IgM Ab F(ab′)2 fragments and under interferon (IFN)-α stimulation. Plasmablast (CD3 − CD19 + CD20 − CD27 hi CD38 hi ) differentiation was assessed by flow cytometry, and IgG Ab titers and interleukin (IL)-6 concentrations in culture supernatants were measured by ELISA Cytometric Bead Array. (A) Percentage of plasmablasts. (B) Representative flow cytometry plots showing plasmablasts. (C) Percentage of plasmablast, IL-6, and IgG concentrations in the presence of baricitinib and other inhibitors. (D) Representative histogram data of annexin V/propidium iodide staining. (E) Rate of viable cells (annexin V neg /propidium iodide neg ). Data are mean ± SD of three different donors per group. * p

    Article Snippet: Cell viability was evaluated by Annexin V and Propidium Iodide (BioLegend, San Diego, CA, USA).

    Techniques: Cell Differentiation, Purification, Cell Culture, Flow Cytometry, Cytometry, Enzyme-linked Immunosorbent Assay, Staining

    Radioresistant TE-1 and Eca-109 cells showed more aggressive malignancies. (A) Wound-healing assay of TE-1 and TE-1/R cells. (B) Wound-healing assay of Eca-109 and Eca-109/R cells. Wound healing was observed 24 h after the treatment. (C) Induction of apoptosis by radiation in TE-1 and TE-1/R cells. (D) Induction of apoptosis by radiation in Eca-109 and Eca-109/R cells. Cells were treated with 6 Gy irradiation, and the apoptosis was measured using propidium iodide (PI)/Annexin-V double staining. Data are normalized to the control cells and presented as the mean ± SEM of three independent experiments, * P

    Journal: Journal of Cancer

    Article Title: mRNA and methylation profiling of radioresistant esophageal cancer cells: the involvement of Sall2 in acquired aggressive phenotypes

    doi: 10.7150/jca.15652

    Figure Lengend Snippet: Radioresistant TE-1 and Eca-109 cells showed more aggressive malignancies. (A) Wound-healing assay of TE-1 and TE-1/R cells. (B) Wound-healing assay of Eca-109 and Eca-109/R cells. Wound healing was observed 24 h after the treatment. (C) Induction of apoptosis by radiation in TE-1 and TE-1/R cells. (D) Induction of apoptosis by radiation in Eca-109 and Eca-109/R cells. Cells were treated with 6 Gy irradiation, and the apoptosis was measured using propidium iodide (PI)/Annexin-V double staining. Data are normalized to the control cells and presented as the mean ± SEM of three independent experiments, * P

    Article Snippet: Apoptosis was measured using propidium iodide (PI)/Annexin-V double staining following the manufacturer's instructions (Keygen Biotech, Nanjing, China).

    Techniques: Wound Healing Assay, Irradiation, Double Staining

    The effect of Sall2 overexpression on the apoptosis, migration and cisplatin sensitivity of esophageal cancer cells. (A) Cells were transiently transfected with control vector or Sall2-overexpression vector (pcDNA3.1-Sall2). The expression of Sall2 was validated by Western blot. Apoptosis of (B) TE-1/R and (C) Eca-109/R cells was measured using propidium iodide (PI)/Annexin-V double staining. (D) Wound-healing assay of cells after the indicated transfection in TE-1/R cells. (E) Wound-healing assay of Eca-109/R cells after the indicated transfection. Wound-healing was observed 24 h after the treatment. (F) TE-1 and (G) Eca-109 cells were treated with cisplatin for 24 h. MTT assay of cell viability. The data are shown as the mean ± SEM for three independent experiments. Statistical analysis between the groups was determined by ANOVA; * P

    Journal: Journal of Cancer

    Article Title: mRNA and methylation profiling of radioresistant esophageal cancer cells: the involvement of Sall2 in acquired aggressive phenotypes

    doi: 10.7150/jca.15652

    Figure Lengend Snippet: The effect of Sall2 overexpression on the apoptosis, migration and cisplatin sensitivity of esophageal cancer cells. (A) Cells were transiently transfected with control vector or Sall2-overexpression vector (pcDNA3.1-Sall2). The expression of Sall2 was validated by Western blot. Apoptosis of (B) TE-1/R and (C) Eca-109/R cells was measured using propidium iodide (PI)/Annexin-V double staining. (D) Wound-healing assay of cells after the indicated transfection in TE-1/R cells. (E) Wound-healing assay of Eca-109/R cells after the indicated transfection. Wound-healing was observed 24 h after the treatment. (F) TE-1 and (G) Eca-109 cells were treated with cisplatin for 24 h. MTT assay of cell viability. The data are shown as the mean ± SEM for three independent experiments. Statistical analysis between the groups was determined by ANOVA; * P

    Article Snippet: Apoptosis was measured using propidium iodide (PI)/Annexin-V double staining following the manufacturer's instructions (Keygen Biotech, Nanjing, China).

    Techniques: Over Expression, Migration, Transfection, Plasmid Preparation, Expressing, Western Blot, Double Staining, Wound Healing Assay, MTT Assay

    PCB-induced arrest is reversible and cells regain sensitivity to VCR after reentering the cell cycle. A . ALL-5 and T98G cells were stained with propidium iodide and subjected to two-parameter flow cytometry to create a gate distinguishing EdU-negative and EdU-positive cells (left panels). ALL-5 and T98G cells were then treated with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G), washed thrice with PBS, and incubated in growth media containing 10 µM EdU for the times indicated. Cells were harvested, fixed, and stained for EdU incorporation and with propidium iodide and analyzed by two-parameter flow cytometry. Data shown are representative of three independent experiments. The percentage of cells positive for EdU is indicated at the top of each panel. B . ALL-5 and T98G cells were treated with 0.1% DMSO or 100 nM (ALL-5) or 1 µM (T98G) VCR for 48 h with or without pretreatment with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G). A third set of cells were pretreated with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G), after which cells were washed thrice with PBS and treated with 0.1% DMSO or 100 nM (ALL-5) or 1 µM (T98G) VCR for 48 h. Cells were stained with propidium iodide and subjected to flow cytometry to determine the percentage of cells with

    Journal: Cell Cycle

    Article Title: Microtubules play an essential role in the survival of primary acute lymphoblastic leukemia cells advancing through G1 phase

    doi: 10.1080/15384101.2018.1496746

    Figure Lengend Snippet: PCB-induced arrest is reversible and cells regain sensitivity to VCR after reentering the cell cycle. A . ALL-5 and T98G cells were stained with propidium iodide and subjected to two-parameter flow cytometry to create a gate distinguishing EdU-negative and EdU-positive cells (left panels). ALL-5 and T98G cells were then treated with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G), washed thrice with PBS, and incubated in growth media containing 10 µM EdU for the times indicated. Cells were harvested, fixed, and stained for EdU incorporation and with propidium iodide and analyzed by two-parameter flow cytometry. Data shown are representative of three independent experiments. The percentage of cells positive for EdU is indicated at the top of each panel. B . ALL-5 and T98G cells were treated with 0.1% DMSO or 100 nM (ALL-5) or 1 µM (T98G) VCR for 48 h with or without pretreatment with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G). A third set of cells were pretreated with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G), after which cells were washed thrice with PBS and treated with 0.1% DMSO or 100 nM (ALL-5) or 1 µM (T98G) VCR for 48 h. Cells were stained with propidium iodide and subjected to flow cytometry to determine the percentage of cells with

    Article Snippet: DNA content was evaluated by propidium iodide staining according to the manufacturer’s instructions (BD Pharmigen) and flow cytometry was carried out by the UAMS Flow Cytometry Core Facility using a FACSCalibur (Becton Dickinson, Mountain View, CA).

    Techniques: Staining, Flow Cytometry, Cytometry, Incubation

    Vinblastine induces death in G1 phase which is prevented by PCB pretreatment. A . ALL-5 cells in G1 phase were isolated using centrifugal elutriation and treated with 0.1% DMSO or 100 nM vinblastine (VBL) for the times indicated. Cells were fixed and stained with propidium iodide and DNA content analyzed by flow cytometry. Data shown are representative of two independent experiments of three replicates each. B . ALL-5 cells were treated with 0.1% DMSO or 100 nM VBL for 48 h with or without pretreatment with 1 µM PCB for 72 h. Cells were fixed and stained with propidium iodide and subjected to flow cytometry to determine the percentage of cells with

    Journal: Cell Cycle

    Article Title: Microtubules play an essential role in the survival of primary acute lymphoblastic leukemia cells advancing through G1 phase

    doi: 10.1080/15384101.2018.1496746

    Figure Lengend Snippet: Vinblastine induces death in G1 phase which is prevented by PCB pretreatment. A . ALL-5 cells in G1 phase were isolated using centrifugal elutriation and treated with 0.1% DMSO or 100 nM vinblastine (VBL) for the times indicated. Cells were fixed and stained with propidium iodide and DNA content analyzed by flow cytometry. Data shown are representative of two independent experiments of three replicates each. B . ALL-5 cells were treated with 0.1% DMSO or 100 nM VBL for 48 h with or without pretreatment with 1 µM PCB for 72 h. Cells were fixed and stained with propidium iodide and subjected to flow cytometry to determine the percentage of cells with

    Article Snippet: DNA content was evaluated by propidium iodide staining according to the manufacturer’s instructions (BD Pharmigen) and flow cytometry was carried out by the UAMS Flow Cytometry Core Facility using a FACSCalibur (Becton Dickinson, Mountain View, CA).

    Techniques: Isolation, Staining, Flow Cytometry, Cytometry

    Cycloheximide treated ALL cells arrest in G1 phase and become insensitive to VCR. A,B . ALL-5 cells isolated in G1 phase by centrifugal elutriation were treated with 1 µg/mL cycloheximide (CHX) or 100 nM VCR or both and stained with propidium iodide and analyzed for DNA content by flow cytometry. The percentage of cells with 2N DNA (G1 phase) ( A ) or sub-G1 DNA ( B ) are shown (mean ± S.D., n = 3). C . Whole cell extracts were prepared from G1 phase ALL-5 cells treated with 1 µM PCB or 1 µg/mL CHX for the times indicated and subjected to immunoblot analysis for RB, phospho-RB (Ser 807/811), cyclin D, or GAPDH as a loading control.

    Journal: Cell Cycle

    Article Title: Microtubules play an essential role in the survival of primary acute lymphoblastic leukemia cells advancing through G1 phase

    doi: 10.1080/15384101.2018.1496746

    Figure Lengend Snippet: Cycloheximide treated ALL cells arrest in G1 phase and become insensitive to VCR. A,B . ALL-5 cells isolated in G1 phase by centrifugal elutriation were treated with 1 µg/mL cycloheximide (CHX) or 100 nM VCR or both and stained with propidium iodide and analyzed for DNA content by flow cytometry. The percentage of cells with 2N DNA (G1 phase) ( A ) or sub-G1 DNA ( B ) are shown (mean ± S.D., n = 3). C . Whole cell extracts were prepared from G1 phase ALL-5 cells treated with 1 µM PCB or 1 µg/mL CHX for the times indicated and subjected to immunoblot analysis for RB, phospho-RB (Ser 807/811), cyclin D, or GAPDH as a loading control.

    Article Snippet: DNA content was evaluated by propidium iodide staining according to the manufacturer’s instructions (BD Pharmigen) and flow cytometry was carried out by the UAMS Flow Cytometry Core Facility using a FACSCalibur (Becton Dickinson, Mountain View, CA).

    Techniques: Isolation, Staining, Flow Cytometry, Cytometry

    PCB causes G1 phase arrest in ALL-5 and T98G but not HeLa cells. A . Cells were treated with vehicle (0.1% DMSO) or 1 µM PCB for 72 h (ALL-5) or 48 h (T98G and HeLa) and DNA content determined by propidium iodide staining and flow cytometry. Data shown are mean ± S.D. (n = 4). B . ALL-5 or T98G cells were treated with 1 μM PCB for 72 h or 48 h, respectively. Cells were fixed and stained for Ki-67 (red) or with DAPI (blue) as a nuclear marker. The scale bar in ALL-5 images is 60 μm while that in T98G images is 120 μm. Images are representative of six fields of vision. C . Mean intensity fluorescence of Ki-67 was quantified for 20 cells per six fields of vision using ImageJ. Cells were selected using DAPI stain while blind to Ki-67 intensity. Each point represents a single cell while horizontal bars represent the mean ± S.D. of all data points (n = 120).

    Journal: Cell Cycle

    Article Title: Microtubules play an essential role in the survival of primary acute lymphoblastic leukemia cells advancing through G1 phase

    doi: 10.1080/15384101.2018.1496746

    Figure Lengend Snippet: PCB causes G1 phase arrest in ALL-5 and T98G but not HeLa cells. A . Cells were treated with vehicle (0.1% DMSO) or 1 µM PCB for 72 h (ALL-5) or 48 h (T98G and HeLa) and DNA content determined by propidium iodide staining and flow cytometry. Data shown are mean ± S.D. (n = 4). B . ALL-5 or T98G cells were treated with 1 μM PCB for 72 h or 48 h, respectively. Cells were fixed and stained for Ki-67 (red) or with DAPI (blue) as a nuclear marker. The scale bar in ALL-5 images is 60 μm while that in T98G images is 120 μm. Images are representative of six fields of vision. C . Mean intensity fluorescence of Ki-67 was quantified for 20 cells per six fields of vision using ImageJ. Cells were selected using DAPI stain while blind to Ki-67 intensity. Each point represents a single cell while horizontal bars represent the mean ± S.D. of all data points (n = 120).

    Article Snippet: DNA content was evaluated by propidium iodide staining according to the manufacturer’s instructions (BD Pharmigen) and flow cytometry was carried out by the UAMS Flow Cytometry Core Facility using a FACSCalibur (Becton Dickinson, Mountain View, CA).

    Techniques: Staining, Flow Cytometry, Cytometry, Marker, Fluorescence

    ALL-5 and T98G cells are refractory to VCR after PCB pretreatment while HeLa cells retain their sensitivity. A . ALL-5, T98G, or HeLa cells were treated with 0.1% DMSO or 100 nM (ALL-5 and HeLa) or 1 µM (T98G cells) VCR for 48 h with or without pretreatment with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G and HeLa). Cells were subjected to propidium iodide staining and flow cytometry to determine the percentage of cells with

    Journal: Cell Cycle

    Article Title: Microtubules play an essential role in the survival of primary acute lymphoblastic leukemia cells advancing through G1 phase

    doi: 10.1080/15384101.2018.1496746

    Figure Lengend Snippet: ALL-5 and T98G cells are refractory to VCR after PCB pretreatment while HeLa cells retain their sensitivity. A . ALL-5, T98G, or HeLa cells were treated with 0.1% DMSO or 100 nM (ALL-5 and HeLa) or 1 µM (T98G cells) VCR for 48 h with or without pretreatment with 1 µM PCB for 72 h (ALL-5) or 48 h (T98G and HeLa). Cells were subjected to propidium iodide staining and flow cytometry to determine the percentage of cells with

    Article Snippet: DNA content was evaluated by propidium iodide staining according to the manufacturer’s instructions (BD Pharmigen) and flow cytometry was carried out by the UAMS Flow Cytometry Core Facility using a FACSCalibur (Becton Dickinson, Mountain View, CA).

    Techniques: Staining, Flow Cytometry, Cytometry

    Pretreatment with PCB does not prevent VCR from disrupting the microtubule network or block downstream apoptotic signaling. A . HeLa or ALL-5 cells were treated with 0.1% DMSO or 100 nM vincristine (VCR) for 14 h, with or without pretreatment with 1 µM PCB for 72 h, as indicated. Cells were fixed and permeabilized and α-tubulin visualized by fluorescence microscopy. B . ALL-5 cells were treated with 0.1% DMSO or 100 nM ABT-263 for 16 h with or without pretreatment with 1 µM PCB for 72 h. Cells were fixed and stained with propidium iodide and subjected to flow cytometry to determine the percentage of cells with

    Journal: Cell Cycle

    Article Title: Microtubules play an essential role in the survival of primary acute lymphoblastic leukemia cells advancing through G1 phase

    doi: 10.1080/15384101.2018.1496746

    Figure Lengend Snippet: Pretreatment with PCB does not prevent VCR from disrupting the microtubule network or block downstream apoptotic signaling. A . HeLa or ALL-5 cells were treated with 0.1% DMSO or 100 nM vincristine (VCR) for 14 h, with or without pretreatment with 1 µM PCB for 72 h, as indicated. Cells were fixed and permeabilized and α-tubulin visualized by fluorescence microscopy. B . ALL-5 cells were treated with 0.1% DMSO or 100 nM ABT-263 for 16 h with or without pretreatment with 1 µM PCB for 72 h. Cells were fixed and stained with propidium iodide and subjected to flow cytometry to determine the percentage of cells with

    Article Snippet: DNA content was evaluated by propidium iodide staining according to the manufacturer’s instructions (BD Pharmigen) and flow cytometry was carried out by the UAMS Flow Cytometry Core Facility using a FACSCalibur (Becton Dickinson, Mountain View, CA).

    Techniques: Blocking Assay, Fluorescence, Microscopy, Staining, Flow Cytometry, Cytometry

    The effect of oridonin nanosuspension and free oridonin on PANC-1 cell apoptosis was measured by annexin V-fluorescein isothiocyanate/propidium iodide staining. The early apoptotic cells stained by annexin-V-fluorescein isothiocyanate are located in the lower right quadrant. Abbreviation: ORI, oridonin.

    Journal: International Journal of Nanomedicine

    Article Title: Oridonin nanosuspension was more effective than free oridonin on G2/M cell cycle arrest and apoptosis in the human pancreatic cancer PANC-1 cell line

    doi: 10.2147/IJN.S29483

    Figure Lengend Snippet: The effect of oridonin nanosuspension and free oridonin on PANC-1 cell apoptosis was measured by annexin V-fluorescein isothiocyanate/propidium iodide staining. The early apoptotic cells stained by annexin-V-fluorescein isothiocyanate are located in the lower right quadrant. Abbreviation: ORI, oridonin.

    Article Snippet: Propidium iodide (PI), ribonuclease A, and annexin V-FITC were purchased from KeyGen Biotechnology ( Nanjing, China).

    Techniques: Staining

    Oridonin nanosuspension-induced and free oridonin-induced morphologic changes of PANC-1 cells. ( A ) Cellular morphology was examined in the presence of different doses of oridonin nanosuspension and free oridonin. Nuclear morphology was determined using ( B ) propidium iodide staining and ( C ) Hoechst 33342 staining. Abbreviation: ORI, oridonin.

    Journal: International Journal of Nanomedicine

    Article Title: Oridonin nanosuspension was more effective than free oridonin on G2/M cell cycle arrest and apoptosis in the human pancreatic cancer PANC-1 cell line

    doi: 10.2147/IJN.S29483

    Figure Lengend Snippet: Oridonin nanosuspension-induced and free oridonin-induced morphologic changes of PANC-1 cells. ( A ) Cellular morphology was examined in the presence of different doses of oridonin nanosuspension and free oridonin. Nuclear morphology was determined using ( B ) propidium iodide staining and ( C ) Hoechst 33342 staining. Abbreviation: ORI, oridonin.

    Article Snippet: Propidium iodide (PI), ribonuclease A, and annexin V-FITC were purchased from KeyGen Biotechnology ( Nanjing, China).

    Techniques: Staining

    Effects of CD54 Knockdown on Cell Proliferation and Apoptosis. ( a ) BrdU analysis of cell cycle progression in prostate cancer patient cells transfected with shCtrl or shCD54-1. ( b ) Ki67 analysis of cell cycle progression in prostate cancer patient cells transfected with shCtrl or shCD54-1. ( c ) FACS analysis of propidium iodide- and annexin V-stained prostate cancer patient cells (from two different patients) transfected with shCtrl or shCD54-1. ( d ) Representative light micrograph fields and comparative quantification of the migratory capacity of prostate cancer patient cells (from two different patients) transfected with shCtrl or shCD54-1 via Transwell assay. ( e ) Representative microscope fields and comparative quantification of apoptosis in vehicle-treated or cisplatin (DDP)-treated PC3 cells transfected with shCtrl or shCD54-1. Arrows indicate apoptotic cells. ( f ) The cisplatin (DDP) IC 50 values of 5 prostate cancer patient cell lines transfected with shCtrl or shCD54-1. * P

    Journal: Theranostics

    Article Title: CD54-NOTCH1 axis controls tumor initiation and cancer stem cell functions in human prostate cancer

    doi: 10.7150/thno.16752

    Figure Lengend Snippet: Effects of CD54 Knockdown on Cell Proliferation and Apoptosis. ( a ) BrdU analysis of cell cycle progression in prostate cancer patient cells transfected with shCtrl or shCD54-1. ( b ) Ki67 analysis of cell cycle progression in prostate cancer patient cells transfected with shCtrl or shCD54-1. ( c ) FACS analysis of propidium iodide- and annexin V-stained prostate cancer patient cells (from two different patients) transfected with shCtrl or shCD54-1. ( d ) Representative light micrograph fields and comparative quantification of the migratory capacity of prostate cancer patient cells (from two different patients) transfected with shCtrl or shCD54-1 via Transwell assay. ( e ) Representative microscope fields and comparative quantification of apoptosis in vehicle-treated or cisplatin (DDP)-treated PC3 cells transfected with shCtrl or shCD54-1. Arrows indicate apoptotic cells. ( f ) The cisplatin (DDP) IC 50 values of 5 prostate cancer patient cell lines transfected with shCtrl or shCD54-1. * P

    Article Snippet: Cells were stained with annexin V-FITC and propidium iodide (BioVision, Milpitas, CA, USA) and incubated for 10 min at room temperature in the dark.

    Techniques: Transfection, FACS, Staining, Transwell Assay, Microscopy