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Becton Dickinson annexin v apoptosis detection kit
GIMAP6 knockdown accelerates the PMA/ionomycin induction of T cell activation. A , Jurkat-derived cell lines ( Ctl , KD1 , and KD2 ) were treated with either PMA/ionomycin or an equal concentration of DMSO to determine the effect of GIMAP6 on T cell activation. To monitor the rate of cell <t>apoptosis/death</t> and T cell activation, PMA/ionomycin-treated cells were harvested at 3, 8, and 24 h and stained for the phosphatidylserine exposure marker Annexin V-FITC and for the T cell activation surface marker CD69-PE. The results were analyzed and quantified by FlowJo 7.6.1. B , the effect of GIMAP6 expression on PMA/ionomycin-induced cell death. All AnV+ cells were taken as apoptotic/dead cells, and the remaining AnV− cells were taken as live cells. C , the effect of GIMAP6 expression on PMA/ionomycin-induced T cell activation. To discriminate activated T cells from inactivated T cells across all live cells, the surface marker CD69 was used to calculate the percentage of CD69+/AnV−. D , the effect of GIMAP6 expression on PMA/ionomycin-induced IL-2 secretion. The data represent the average of at least two independent experiments with triplicate measurements (mean ± S.D.). P/I indicates that the cells were treated with both PMA (10 ng/ml) and ionomycin (1 μg/ml).
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1) Product Images from "Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6"

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M116.768689

GIMAP6 knockdown accelerates the PMA/ionomycin induction of T cell activation. A , Jurkat-derived cell lines ( Ctl , KD1 , and KD2 ) were treated with either PMA/ionomycin or an equal concentration of DMSO to determine the effect of GIMAP6 on T cell activation. To monitor the rate of cell apoptosis/death and T cell activation, PMA/ionomycin-treated cells were harvested at 3, 8, and 24 h and stained for the phosphatidylserine exposure marker Annexin V-FITC and for the T cell activation surface marker CD69-PE. The results were analyzed and quantified by FlowJo 7.6.1. B , the effect of GIMAP6 expression on PMA/ionomycin-induced cell death. All AnV+ cells were taken as apoptotic/dead cells, and the remaining AnV− cells were taken as live cells. C , the effect of GIMAP6 expression on PMA/ionomycin-induced T cell activation. To discriminate activated T cells from inactivated T cells across all live cells, the surface marker CD69 was used to calculate the percentage of CD69+/AnV−. D , the effect of GIMAP6 expression on PMA/ionomycin-induced IL-2 secretion. The data represent the average of at least two independent experiments with triplicate measurements (mean ± S.D.). P/I indicates that the cells were treated with both PMA (10 ng/ml) and ionomycin (1 μg/ml).
Figure Legend Snippet: GIMAP6 knockdown accelerates the PMA/ionomycin induction of T cell activation. A , Jurkat-derived cell lines ( Ctl , KD1 , and KD2 ) were treated with either PMA/ionomycin or an equal concentration of DMSO to determine the effect of GIMAP6 on T cell activation. To monitor the rate of cell apoptosis/death and T cell activation, PMA/ionomycin-treated cells were harvested at 3, 8, and 24 h and stained for the phosphatidylserine exposure marker Annexin V-FITC and for the T cell activation surface marker CD69-PE. The results were analyzed and quantified by FlowJo 7.6.1. B , the effect of GIMAP6 expression on PMA/ionomycin-induced cell death. All AnV+ cells were taken as apoptotic/dead cells, and the remaining AnV− cells were taken as live cells. C , the effect of GIMAP6 expression on PMA/ionomycin-induced T cell activation. To discriminate activated T cells from inactivated T cells across all live cells, the surface marker CD69 was used to calculate the percentage of CD69+/AnV−. D , the effect of GIMAP6 expression on PMA/ionomycin-induced IL-2 secretion. The data represent the average of at least two independent experiments with triplicate measurements (mean ± S.D.). P/I indicates that the cells were treated with both PMA (10 ng/ml) and ionomycin (1 μg/ml).

Techniques Used: Activation Assay, Derivative Assay, CTL Assay, Concentration Assay, Staining, Marker, Expressing

2) Product Images from "Ubiquitin-like (UBX)-domain-containing protein, UBXN2A, promotes cell death by interfering with the p53-Mortalin interactions in colon cancer cells"

Article Title: Ubiquitin-like (UBX)-domain-containing protein, UBXN2A, promotes cell death by interfering with the p53-Mortalin interactions in colon cancer cells

Journal: Cell Death & Disease

doi: 10.1038/cddis.2014.100

UBXN2A overexpression induces apoptosis in colon cancer cell lines but not in normal, non-transformed cells. CCD-18Co, HCT-116, and SW48 were transfected with GFP-empty or GFP-UBXN2A. 48 h after transfection, cells were stained with Annexin V and early apoptosis was determined using flow cytometry as described in Materials and Methods. ( a ) Representative flow-cytometry analysis data from an Annexin V assay. The histograms show a comparison of the distribution of Annexin V positive cells (M1) after transient transfection of cells with GFP-empty vector or GFP-UBXN2A. The data were gated on GFP-UBXN2A positive cells prior to Annexin V analysis. ( b ) UBXN2A expression for 48 h significantly increased the number of apoptotic cells in HCT-116 and SW48 cell lines, while CCD-18Co normal colon cells were unaffected. ( c ) HCT-116 were transfected with scrambled shRNA or shRNA against UBXN2A (clone I and II). 48 h after silencing, superconfluent cultures of HCT-116 were analyzed by Annexin V assay using flow cytometry. Expression of GFP containing shRNA against UBXN2A resulted in 50% less apoptosis in comparison with cells transfected with scrambled shRNA. Values are expressed as mean (±S.E.M.) from three independent experiments (** P
Figure Legend Snippet: UBXN2A overexpression induces apoptosis in colon cancer cell lines but not in normal, non-transformed cells. CCD-18Co, HCT-116, and SW48 were transfected with GFP-empty or GFP-UBXN2A. 48 h after transfection, cells were stained with Annexin V and early apoptosis was determined using flow cytometry as described in Materials and Methods. ( a ) Representative flow-cytometry analysis data from an Annexin V assay. The histograms show a comparison of the distribution of Annexin V positive cells (M1) after transient transfection of cells with GFP-empty vector or GFP-UBXN2A. The data were gated on GFP-UBXN2A positive cells prior to Annexin V analysis. ( b ) UBXN2A expression for 48 h significantly increased the number of apoptotic cells in HCT-116 and SW48 cell lines, while CCD-18Co normal colon cells were unaffected. ( c ) HCT-116 were transfected with scrambled shRNA or shRNA against UBXN2A (clone I and II). 48 h after silencing, superconfluent cultures of HCT-116 were analyzed by Annexin V assay using flow cytometry. Expression of GFP containing shRNA against UBXN2A resulted in 50% less apoptosis in comparison with cells transfected with scrambled shRNA. Values are expressed as mean (±S.E.M.) from three independent experiments (** P

Techniques Used: Over Expression, Transformation Assay, Transfection, Staining, Flow Cytometry, Cytometry, Annexin V Assay, Plasmid Preparation, Expressing, shRNA

Interaction of the SEP domain of UBXN2A with the p53-binding site (SBD domain) of mot-2 is sufficient to induce apoptosis. ( a , b ) Schematic diagram of UBXN2A and mot-2 protein domain structures. ( c – f ) Comprehensive mapping of protein–protein interaction sites by Y2H method using α-galactosidase activity and nutritional selection verified that (i) WT-UBXN2A interacts with WT-mot-2, (ii) the SEP domain of UBXN2A is sufficient to interact with WT-mot-2, and (iii) a partial section of p53 binding site on the SBD domain of mot-2 (aa:438-506) is sufficient for binding to WT-UBXN2A. ( g ) An apoptosis assay using Annexin V staining followed by flow cytometry analysis confirmed that only the SEP domain of UBXN2A is required to induce apoptosis in HCT-116 cells, similar to full WT-UBXN2A. No increase in apoptosis was seen with the GFP-UBX domain. ( h ) HCT-116 cells were transfected with GFP-UBXN2A (WT) or its truncated forms (GFP-SEP or GFP-UBX domains). After 48 h, cells were subjected to a crystal violet cell cytotoxicity assay. Counting the remaining colonies showed both WT-UBXN2A and GFP-SEP domains significantly induce cell cytotoxicity (* P
Figure Legend Snippet: Interaction of the SEP domain of UBXN2A with the p53-binding site (SBD domain) of mot-2 is sufficient to induce apoptosis. ( a , b ) Schematic diagram of UBXN2A and mot-2 protein domain structures. ( c – f ) Comprehensive mapping of protein–protein interaction sites by Y2H method using α-galactosidase activity and nutritional selection verified that (i) WT-UBXN2A interacts with WT-mot-2, (ii) the SEP domain of UBXN2A is sufficient to interact with WT-mot-2, and (iii) a partial section of p53 binding site on the SBD domain of mot-2 (aa:438-506) is sufficient for binding to WT-UBXN2A. ( g ) An apoptosis assay using Annexin V staining followed by flow cytometry analysis confirmed that only the SEP domain of UBXN2A is required to induce apoptosis in HCT-116 cells, similar to full WT-UBXN2A. No increase in apoptosis was seen with the GFP-UBX domain. ( h ) HCT-116 cells were transfected with GFP-UBXN2A (WT) or its truncated forms (GFP-SEP or GFP-UBX domains). After 48 h, cells were subjected to a crystal violet cell cytotoxicity assay. Counting the remaining colonies showed both WT-UBXN2A and GFP-SEP domains significantly induce cell cytotoxicity (* P

Techniques Used: Binding Assay, Activity Assay, Selection, Apoptosis Assay, Staining, Flow Cytometry, Cytometry, Transfection, Cytotoxicity Assay

Induction of apoptosis by UBXN2A is p53 dependent and caspase-mediated in colon cancer cell lines. ( a , b ) HCT-116 (p53+/+) or HCT-116 (p53−/−) cells were transiently transfected with GFP-empty or GFP-UBXN2A for 48 h. An Annexin V apoptosis assay ( a ) and Prestoblue cell viability ( b ) assay show that overexpression of UBXN2A leads to a significant increase in cell apoptosis ( c ) and reduction of cell viability ( d ) in HCT-116 with WT-p53 (p53+/+). There was not a significant change between GFP-empty and GFP-UBXN2A in p53-KO cells (* P
Figure Legend Snippet: Induction of apoptosis by UBXN2A is p53 dependent and caspase-mediated in colon cancer cell lines. ( a , b ) HCT-116 (p53+/+) or HCT-116 (p53−/−) cells were transiently transfected with GFP-empty or GFP-UBXN2A for 48 h. An Annexin V apoptosis assay ( a ) and Prestoblue cell viability ( b ) assay show that overexpression of UBXN2A leads to a significant increase in cell apoptosis ( c ) and reduction of cell viability ( d ) in HCT-116 with WT-p53 (p53+/+). There was not a significant change between GFP-empty and GFP-UBXN2A in p53-KO cells (* P

Techniques Used: Transfection, Apoptosis Assay, Over Expression

3) Product Images from "Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS"

Article Title: Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS

Journal: BMC Cancer

doi: 10.1186/1471-2407-14-139

Pharmacological inhibition of EZH2 restores myogenic differentiation of embryonal RMS cells in the presence of growth medium (GM). RD cells were analyzed for the induction of muscle-like differentiation after 6 days of 5 μM DZNep (a) and MC1945 (c) treatments. Representative immunofluorescence showing de novo expression of endogenous Myosin Heavy Chain (MHC, red) in multinucleated fibers of DZNep and MC1945 treated RD cells. Untreated (UN) and control cells treated with vehicle (i.e., water or DMSO) are shown. Representative immunofluorescence of three assays. mRNA levels (real time qRT-PCR) of Myogenin and MCK in RD treated for 72 h with 5 μM DZNep (b) and 5 μM MC1954 (d) were normalized to GAPDH levels and expressed as fold increase over Untreated condition (1 arbitrary unit, not reported). Columns, means; Bars, SD. Results from two independent experiments are shown. (e) RD cells Untreated or treated for 96 h with DZNep (left) or MC1945 (right) at the indicated concentrations were stained for Annexin V and 7-AAD, and the frequency of Annexin V and 7-AAD-positive labeling (% cell death) was recorded by flow cytometry. Representative cytofluorometric plots are shown. Annexin V+/7-AAD- events (lower right quadrants) represent early stages of apoptosis, whereas Annexin V+/7-AAD + events (upper right quadrants) stand for late apoptotic cells. Representative of three independent experiments run in duplicate.
Figure Legend Snippet: Pharmacological inhibition of EZH2 restores myogenic differentiation of embryonal RMS cells in the presence of growth medium (GM). RD cells were analyzed for the induction of muscle-like differentiation after 6 days of 5 μM DZNep (a) and MC1945 (c) treatments. Representative immunofluorescence showing de novo expression of endogenous Myosin Heavy Chain (MHC, red) in multinucleated fibers of DZNep and MC1945 treated RD cells. Untreated (UN) and control cells treated with vehicle (i.e., water or DMSO) are shown. Representative immunofluorescence of three assays. mRNA levels (real time qRT-PCR) of Myogenin and MCK in RD treated for 72 h with 5 μM DZNep (b) and 5 μM MC1954 (d) were normalized to GAPDH levels and expressed as fold increase over Untreated condition (1 arbitrary unit, not reported). Columns, means; Bars, SD. Results from two independent experiments are shown. (e) RD cells Untreated or treated for 96 h with DZNep (left) or MC1945 (right) at the indicated concentrations were stained for Annexin V and 7-AAD, and the frequency of Annexin V and 7-AAD-positive labeling (% cell death) was recorded by flow cytometry. Representative cytofluorometric plots are shown. Annexin V+/7-AAD- events (lower right quadrants) represent early stages of apoptosis, whereas Annexin V+/7-AAD + events (upper right quadrants) stand for late apoptotic cells. Representative of three independent experiments run in duplicate.

Techniques Used: Inhibition, Immunofluorescence, Expressing, Quantitative RT-PCR, Staining, Labeling, Flow Cytometry, Cytometry

4) Product Images from "Moscatilin Inhibits Lung Cancer Cell Motility and Invasion via Suppression of Endogenous Reactive Oxygen Species"

Article Title: Moscatilin Inhibits Lung Cancer Cell Motility and Invasion via Suppression of Endogenous Reactive Oxygen Species

Journal: BioMed Research International

doi: 10.1155/2013/765894

Cytotoxicity of moscatilin on human lung H23 cells. (a) Cells were treated with various concentrations of moscatilin (0–5 μ M) for 24 h. (b) Cells were treated with moscatilin (0-1 μ M) for various times (0–72 h). Cytotoxicity was determined by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. (c) After indicated treatment for 24 h, mode of cell death was examined by Hoechst33342/PI costaining assay and Annexin-V staining assay. (d) Cellular apoptosis was determined by DNA content analysis using flow cytometry. Data represent the means ± SD ( n = 4). * P
Figure Legend Snippet: Cytotoxicity of moscatilin on human lung H23 cells. (a) Cells were treated with various concentrations of moscatilin (0–5 μ M) for 24 h. (b) Cells were treated with moscatilin (0-1 μ M) for various times (0–72 h). Cytotoxicity was determined by 3-(4,5-dimethyl-thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. (c) After indicated treatment for 24 h, mode of cell death was examined by Hoechst33342/PI costaining assay and Annexin-V staining assay. (d) Cellular apoptosis was determined by DNA content analysis using flow cytometry. Data represent the means ± SD ( n = 4). * P

Techniques Used: MTT Assay, Annexin V Staining Assay, Flow Cytometry, Cytometry

5) Product Images from "Downregulation of A20 increases the cytotoxicity of IFN-γ in hepatocellular carcinoma cells"

Article Title: Downregulation of A20 increases the cytotoxicity of IFN-γ in hepatocellular carcinoma cells

Journal: Drug Design, Development and Therapy

doi: 10.2147/DDDT.S135993

A20 knockdown increases the apoptosis induced by IFN-γ in HepG2 cells. Notes: ( A ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were then subjected to flow cytometry analysis using Annexin-V/propidium iodide staining. Quantification of apoptosis was derived from three independent experiments (right). ( B ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cellular lysates were subjected to Western blot analysis with the indicated antibodies. Graphs represent quantification of target protein bands relative to GAPDH. ( C ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were lysed and assayed for caspase-3, -8 and -9 activity. Graphs represent the mean ± SD of three independent experiments. ( D ) sh-A20 cells were incubated with 100 ng/mL of IFN-γ for 24 h after 1 h pretreatment with z-DEVD-fmk, z-LEHD-fmk and z-IETD-fmk, respectively. Cell viability (left) was determined by MTT assay and apoptosis (right) was determined by flow cytometry. The data are represented as the mean ± SD of three independent experiments. The significance was determined by the Student’s t -test (** P
Figure Legend Snippet: A20 knockdown increases the apoptosis induced by IFN-γ in HepG2 cells. Notes: ( A ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were then subjected to flow cytometry analysis using Annexin-V/propidium iodide staining. Quantification of apoptosis was derived from three independent experiments (right). ( B ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cellular lysates were subjected to Western blot analysis with the indicated antibodies. Graphs represent quantification of target protein bands relative to GAPDH. ( C ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were lysed and assayed for caspase-3, -8 and -9 activity. Graphs represent the mean ± SD of three independent experiments. ( D ) sh-A20 cells were incubated with 100 ng/mL of IFN-γ for 24 h after 1 h pretreatment with z-DEVD-fmk, z-LEHD-fmk and z-IETD-fmk, respectively. Cell viability (left) was determined by MTT assay and apoptosis (right) was determined by flow cytometry. The data are represented as the mean ± SD of three independent experiments. The significance was determined by the Student’s t -test (** P

Techniques Used: Flow Cytometry, Cytometry, Staining, Derivative Assay, Western Blot, Activity Assay, Incubation, MTT Assay

6) Product Images from "The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia"

Article Title: The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia

Journal: Blood Advances

doi: 10.1182/bloodadvances.2019032029

HOXA10-AS is required for the maintenance of KMT2A -rearranged AML cells. (A) Percentage of cells transduced with shRNAs against HOXA10-AS after 21 days in culture compared with day 5 and a nontargeting shRNA (ctrl; n = 3; unpaired Student t test; presented as mean ± SEM). (B) Cell counts on day 21 following treatment with LNA-GapmeRs against HOXA10-AS compared with day 2 and the nontargeting control (ctrl; n = 2; unpaired Student t test; presented as mean ± SEM). (A-B) EOL-1, MOLM-13, and MV4-11 were used as KMT2A-r cell lines, while NB-4 was used as the non- KMT2A-r cell line. (C-D) 5-Bromo-2′-deoxyuridine cell cycle (C) and Annexin V apoptosis (D) assays in EOL-1 and NB-4 cells after shRNA-mediated HOXA10-AS knockdown using 3 different shRNAs. The percentage of cells is shown as mean ± SD (n = 2; * P
Figure Legend Snippet: HOXA10-AS is required for the maintenance of KMT2A -rearranged AML cells. (A) Percentage of cells transduced with shRNAs against HOXA10-AS after 21 days in culture compared with day 5 and a nontargeting shRNA (ctrl; n = 3; unpaired Student t test; presented as mean ± SEM). (B) Cell counts on day 21 following treatment with LNA-GapmeRs against HOXA10-AS compared with day 2 and the nontargeting control (ctrl; n = 2; unpaired Student t test; presented as mean ± SEM). (A-B) EOL-1, MOLM-13, and MV4-11 were used as KMT2A-r cell lines, while NB-4 was used as the non- KMT2A-r cell line. (C-D) 5-Bromo-2′-deoxyuridine cell cycle (C) and Annexin V apoptosis (D) assays in EOL-1 and NB-4 cells after shRNA-mediated HOXA10-AS knockdown using 3 different shRNAs. The percentage of cells is shown as mean ± SD (n = 2; * P

Techniques Used: Transduction, shRNA

7) Product Images from "Small interfering RNA targeting S100A4 sensitizes non-small-cell lung cancer cells (A549) to radiation treatment"

Article Title: Small interfering RNA targeting S100A4 sensitizes non-small-cell lung cancer cells (A549) to radiation treatment

Journal: OncoTargets and therapy

doi: 10.2147/OTT.S106557

Flow cytometry analysis for apoptotic cells with annexin V/PI. Notes: ( A ) Representative flow cytometry results of cell apoptosis. B1, dead cells; B2, late apoptosis; B3, viable cells; B4, early apoptotic cells. ( B ) The columns illustrate the flow cytometric results of the apoptosis rate. Data are presented as the mean ± SD of three independent experiments. * P
Figure Legend Snippet: Flow cytometry analysis for apoptotic cells with annexin V/PI. Notes: ( A ) Representative flow cytometry results of cell apoptosis. B1, dead cells; B2, late apoptosis; B3, viable cells; B4, early apoptotic cells. ( B ) The columns illustrate the flow cytometric results of the apoptosis rate. Data are presented as the mean ± SD of three independent experiments. * P

Techniques Used: Flow Cytometry, Cytometry

8) Product Images from "Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma"

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma

Journal: Oncotarget

doi: 10.18632/oncotarget.6831

SEMA3A over-expression induces apoptosis in HNSCC cells in a caspase-dependent manner ( A ) A gain of apoptosis-like phenotype of SEMA3A-transfected cells on 48 and 96 hours after transfection. Red arrowheads indicated apoptotic cells. Scale bar: 100 μm. ( B ) Flow cytometric analysis of apoptosis in Ad-SEMA3A/Con-CAL27/HN6. Cells were stained with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD), followed by FACS (fluorescence-activated cell sorting) analysis (up). Apoptosis was determined by FACS analysis (early apoptotic death cells in lower right plot quadrants and late apoptotic death cells in upper right plot quadrants) and plotted (down). As showed, apoptosis rate in Ad-SEMA3A cells was significantly higher than controls (** P
Figure Legend Snippet: SEMA3A over-expression induces apoptosis in HNSCC cells in a caspase-dependent manner ( A ) A gain of apoptosis-like phenotype of SEMA3A-transfected cells on 48 and 96 hours after transfection. Red arrowheads indicated apoptotic cells. Scale bar: 100 μm. ( B ) Flow cytometric analysis of apoptosis in Ad-SEMA3A/Con-CAL27/HN6. Cells were stained with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD), followed by FACS (fluorescence-activated cell sorting) analysis (up). Apoptosis was determined by FACS analysis (early apoptotic death cells in lower right plot quadrants and late apoptotic death cells in upper right plot quadrants) and plotted (down). As showed, apoptosis rate in Ad-SEMA3A cells was significantly higher than controls (** P

Techniques Used: Over Expression, Transfection, Flow Cytometry, Staining, FACS, Fluorescence

9) Product Images from "Molecular Mechanisms of Action of Herbal Antifungal Alkaloid Berberine, in Candida albicans"

Article Title: Molecular Mechanisms of Action of Herbal Antifungal Alkaloid Berberine, in Candida albicans

Journal: PLoS ONE

doi: 10.1371/journal.pone.0104554

Determination of endogenous ROS generation by BER and induction of apoptosis (a) (upper panel) bar graph representing relative fluorescent units when cells were treated with DCFDA in presence and absence of BER, AA is added to revert the ROS production, (lower panel) fluorescent microscopy images of WT C. albicans cells labeled with DCFDA, (b) Cytometric determination FITC Annexin V labeling in WT cells treated with BER.
Figure Legend Snippet: Determination of endogenous ROS generation by BER and induction of apoptosis (a) (upper panel) bar graph representing relative fluorescent units when cells were treated with DCFDA in presence and absence of BER, AA is added to revert the ROS production, (lower panel) fluorescent microscopy images of WT C. albicans cells labeled with DCFDA, (b) Cytometric determination FITC Annexin V labeling in WT cells treated with BER.

Techniques Used: Microscopy, Labeling

10) Product Images from "Plk2 Regulated by miR-128 Induces Ischemia-Reperfusion Injury in Cardiac Cells"

Article Title: Plk2 Regulated by miR-128 Induces Ischemia-Reperfusion Injury in Cardiac Cells

Journal: Molecular Therapy. Nucleic Acids

doi: 10.1016/j.omtn.2019.11.029

In Vitro I/R Stimulated with AA Induced Cardiac Cell Apoptosis/Death, Increased Plk2 Expression, and Reduced miR-128 (A) Cell apoptosis. H9c2 cells were cultured and treated with 20 μM AA or PBS (as a control) for 3 h, followed by 3-h reperfusion with complete medium. Cells were double stained with FITC-Annexin V and PI, followed by flow cytometry analysis. Left panel: representative images from flow cytometry result; right panel: summarized results of flow cytometry. n = 5; ***p
Figure Legend Snippet: In Vitro I/R Stimulated with AA Induced Cardiac Cell Apoptosis/Death, Increased Plk2 Expression, and Reduced miR-128 (A) Cell apoptosis. H9c2 cells were cultured and treated with 20 μM AA or PBS (as a control) for 3 h, followed by 3-h reperfusion with complete medium. Cells were double stained with FITC-Annexin V and PI, followed by flow cytometry analysis. Left panel: representative images from flow cytometry result; right panel: summarized results of flow cytometry. n = 5; ***p

Techniques Used: In Vitro, Expressing, Cell Culture, Staining, Flow Cytometry, Cytometry

11) Product Images from "LINC00365-SCGB2A1 axis inhibits the viability of breast cancer through targeting NF-κB signaling"

Article Title: LINC00365-SCGB2A1 axis inhibits the viability of breast cancer through targeting NF-κB signaling

Journal: Oncology Letters

doi: 10.3892/ol.2019.11166

Overexpression of LINC00365 and SCGB2A1 induces apoptosis in breast cancer cells. (A) Apoptotic rate was determined by Annexin V/propidium iodide assay following transfection of MCF-7 and T47D cells with LINC00365- and SCGB2A1-expressing vectors. (B) Quantitative analysis of the apoptosis results. (C) Western blotting analysis of cleaved-caspase-3, Bcl-xl, Bcl-2 and Bax in MCF-7 and T47D cells following transfection with LINC00365- and SCGB2A1-expressing vectors. β-actin was used as an internal control. (D-F) The ratio of protein/β-actin in T47D and MCF-7 cells. *P
Figure Legend Snippet: Overexpression of LINC00365 and SCGB2A1 induces apoptosis in breast cancer cells. (A) Apoptotic rate was determined by Annexin V/propidium iodide assay following transfection of MCF-7 and T47D cells with LINC00365- and SCGB2A1-expressing vectors. (B) Quantitative analysis of the apoptosis results. (C) Western blotting analysis of cleaved-caspase-3, Bcl-xl, Bcl-2 and Bax in MCF-7 and T47D cells following transfection with LINC00365- and SCGB2A1-expressing vectors. β-actin was used as an internal control. (D-F) The ratio of protein/β-actin in T47D and MCF-7 cells. *P

Techniques Used: Over Expression, Transfection, Expressing, Western Blot

12) Product Images from "MicroRNA-125b promotes tumor growth and suppresses apoptosis by targeting DRAM2 in retinoblastoma"

Article Title: MicroRNA-125b promotes tumor growth and suppresses apoptosis by targeting DRAM2 in retinoblastoma

Journal: Eye

doi: 10.1038/eye.2016.189

MiR-125b affects retinoblastoma cell migration, invasion, and apoptosis in vitro . (a) The effect of miR-125b apoptosis was examined by flow cytometric analysis. Cells were analyzed for apoptotic rate after staining with Annexin V-FITC and PI staining, as described in the Materials and Methods section. (b) Quantification of relative apoptotic in HXO-Rb44 cells transfected with miR-125b mimic or miR-125b inhibitor for 48 h. Data represent mean±SD from three independent experiments. (c) Wound-healing assays were used to analyze the effect of miR-125b on cell migration of HXO-Rb44 cells. (d) Transwell assays were used to analyze the effect of miR-125b on cell invasion of HXO-Rb44 cells. The symbols ** and * indicate differences from their respective controls at P
Figure Legend Snippet: MiR-125b affects retinoblastoma cell migration, invasion, and apoptosis in vitro . (a) The effect of miR-125b apoptosis was examined by flow cytometric analysis. Cells were analyzed for apoptotic rate after staining with Annexin V-FITC and PI staining, as described in the Materials and Methods section. (b) Quantification of relative apoptotic in HXO-Rb44 cells transfected with miR-125b mimic or miR-125b inhibitor for 48 h. Data represent mean±SD from three independent experiments. (c) Wound-healing assays were used to analyze the effect of miR-125b on cell migration of HXO-Rb44 cells. (d) Transwell assays were used to analyze the effect of miR-125b on cell invasion of HXO-Rb44 cells. The symbols ** and * indicate differences from their respective controls at P

Techniques Used: Migration, In Vitro, Flow Cytometry, Staining, Transfection

13) Product Images from "Immune-mediated bone marrow failure in C57BL/6 mice"

Article Title: Immune-mediated bone marrow failure in C57BL/6 mice

Journal: Experimental hematology

doi: 10.1016/j.exphem.2014.12.006

Elevation in hematopoietic cell apoptosis and BM destruction. BM cells from TBI + FVB LN treated mice (N=5) had significantly higher proportions of Annexin V + (including both 7AAD high and 7AAD low ) apoptotic cells in KSL (P
Figure Legend Snippet: Elevation in hematopoietic cell apoptosis and BM destruction. BM cells from TBI + FVB LN treated mice (N=5) had significantly higher proportions of Annexin V + (including both 7AAD high and 7AAD low ) apoptotic cells in KSL (P

Techniques Used: Mouse Assay

14) Product Images from "Five-aza-2′-deoxycytidine-induced hypomethylation of cholesterol 25-hydroxylase gene is responsible for cell death of myelodysplasia/leukemia cells"

Article Title: Five-aza-2′-deoxycytidine-induced hypomethylation of cholesterol 25-hydroxylase gene is responsible for cell death of myelodysplasia/leukemia cells

Journal: Scientific Reports

doi: 10.1038/srep16709

The role of CH25H in DAC-induced cell death. To investigate the role of CH25H in detail, we established CH25H -knockdown HL-60 cell clones (HL-60- CH25H shRNA) and MDS-L cell clones (MDS-L- CH25H shRNA) by transfecting shRNA- CH25H into each cell line using a lentivirus vector. ( a ) Displayed are histograms of CH25H expression level in HL-60-CV (CV) and HL60- CH25H shRNA (shRNA) treated with or without DAC for 4 days, respectively. ( b ) Shown is the survival (%) from DAC-induced HL-60 cell death by CH25H -knockdown. HL-60-CV cells (CV) and HL-60- CH25H shRNA cells (shRNA) were treated daily with DAC (0–500 nM) for 3 to 4 days, and the change in the cell number was evaluated by MTT assay. ( c ) HL-60-CV cells (CV) and HL-60- CH25H shRNA cells (shRNA) were treated with different concentrations (0 and 500 nM) of DAC for 96 h, and apoptosis was assessed by flow cytometry using annexin V and propidium iodide (PI) staining. Positive staining for annexin V or for annexin V and PI show early or late apoptosis, respectively. The value of right area indicates the percentage of the cells in early and late apoptosis. ( d ) Displayed are the histograms of CH25H expression level in MDS-L-CV (CV) and MDS-L- CH25H shRNA (shRNA) treated with or without DAC for 4 days, respectively. ( e ) Shown is the survival (%) from DAC-induced MDS-L cell death by CH25H -knockdown. MDS-L-CV cells (CV) and MDS-L- CH25H shRNA cells (shRNA) were treated daily with DAC (0–100 nM) for 4 days, and the change in the cell number was evaluated by MTT assay. The data shown are the average ± SD of three ( a,d ) or five ( b,e ) independent experiments. *p
Figure Legend Snippet: The role of CH25H in DAC-induced cell death. To investigate the role of CH25H in detail, we established CH25H -knockdown HL-60 cell clones (HL-60- CH25H shRNA) and MDS-L cell clones (MDS-L- CH25H shRNA) by transfecting shRNA- CH25H into each cell line using a lentivirus vector. ( a ) Displayed are histograms of CH25H expression level in HL-60-CV (CV) and HL60- CH25H shRNA (shRNA) treated with or without DAC for 4 days, respectively. ( b ) Shown is the survival (%) from DAC-induced HL-60 cell death by CH25H -knockdown. HL-60-CV cells (CV) and HL-60- CH25H shRNA cells (shRNA) were treated daily with DAC (0–500 nM) for 3 to 4 days, and the change in the cell number was evaluated by MTT assay. ( c ) HL-60-CV cells (CV) and HL-60- CH25H shRNA cells (shRNA) were treated with different concentrations (0 and 500 nM) of DAC for 96 h, and apoptosis was assessed by flow cytometry using annexin V and propidium iodide (PI) staining. Positive staining for annexin V or for annexin V and PI show early or late apoptosis, respectively. The value of right area indicates the percentage of the cells in early and late apoptosis. ( d ) Displayed are the histograms of CH25H expression level in MDS-L-CV (CV) and MDS-L- CH25H shRNA (shRNA) treated with or without DAC for 4 days, respectively. ( e ) Shown is the survival (%) from DAC-induced MDS-L cell death by CH25H -knockdown. MDS-L-CV cells (CV) and MDS-L- CH25H shRNA cells (shRNA) were treated daily with DAC (0–100 nM) for 4 days, and the change in the cell number was evaluated by MTT assay. The data shown are the average ± SD of three ( a,d ) or five ( b,e ) independent experiments. *p

Techniques Used: Clone Assay, shRNA, Plasmid Preparation, Expressing, MTT Assay, Flow Cytometry, Cytometry, Staining

Growth suppressive effect of 25-hydroxycholesterol (25-OHC) on MDS/leukemia cell lines. ( a ) The indicated cell lines were cultured for 3 days in the presence of various concentrations of 25-OHC or 27-OHC, and growth suppression was evaluated by MTT assay. ( b ) MDS-L cells were treated with 0, 1 or 5 μM of 25-OHC for 72 h, and apoptosis was assessed by flow cytometry using annexin V and PI staining. Positive staining for annexin V or for annexin V and PI show early or late apoptosis, respectively. The value of lower right area and upper right area indicate the percentage of cells in early apoptosis and late apoptosis, respectively. ( c ) MDS-L cells were treated with 5 nM DAC in the presence or absence of 10 μM desmosterol for 4 days. Shown is the survival (%) from DAC-induced MDS-L cell death by desmosterol. The change in the cell number was evaluated by MTT assay. The data shown are the average ± SD of five independent experiments and there was significant difference between DAC-treated and DAC plus desmosterol-treated cells. *p
Figure Legend Snippet: Growth suppressive effect of 25-hydroxycholesterol (25-OHC) on MDS/leukemia cell lines. ( a ) The indicated cell lines were cultured for 3 days in the presence of various concentrations of 25-OHC or 27-OHC, and growth suppression was evaluated by MTT assay. ( b ) MDS-L cells were treated with 0, 1 or 5 μM of 25-OHC for 72 h, and apoptosis was assessed by flow cytometry using annexin V and PI staining. Positive staining for annexin V or for annexin V and PI show early or late apoptosis, respectively. The value of lower right area and upper right area indicate the percentage of cells in early apoptosis and late apoptosis, respectively. ( c ) MDS-L cells were treated with 5 nM DAC in the presence or absence of 10 μM desmosterol for 4 days. Shown is the survival (%) from DAC-induced MDS-L cell death by desmosterol. The change in the cell number was evaluated by MTT assay. The data shown are the average ± SD of five independent experiments and there was significant difference between DAC-treated and DAC plus desmosterol-treated cells. *p

Techniques Used: Cell Culture, MTT Assay, Flow Cytometry, Cytometry, Staining

15) Product Images from "Dual Specificity Phosphatase 5 Is Essential for T Cell Survival"

Article Title: Dual Specificity Phosphatase 5 Is Essential for T Cell Survival

Journal: PLoS ONE

doi: 10.1371/journal.pone.0167246

T cell survival is regulated by DUSP5. A: Legend for Annexin V/ Propidium Iodide flow plots (left) and representative flow plots of WT and Dusp5 -/- samples (right). B: Overall comparison of apoptotic populations in T cells after 6 days culture in vitro . C: Individual comparison of live cell, early apoptosis, and necrotic cell populations. Dusp5 -/- CD8 + T cells show significant reductions in live cells in both SLEC and MPEC conditions and significant increases in necrotic cells in MPEC culture conditions. D: Overall comparison of cell cycle populations in T cells after 4 days culture in vitro and specific comparison of G1, S, and G2 populations. Dusp5 -/- T cells show increases in proportion of cells in S phase and decreases in proportions in G1 and G2 phases in both SLEC and MPEC cultured T cells. E: Dusp5 -/- T cells have increased spare respiratory capacity (SRC) compared to WT T cells. SRC is a key survival mechanism in memory T cells. Together, these data show that DUSP5 plays an important role in T cell survival. Plots are means with error bars ±SEM. Comparisons were made using a two-tailed t-test, *: p
Figure Legend Snippet: T cell survival is regulated by DUSP5. A: Legend for Annexin V/ Propidium Iodide flow plots (left) and representative flow plots of WT and Dusp5 -/- samples (right). B: Overall comparison of apoptotic populations in T cells after 6 days culture in vitro . C: Individual comparison of live cell, early apoptosis, and necrotic cell populations. Dusp5 -/- CD8 + T cells show significant reductions in live cells in both SLEC and MPEC conditions and significant increases in necrotic cells in MPEC culture conditions. D: Overall comparison of cell cycle populations in T cells after 4 days culture in vitro and specific comparison of G1, S, and G2 populations. Dusp5 -/- T cells show increases in proportion of cells in S phase and decreases in proportions in G1 and G2 phases in both SLEC and MPEC cultured T cells. E: Dusp5 -/- T cells have increased spare respiratory capacity (SRC) compared to WT T cells. SRC is a key survival mechanism in memory T cells. Together, these data show that DUSP5 plays an important role in T cell survival. Plots are means with error bars ±SEM. Comparisons were made using a two-tailed t-test, *: p

Techniques Used: Flow Cytometry, In Vitro, Cell Culture, Two Tailed Test

16) Product Images from "Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells"

Article Title: Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells

Journal: Journal of Translational Medicine

doi: 10.1186/s12967-016-1096-1

Functional change in iNOS+ MDSCs. a Total iNOS + cells were harvested from the spleen of mice treated by VACV (10 d.p.i) ( filled square ) or untreated mice ( open square ). Isolated cells were mixed at the indicated ratios with HCT-116 target cells in a transwell system for 12 h. In addition, where indicated, 0.5 mM L-NIL was added to inhibit NO production. Cytotoxicity was measured by Alamar blue CTL assay and tumor killing was shown as % lysis. Experiments were performed in triplicate, n = 2. Error bars represent mean and SD. b In a separate experiment, HCT-116 cells isolated from the lower transwell chamber were analyzed by flow cytometry for apoptosis at the end of 12 h of culture period. The data is presented as % Annexin V + cells. This is representative data out of two distinct experiments with similar results. c NO level was determined by nitrite measurement in the media. The culture supernatant obtained at 12 h of incubation period was used for the determination of nitrite level
Figure Legend Snippet: Functional change in iNOS+ MDSCs. a Total iNOS + cells were harvested from the spleen of mice treated by VACV (10 d.p.i) ( filled square ) or untreated mice ( open square ). Isolated cells were mixed at the indicated ratios with HCT-116 target cells in a transwell system for 12 h. In addition, where indicated, 0.5 mM L-NIL was added to inhibit NO production. Cytotoxicity was measured by Alamar blue CTL assay and tumor killing was shown as % lysis. Experiments were performed in triplicate, n = 2. Error bars represent mean and SD. b In a separate experiment, HCT-116 cells isolated from the lower transwell chamber were analyzed by flow cytometry for apoptosis at the end of 12 h of culture period. The data is presented as % Annexin V + cells. This is representative data out of two distinct experiments with similar results. c NO level was determined by nitrite measurement in the media. The culture supernatant obtained at 12 h of incubation period was used for the determination of nitrite level

Techniques Used: Functional Assay, Mouse Assay, Isolation, CTL Assay, Lysis, Flow Cytometry, Cytometry, Incubation

17) Product Images from "Expression and Regulation of Prostate Apoptosis Response-4 (Par-4) in Human Glioma Stem Cells in Drug-Induced Apoptosis"

Article Title: Expression and Regulation of Prostate Apoptosis Response-4 (Par-4) in Human Glioma Stem Cells in Drug-Induced Apoptosis

Journal: PLoS ONE

doi: 10.1371/journal.pone.0088505

Tamoxifen induces apoptosis and Par-4 expression in Primary GBM cells. (A) G1 cells were treated with TAM for 12 h and analyzed for apoptosis by staining cells with Annexin V and PI followed by flow cytometry analysis. The bars in the histogram represent mean ±SE of three independent experiments. * p
Figure Legend Snippet: Tamoxifen induces apoptosis and Par-4 expression in Primary GBM cells. (A) G1 cells were treated with TAM for 12 h and analyzed for apoptosis by staining cells with Annexin V and PI followed by flow cytometry analysis. The bars in the histogram represent mean ±SE of three independent experiments. * p

Techniques Used: Expressing, Staining, Flow Cytometry, Cytometry

Tamoxifen induces cell death by apoptosis. (A) HNGC-2 cells were exposed to lomustine (Lom), carmustine (Carm), UCN-01, oxaliplatin (Ox), tamoxifen (TAM) and temozolomide (TMZ) for 12 h and stained by Annexin V (FL-1) and Propidium Iodide (FL-2) to perform flowcytometry analysis. The graph represents percent apoptotic population (Annexin V and PI positive). The bars represent mean± SE of three independent experiments. *p
Figure Legend Snippet: Tamoxifen induces cell death by apoptosis. (A) HNGC-2 cells were exposed to lomustine (Lom), carmustine (Carm), UCN-01, oxaliplatin (Ox), tamoxifen (TAM) and temozolomide (TMZ) for 12 h and stained by Annexin V (FL-1) and Propidium Iodide (FL-2) to perform flowcytometry analysis. The graph represents percent apoptotic population (Annexin V and PI positive). The bars represent mean± SE of three independent experiments. *p

Techniques Used: Staining

18) Product Images from "LPS promotes resistance to TRAIL-induced apoptosis in pancreatic cancer"

Article Title: LPS promotes resistance to TRAIL-induced apoptosis in pancreatic cancer

Journal: Infectious Agents and Cancer

doi: 10.1186/s13027-017-0139-4

LPS-stimulation promoted resistance to TRAIL-induced apoptosis. a Cells cultures of COLO357, BxPC-3 and PANC-1 were stimulated with TRAIL for 24 h and cell viability was determined using a Cell titer blue assay. Mean fluorescence intensities following TRAIL-stimulation are shown. TRAIL – stimulation decreased viability of COLO 357 and BxPC-3 whereas viability of PANC-1 cells remained almost unchanged. N = 5/group. b Cell cultures of COLO357, BxPC-3 and PANC-1 were TRAIL-stimulated for 24 h and the fraction of apoptotic cells was determined via FACS analyses employing a Annexin V assay. TRAIL-stimulation induced apoptosis in COLO 357 and, to a lesser degree, in BxPC-3 cells, whereas PANC-1 cells were TRAIL-resistant. N = 5/group. c Cell cultures of COLO357, BxPC-3 and PANC-1 were stimulated with 300 ng/ml TRAIL, 1 μg/ml LPS and 300 ng/ml TRAIL + 1 μg/ml LPS for 24 h. Non-stimulated cell cultures served for controls. Thereafter, fractions of apoptotic cells were determined. Co-stimulation with TRAIL and LPS significantly decreased the number of TRAIL-induced apoptotic cells in COLO357 and BxPC-3. N = 5/group. d Representative histograms of FACS analyses employing the Annexin V assay of COLO357 and BxPC-3 are shown. Means and standard errors of the mean are shown. *) p
Figure Legend Snippet: LPS-stimulation promoted resistance to TRAIL-induced apoptosis. a Cells cultures of COLO357, BxPC-3 and PANC-1 were stimulated with TRAIL for 24 h and cell viability was determined using a Cell titer blue assay. Mean fluorescence intensities following TRAIL-stimulation are shown. TRAIL – stimulation decreased viability of COLO 357 and BxPC-3 whereas viability of PANC-1 cells remained almost unchanged. N = 5/group. b Cell cultures of COLO357, BxPC-3 and PANC-1 were TRAIL-stimulated for 24 h and the fraction of apoptotic cells was determined via FACS analyses employing a Annexin V assay. TRAIL-stimulation induced apoptosis in COLO 357 and, to a lesser degree, in BxPC-3 cells, whereas PANC-1 cells were TRAIL-resistant. N = 5/group. c Cell cultures of COLO357, BxPC-3 and PANC-1 were stimulated with 300 ng/ml TRAIL, 1 μg/ml LPS and 300 ng/ml TRAIL + 1 μg/ml LPS for 24 h. Non-stimulated cell cultures served for controls. Thereafter, fractions of apoptotic cells were determined. Co-stimulation with TRAIL and LPS significantly decreased the number of TRAIL-induced apoptotic cells in COLO357 and BxPC-3. N = 5/group. d Representative histograms of FACS analyses employing the Annexin V assay of COLO357 and BxPC-3 are shown. Means and standard errors of the mean are shown. *) p

Techniques Used: Fluorescence, FACS, Annexin V Assay

19) Product Images from "Inhibition of mitochondrial matrix chaperones and anti-apoptotic Bcl-2 family proteins empower antitumor therapeutic responses"

Article Title: Inhibition of mitochondrial matrix chaperones and anti-apoptotic Bcl-2 family proteins empower antitumor therapeutic responses

Journal: Cancer research

doi: 10.1158/0008-5472.CAN-16-3424

Combined treatment with G-TPP and broad or selective BH3-mimetics yields enhanced induction of apoptosis. A, Representative histograms of U87MG and T98G glioblastoma cells treated with solvent, ABT263, G-TPP or the combination as indicated for 48h prior to staining for Annexin V and propidium iodide and flowcytometric analysis. B, Representative histograms of U87MG and T98G glioblastoma cells that were treated for 24 h with G-TPP, ABT263, both or solvent prior to staining with JC-1 and flow cytometric analysis. C, T98G and LN229 glioblastoma cells were treated for 7 h with G-TPP, ABT263, both agents and solvent under serum starvation. Whole-cell extracts were examined by Western blot analysis for caspase 9 (CP9) and cleaved caspase 3 (cCP3). Actin Western blot analysis was performed to confirm equal protein loading. D, LN229 cells were treated with the combination of ABT263 and G-TPP as indicated in the presence or absence of the pan-caspase inhibitor zVAD.fmk (20µM). Staining for propidium iodide and flowcytometric analysis was performed to determine the fraction of subG1 cells. E, LN229 cells were treated with the combination of ABT263 and G-TPP as indicated in the presence or absence of necrostatin (20µM). Staining for propidium iodide and flowcytometric analysis was performed to determine the fraction of subG1 cells. F–G, U251 glioblastoma (F) or MeWo melanoma (G) cells were treated with selective BH-3 mimetics, G-TPP or the combination of both for 24 h (U251) or 48 h (MeWo). Thereafter, cells were stained with annexin V and propidium iodide and analyzed by flow cytometry. Shown are representative flow plots.
Figure Legend Snippet: Combined treatment with G-TPP and broad or selective BH3-mimetics yields enhanced induction of apoptosis. A, Representative histograms of U87MG and T98G glioblastoma cells treated with solvent, ABT263, G-TPP or the combination as indicated for 48h prior to staining for Annexin V and propidium iodide and flowcytometric analysis. B, Representative histograms of U87MG and T98G glioblastoma cells that were treated for 24 h with G-TPP, ABT263, both or solvent prior to staining with JC-1 and flow cytometric analysis. C, T98G and LN229 glioblastoma cells were treated for 7 h with G-TPP, ABT263, both agents and solvent under serum starvation. Whole-cell extracts were examined by Western blot analysis for caspase 9 (CP9) and cleaved caspase 3 (cCP3). Actin Western blot analysis was performed to confirm equal protein loading. D, LN229 cells were treated with the combination of ABT263 and G-TPP as indicated in the presence or absence of the pan-caspase inhibitor zVAD.fmk (20µM). Staining for propidium iodide and flowcytometric analysis was performed to determine the fraction of subG1 cells. E, LN229 cells were treated with the combination of ABT263 and G-TPP as indicated in the presence or absence of necrostatin (20µM). Staining for propidium iodide and flowcytometric analysis was performed to determine the fraction of subG1 cells. F–G, U251 glioblastoma (F) or MeWo melanoma (G) cells were treated with selective BH-3 mimetics, G-TPP or the combination of both for 24 h (U251) or 48 h (MeWo). Thereafter, cells were stained with annexin V and propidium iodide and analyzed by flow cytometry. Shown are representative flow plots.

Techniques Used: Staining, Flow Cytometry, Western Blot, Cytometry

20) Product Images from "Honokiol Eliminates Human Oral Cancer Stem-Like Cells Accompanied with Suppression of Wnt/β-Catenin Signaling and Apoptosis Induction"

Article Title: Honokiol Eliminates Human Oral Cancer Stem-Like Cells Accompanied with Suppression of Wnt/β-Catenin Signaling and Apoptosis Induction

Journal: Evidence-based Complementary and Alternative Medicine : eCAM

doi: 10.1155/2013/146136

Honokiol-induced apoptosis of SAS SP cells. (a) Annexin V-PI staining of SP cells after treatment with honokiol for 48 h. Q1: quadrant (annexin V−, PI+) represents dead cells; Q2: quadrant (annexin V+, PI+) represents late apoptotic cells; Q3: quadrant (annexin V+, PI–) represents early apoptotic cells; Q4: quadrant (annexin V−, PI−) represents live cells. The percentage of cell population in each quadrant is calculated and shown in histograms and a dose-dependent increase of honokiol-induced apoptosis was displayed. (b) Decrease of Survivin and Bcl-2 protein levels and increase of caspase-3 cleavage in honokiol-treated SP cells. Cells were treated with honokiol for 48 h and the lysates were analyzed by Western blot using β -actin as loading control.
Figure Legend Snippet: Honokiol-induced apoptosis of SAS SP cells. (a) Annexin V-PI staining of SP cells after treatment with honokiol for 48 h. Q1: quadrant (annexin V−, PI+) represents dead cells; Q2: quadrant (annexin V+, PI+) represents late apoptotic cells; Q3: quadrant (annexin V+, PI–) represents early apoptotic cells; Q4: quadrant (annexin V−, PI−) represents live cells. The percentage of cell population in each quadrant is calculated and shown in histograms and a dose-dependent increase of honokiol-induced apoptosis was displayed. (b) Decrease of Survivin and Bcl-2 protein levels and increase of caspase-3 cleavage in honokiol-treated SP cells. Cells were treated with honokiol for 48 h and the lysates were analyzed by Western blot using β -actin as loading control.

Techniques Used: Staining, Western Blot

21) Product Images from "cAMP induces cell apoptosis in multiple myeloma and overcomes bortezomib resistance"

Article Title: cAMP induces cell apoptosis in multiple myeloma and overcomes bortezomib resistance

Journal: American Journal of Cancer Research

doi:

The combination of 8-CPT-cAMP and bortezomib synergistically induced apoptosis in bortezomib-sensitive and -resistant multiple myeloma cells. A. Giemsa staining of U266, H929, U266-R, and H929-R cells following the treatment with 8-CPT-cAMP and bortezomib. Magnification: 400 ×; B. Cell apoptosis ratio of U266 and H929 cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h and 72 h, respectively. Cell apoptosis was evaluated by Annexin V/PI staining; C. Expression of PARP, caspase-3, and cleaved caspase-3 in U266 and H929 cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h; D. Cell apoptosis ratio of U266-R and H929-R cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h and 72 h, respectively; cell apoptosis was evaluated by Annexin V/PI staining; E. Expression of PARP, caspase-3, and cleaved caspase-3 in U266-R and H929-R cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h. U266-R, U266 bortezomib-resistant cells; H929-R, H929 bortezomib-resistant cells. The experiments were performed in triplicate. ** P
Figure Legend Snippet: The combination of 8-CPT-cAMP and bortezomib synergistically induced apoptosis in bortezomib-sensitive and -resistant multiple myeloma cells. A. Giemsa staining of U266, H929, U266-R, and H929-R cells following the treatment with 8-CPT-cAMP and bortezomib. Magnification: 400 ×; B. Cell apoptosis ratio of U266 and H929 cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h and 72 h, respectively. Cell apoptosis was evaluated by Annexin V/PI staining; C. Expression of PARP, caspase-3, and cleaved caspase-3 in U266 and H929 cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h; D. Cell apoptosis ratio of U266-R and H929-R cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h and 72 h, respectively; cell apoptosis was evaluated by Annexin V/PI staining; E. Expression of PARP, caspase-3, and cleaved caspase-3 in U266-R and H929-R cells exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h. U266-R, U266 bortezomib-resistant cells; H929-R, H929 bortezomib-resistant cells. The experiments were performed in triplicate. ** P

Techniques Used: Cycling Probe Technology, Staining, Expressing

PKA is involved in the cell growth inhibition and apoptosis induced by bortezomib and cAMP. A. Cell apoptosis ratio of H929 and H929-R cells exposed to 8-p-O-cAMP (Epac activator), bortezomib, or their combination for 48 h, respectively. Cell apoptosis was evaluated by Annexin V/PI staining; B. Cell apoptosis ratio of H929 and H929-R cells exposed to 6-bnz-cAMP (PKA activator), bortezomib, or their combination for 48 h, respectively. Apoptosis was evaluated by Annexin V/PI staining; C. H929 cells were exposed to bortezomib, 6-bnz-cAMP, or their combination for 48 h, and the expression of PARP and caspase-3 was measured by Western blot; D. H929-R cells were exposed to bortezomib, 6-bnz-cAMP, or their combination for 48 h, and the expression of PARP and caspase-3 was measured by Western blot; E. Knockdown of PKA in H929 cells by stably transfected with non-specific shRNA pairs (normal control, NC) or shRNA pairs against PKA-Cα; F. PKA-silenced H929 cells were exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h. Apoptosis was evaluated by Annexin V/PI staining; G. The expression of PARP and caspase-3 was measured by Western blot after U266 cells were treated with 8-CPT-cAMP, bortezomib, H89 (PKA inhibitor), or their combination for 48 h. H929-R, H929 bortezomib-resistant cells. Epac, exchange protein directly activated by cAMP; PKA, protein kinase A. ** P
Figure Legend Snippet: PKA is involved in the cell growth inhibition and apoptosis induced by bortezomib and cAMP. A. Cell apoptosis ratio of H929 and H929-R cells exposed to 8-p-O-cAMP (Epac activator), bortezomib, or their combination for 48 h, respectively. Cell apoptosis was evaluated by Annexin V/PI staining; B. Cell apoptosis ratio of H929 and H929-R cells exposed to 6-bnz-cAMP (PKA activator), bortezomib, or their combination for 48 h, respectively. Apoptosis was evaluated by Annexin V/PI staining; C. H929 cells were exposed to bortezomib, 6-bnz-cAMP, or their combination for 48 h, and the expression of PARP and caspase-3 was measured by Western blot; D. H929-R cells were exposed to bortezomib, 6-bnz-cAMP, or their combination for 48 h, and the expression of PARP and caspase-3 was measured by Western blot; E. Knockdown of PKA in H929 cells by stably transfected with non-specific shRNA pairs (normal control, NC) or shRNA pairs against PKA-Cα; F. PKA-silenced H929 cells were exposed to 8-CPT-cAMP, bortezomib, or their combination for 48 h. Apoptosis was evaluated by Annexin V/PI staining; G. The expression of PARP and caspase-3 was measured by Western blot after U266 cells were treated with 8-CPT-cAMP, bortezomib, H89 (PKA inhibitor), or their combination for 48 h. H929-R, H929 bortezomib-resistant cells. Epac, exchange protein directly activated by cAMP; PKA, protein kinase A. ** P

Techniques Used: Inhibition, Staining, Expressing, Western Blot, Stable Transfection, Transfection, shRNA, Cycling Probe Technology

The combination of 8-CPT-cAMP and bortezomib synergistically induced apoptosis in primary CD138 + MM cells. A. Primary CD138 + cells isolated from MM patients were exposed to 8-CPT-cAMP, bortezomib, or their combinations for 24 h; cell apoptosis was evaluated by Annexin V/PI staining; B. CD138 + MM cell co-cultured with bone marrow stromal cells (BMSCs) for 24 h and treated with 8-CPT-cAMP, bortezomib, or their combination for 24 h; cell apoptosis was evaluated by Annexin V/PI staining. * P
Figure Legend Snippet: The combination of 8-CPT-cAMP and bortezomib synergistically induced apoptosis in primary CD138 + MM cells. A. Primary CD138 + cells isolated from MM patients were exposed to 8-CPT-cAMP, bortezomib, or their combinations for 24 h; cell apoptosis was evaluated by Annexin V/PI staining; B. CD138 + MM cell co-cultured with bone marrow stromal cells (BMSCs) for 24 h and treated with 8-CPT-cAMP, bortezomib, or their combination for 24 h; cell apoptosis was evaluated by Annexin V/PI staining. * P

Techniques Used: Cycling Probe Technology, Isolation, Staining, Cell Culture

22) Product Images from "miR-146a controls the resolution of T cell responses in mice"

Article Title: miR-146a controls the resolution of T cell responses in mice

Journal: The Journal of Experimental Medicine

doi: 10.1084/jem.20112218

miR-146a–deficient T cells are hyperactive in chronic inflammatory response in vivo and promote the development of T cell–associated autoimmunity. (A) Schematic representation of the experimental design to study the role of miR-146a as an autonomous factor to regulate T cell response in a chronic T cell inflammatory autoimmune mouse disease model. (B) Flow cytometry analysis to quantify the number of WT and miR-146a −/− CD4 or CD8 T cells (gated as CD4 + and CD8 + , respectively) in the SP of recipient mice on day 29 after adoptive transfer. (C) BrdU incorporation analysis of the in vivo proliferation of WT and miR-146a −/− CD4 and CD8 T cells (gated as CD4 + BrdU + and CD8 + BrdU + , respectively) in the SP of recipient mice on day 29 after adoptive transfer. BrdU was injected into recipient mice 16 h before the analysis. (D) Annexin V staining analysis of the in vivo apoptosis of WT and miR-146a −/− CD4 and CD8 T cells (gated as CD4 + Annexin V + and CD8 + Annexin V + , respectively) in the SP of recipient mice on day 29 after adoptive transfer. (E and F) Immunohistology analysis of tissue sections from recipient mice on day 29 after adoptive transfer. Tissue sections from RAG1 −/− mice that did not receive cell transfer were included as control. (E) Hematoxylin and eosin staining. Arrows point to mononuclear cell infiltrates. (F) Anti–mouse CD3 staining. CD3 is shown in brown. Bars: (E, lung, liver, and kidney) 200 µm; (E, stomach) 400 µm; (F) 40 µm. Data are presented as mean ± SEM ( n = 4) and are representative of three independent experiments. *, P
Figure Legend Snippet: miR-146a–deficient T cells are hyperactive in chronic inflammatory response in vivo and promote the development of T cell–associated autoimmunity. (A) Schematic representation of the experimental design to study the role of miR-146a as an autonomous factor to regulate T cell response in a chronic T cell inflammatory autoimmune mouse disease model. (B) Flow cytometry analysis to quantify the number of WT and miR-146a −/− CD4 or CD8 T cells (gated as CD4 + and CD8 + , respectively) in the SP of recipient mice on day 29 after adoptive transfer. (C) BrdU incorporation analysis of the in vivo proliferation of WT and miR-146a −/− CD4 and CD8 T cells (gated as CD4 + BrdU + and CD8 + BrdU + , respectively) in the SP of recipient mice on day 29 after adoptive transfer. BrdU was injected into recipient mice 16 h before the analysis. (D) Annexin V staining analysis of the in vivo apoptosis of WT and miR-146a −/− CD4 and CD8 T cells (gated as CD4 + Annexin V + and CD8 + Annexin V + , respectively) in the SP of recipient mice on day 29 after adoptive transfer. (E and F) Immunohistology analysis of tissue sections from recipient mice on day 29 after adoptive transfer. Tissue sections from RAG1 −/− mice that did not receive cell transfer were included as control. (E) Hematoxylin and eosin staining. Arrows point to mononuclear cell infiltrates. (F) Anti–mouse CD3 staining. CD3 is shown in brown. Bars: (E, lung, liver, and kidney) 200 µm; (E, stomach) 400 µm; (F) 40 µm. Data are presented as mean ± SEM ( n = 4) and are representative of three independent experiments. *, P

Techniques Used: In Vivo, Flow Cytometry, Cytometry, Mouse Assay, Adoptive Transfer Assay, BrdU Incorporation Assay, Injection, Staining

miR-146a–deficient T cells are hyperresponsive to acute antigen stimulation in vivo. (A) Schematic representation of the experimental design to study the influence of miR-146a deficiency on OVA-specific OT1 T cells in response to antigen stimulation in vivo. (B–F) Flow cytometry analysis of the WT and miR-146a −/− OT1 T cells (gated as CD8 + Vβ5 + CD45.2 + ) in the PB of recipient CD45.1 congenic mice receiving 5 × 10 6 WT or miR-146a −/− OT1 Tg T cells (day −1) followed by prime (day 0) and boost (day 46) immunizations. (B) Percentage of WT and miR-146a −/− OT1 T cells of total CD8 + T cells at the indicated time points. (C) Representative contour plots showing the quantitation of WT and miR-146a −/− OT1 T cells (pregated on CD8 + ) at the indicated time points. (D) Surface expression of activation marker CD62L on WT and miR-146a −/− OT1 T cells on day 7 (7 d after prime immunization). Representative histogram plots and measurements of mean fluorescence intensity (MFI) are shown. (E) Apoptosis of WT and miR-146a −/− OT1 T cells on day 14 (14 d after prime immunization) measured by Annexin V staining. Representative histogram plots and measurements of the percentage of Annexin V + OT1 T cells are shown. (F) Phenotype of the WT and miR-146a −/− OT1 T cells on day 45 (45 d after prime immunization). Representative histogram plots are shown. Data are presented as mean ± SEM ( n = 8) and are representative of three independent experiments. *, P
Figure Legend Snippet: miR-146a–deficient T cells are hyperresponsive to acute antigen stimulation in vivo. (A) Schematic representation of the experimental design to study the influence of miR-146a deficiency on OVA-specific OT1 T cells in response to antigen stimulation in vivo. (B–F) Flow cytometry analysis of the WT and miR-146a −/− OT1 T cells (gated as CD8 + Vβ5 + CD45.2 + ) in the PB of recipient CD45.1 congenic mice receiving 5 × 10 6 WT or miR-146a −/− OT1 Tg T cells (day −1) followed by prime (day 0) and boost (day 46) immunizations. (B) Percentage of WT and miR-146a −/− OT1 T cells of total CD8 + T cells at the indicated time points. (C) Representative contour plots showing the quantitation of WT and miR-146a −/− OT1 T cells (pregated on CD8 + ) at the indicated time points. (D) Surface expression of activation marker CD62L on WT and miR-146a −/− OT1 T cells on day 7 (7 d after prime immunization). Representative histogram plots and measurements of mean fluorescence intensity (MFI) are shown. (E) Apoptosis of WT and miR-146a −/− OT1 T cells on day 14 (14 d after prime immunization) measured by Annexin V staining. Representative histogram plots and measurements of the percentage of Annexin V + OT1 T cells are shown. (F) Phenotype of the WT and miR-146a −/− OT1 T cells on day 45 (45 d after prime immunization). Representative histogram plots are shown. Data are presented as mean ± SEM ( n = 8) and are representative of three independent experiments. *, P

Techniques Used: In Vivo, Flow Cytometry, Cytometry, Mouse Assay, Quantitation Assay, Expressing, Activation Assay, Marker, Fluorescence, Staining

miR-146a regulates the various aspects of T cell activation induced by TCR stimulation. (A) [ 3 H]Thymidine incorporation analysis of proliferation of purified WT or miR-146a −/− CD4 and CD8 T cells stimulated with plate-bound anti-CD3 (10 µg/ml) + anti-CD28 (1 µg/ml) for 4 d. (B) Annexin V staining analysis for apoptosis of WT or miR-146a −/− CD4 and CD8 T cells (gated as CD4 + and CD8 + from the SP/LN cell culture, respectively) stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 4 d. (C) Flow cytometry analysis of surface activation marker expression on WT or miR-146a −/− CD4 and CD8 T cells (gated as CD4 + and CD8 + from the SP/LN cell culture, respectively) stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d. (D) ELISA analysis of effector cytokine production of purified WT or miR-146a −/− CD4 and CD8 T cells stimulated with plate-bound anti-CD3 (10 µg/ml) + anti-CD28 (1 µg/ml) for 4 d. (E) Flow cytometry analysis of surface activation marker expression on WT CD4 and CD8 T cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (gated as CD4 + GFP + and CD8 + GFP + from the SP/LN cell culture, respectively). (F) Relative cell number fold change of WT CD4 and CD8 T cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (gated as CD4 + and CD8 + from the SP/LN cell culture, respectively; counts of T cells transduced with MIG = 1). (G) Annexin V staining of WT CD4 and CD8 T cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 4 d (gated as CD4 + GFP + and CD8 + GFP + from the SP/LN cell culture, respectively). (H) ELISA analysis of effector cytokine production of WT SP/LN cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 4 d. (I) TaqMan Q-PCR analysis of the expression of NF-κB responsive genes in WT and miR-146a −/− SP/LN cells stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (top) or in WT SP/LN cells transduced with either MIG-miR-146a or control MIG retroviral vector and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (bottom). RE, relative expression. (J) TaqMan Q-PCR analysis of the time course expression of NF-κB responsive genes in WT and miR-146a −/− SP/LN cells stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 5 d. Data are presented as mean of duplicate culture ± SEM (A, B, D, and F–I) and are representative of three independent experiments. *, P
Figure Legend Snippet: miR-146a regulates the various aspects of T cell activation induced by TCR stimulation. (A) [ 3 H]Thymidine incorporation analysis of proliferation of purified WT or miR-146a −/− CD4 and CD8 T cells stimulated with plate-bound anti-CD3 (10 µg/ml) + anti-CD28 (1 µg/ml) for 4 d. (B) Annexin V staining analysis for apoptosis of WT or miR-146a −/− CD4 and CD8 T cells (gated as CD4 + and CD8 + from the SP/LN cell culture, respectively) stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 4 d. (C) Flow cytometry analysis of surface activation marker expression on WT or miR-146a −/− CD4 and CD8 T cells (gated as CD4 + and CD8 + from the SP/LN cell culture, respectively) stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d. (D) ELISA analysis of effector cytokine production of purified WT or miR-146a −/− CD4 and CD8 T cells stimulated with plate-bound anti-CD3 (10 µg/ml) + anti-CD28 (1 µg/ml) for 4 d. (E) Flow cytometry analysis of surface activation marker expression on WT CD4 and CD8 T cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (gated as CD4 + GFP + and CD8 + GFP + from the SP/LN cell culture, respectively). (F) Relative cell number fold change of WT CD4 and CD8 T cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (gated as CD4 + and CD8 + from the SP/LN cell culture, respectively; counts of T cells transduced with MIG = 1). (G) Annexin V staining of WT CD4 and CD8 T cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 4 d (gated as CD4 + GFP + and CD8 + GFP + from the SP/LN cell culture, respectively). (H) ELISA analysis of effector cytokine production of WT SP/LN cells transduced with MIG–miR-146a or control MIG retroviral vectors and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 4 d. (I) TaqMan Q-PCR analysis of the expression of NF-κB responsive genes in WT and miR-146a −/− SP/LN cells stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (top) or in WT SP/LN cells transduced with either MIG-miR-146a or control MIG retroviral vector and stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 3 d (bottom). RE, relative expression. (J) TaqMan Q-PCR analysis of the time course expression of NF-κB responsive genes in WT and miR-146a −/− SP/LN cells stimulated with soluble anti-CD3 + anti-CD28 (1 µg/ml each) for 5 d. Data are presented as mean of duplicate culture ± SEM (A, B, D, and F–I) and are representative of three independent experiments. *, P

Techniques Used: Activation Assay, Purification, Staining, Cell Culture, Flow Cytometry, Cytometry, Marker, Expressing, Enzyme-linked Immunosorbent Assay, Transduction, Polymerase Chain Reaction, Plasmid Preparation

23) Product Images from "Ruxolitinib synergistically enhances the anti-tumor activity of paclitaxel in human ovarian cancer"

Article Title: Ruxolitinib synergistically enhances the anti-tumor activity of paclitaxel in human ovarian cancer

Journal: Oncotarget

doi: 10.18632/oncotarget.24368

Dose dependent induction of apoptosis (A) and (B) OVCAR-8 and MDAH 2774 cells were incubated with various concentrations of ruxolitinib for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A) or using cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (B). * P
Figure Legend Snippet: Dose dependent induction of apoptosis (A) and (B) OVCAR-8 and MDAH 2774 cells were incubated with various concentrations of ruxolitinib for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A) or using cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (B). * P

Techniques Used: Incubation, Flow Cytometry, Cytometry, Staining, Western Blot

Ruxolitinib enhanced paclitaxel-induced apoptosis in human ovarian cancer cells (A) OVCAR-8 and (B) MDAH2774 cells were treated with ruxolitinib (30μM), paclitaxel (10nM) either alone or together, for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A B) or by cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (C) . ns: not significant; # P
Figure Legend Snippet: Ruxolitinib enhanced paclitaxel-induced apoptosis in human ovarian cancer cells (A) OVCAR-8 and (B) MDAH2774 cells were treated with ruxolitinib (30μM), paclitaxel (10nM) either alone or together, for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A B) or by cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (C) . ns: not significant; # P

Techniques Used: Flow Cytometry, Cytometry, Staining, Western Blot

24) Product Images from "High Expression of hTERT and Stemness Genes in BORIS/CTCFL Positive Cells Isolated from Embryonic Cancer Cells"

Article Title: High Expression of hTERT and Stemness Genes in BORIS/CTCFL Positive Cells Isolated from Embryonic Cancer Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0109921

BORIS knockdown impairs cell senescence but not apoptosis. (A) Cell proliferation over 1 moth of dox-induced BORIS shRNA cells were analyzed by MTT assay. Results of the two specific BORIS-shRNA (BORIS sh-3 and sh-4) NCCIT-derived cells are indicated as a percentage compared to the cell proliferation of control cells (scrambled shRNA, CTR sh). Error bars represent the mean ± SD of 3 independent experiments. (B) After dox-induction of the BORIS specific shRNA in NCCIT cells, apoptosis was tested at each week using Annexin V Apoptosis Detection Kit. Results show the percentage of apoptotic cells (late apoptotic AnnexinV + /7AAD + and early apoptotic AnnexinV + /7AAD − ) of BORIS sh-3 and sh-4 cells compared to the control cells. Error bars represent the mean ± SD of 3 experiments. (C) The senescence-associated β-galactosidase (SA-β-gal) staining was performed using β-galactosidase staining kit. SA-β-gal was analyzed after 2 and 4 weeks of dox-induction of the BORIS specific shRNA in NCCIT cells. Results show the percentage of senescent cells of BORIS sh-3, BORIS sh4 and control cells. Error bars represent the mean ± SD of 3 experiments. Asterisks indicate statistically significant difference (p
Figure Legend Snippet: BORIS knockdown impairs cell senescence but not apoptosis. (A) Cell proliferation over 1 moth of dox-induced BORIS shRNA cells were analyzed by MTT assay. Results of the two specific BORIS-shRNA (BORIS sh-3 and sh-4) NCCIT-derived cells are indicated as a percentage compared to the cell proliferation of control cells (scrambled shRNA, CTR sh). Error bars represent the mean ± SD of 3 independent experiments. (B) After dox-induction of the BORIS specific shRNA in NCCIT cells, apoptosis was tested at each week using Annexin V Apoptosis Detection Kit. Results show the percentage of apoptotic cells (late apoptotic AnnexinV + /7AAD + and early apoptotic AnnexinV + /7AAD − ) of BORIS sh-3 and sh-4 cells compared to the control cells. Error bars represent the mean ± SD of 3 experiments. (C) The senescence-associated β-galactosidase (SA-β-gal) staining was performed using β-galactosidase staining kit. SA-β-gal was analyzed after 2 and 4 weeks of dox-induction of the BORIS specific shRNA in NCCIT cells. Results show the percentage of senescent cells of BORIS sh-3, BORIS sh4 and control cells. Error bars represent the mean ± SD of 3 experiments. Asterisks indicate statistically significant difference (p

Techniques Used: shRNA, MTT Assay, Derivative Assay, Staining

25) Product Images from "p15INK4b plays a crucial role in murine lymphoid development and tumorigenesis"

Article Title: p15INK4b plays a crucial role in murine lymphoid development and tumorigenesis

Journal: Carcinogenesis

doi: 10.1093/carcin/bgs003

Modulation of apoptosis by Rgr and p15 status. ( A ) Overall characterization of the apoptotic stages (no apoptosis, Annexin V − /7AAD − ; early apoptosis, Annexin V + /7AAD − ; and late apoptosis Annexin V + /7AAD + ) as determined by subjecting
Figure Legend Snippet: Modulation of apoptosis by Rgr and p15 status. ( A ) Overall characterization of the apoptotic stages (no apoptosis, Annexin V − /7AAD − ; early apoptosis, Annexin V + /7AAD − ; and late apoptosis Annexin V + /7AAD + ) as determined by subjecting

Techniques Used:

26) Product Images from "Ectopic Otoconin 90 expression in triple negative breast cancer cell lines is associated with metastasis functions"

Article Title: Ectopic Otoconin 90 expression in triple negative breast cancer cell lines is associated with metastasis functions

Journal: PLoS ONE

doi: 10.1371/journal.pone.0211737

OC90 siRNA treatment promotes apoptosis in HCC38 cells. A) Flow cytometry graphs showing distribution of HCC38 cells based on Annexin V (PE, X-axis) and 7-AAD (PerCP-Cy5.5, Y-axis) intensity 72 hours after control siRNA or OC90 siRNA treatment. B) C) Western blot showing increase in cleaved Caspase 3 7 (respectively) in HCC38 cell line after OC90 knockdown. Based on these findings, significant reduction in cellular viability in TNBC cell lines after OC90 knockdown may be due to its function as an anti-apoptotic gene in TNBC and may also have some role in cellular proliferation.
Figure Legend Snippet: OC90 siRNA treatment promotes apoptosis in HCC38 cells. A) Flow cytometry graphs showing distribution of HCC38 cells based on Annexin V (PE, X-axis) and 7-AAD (PerCP-Cy5.5, Y-axis) intensity 72 hours after control siRNA or OC90 siRNA treatment. B) C) Western blot showing increase in cleaved Caspase 3 7 (respectively) in HCC38 cell line after OC90 knockdown. Based on these findings, significant reduction in cellular viability in TNBC cell lines after OC90 knockdown may be due to its function as an anti-apoptotic gene in TNBC and may also have some role in cellular proliferation.

Techniques Used: Flow Cytometry, Cytometry, Western Blot

27) Product Images from "The Effects of Silica Nanoparticles on Apoptosis and Autophagy of Glioblastoma Cell Lines"

Article Title: The Effects of Silica Nanoparticles on Apoptosis and Autophagy of Glioblastoma Cell Lines

Journal: Nanomaterials

doi: 10.3390/nano7080230

The effect of SiNPs on apoptosis ( A , B , D , E ) and necrosis ( A , C , D , F ) of LBC3 ( A – C ) and LN-18 ( D – F ) cell lines evaluated by annexin V assay. The cells were incubated for 24 and 48 h in DMEM with 50 and 100 μg/mL of 5–15 nm SiNPs. The cells were double-stained with FITC-Annexin V and PI. Representative dot plots for Annexin V-FITC/propidium iodide (PI) staining are shown ( A , D ). Following acquisition of sample, the cells were gated through the forward scatter FSC and side scatter SSC and analyzed for fluorescence intensity of FITC-Annexin V and PI. The cells were divided into four subpopulations: live cells—Q3 (annexin V-FITC−/PI−), early apoptotic cells—Q4 (annexin V FITC+/PI−), late apoptotic cells—Q2 (annexin V-FITC+/PI+), and necrotic cells—Q1 (annexin V FITC−/ PI+). Percentage of apoptotic cells was the sum of percentage early apoptotic (Q4) and late apoptotic cells (Q2). Mean values of the percentage of apoptotic and necrotic cells, from three independent experiments ± SD are presented. Significant alterations are expressed relative to controls and marked with asterisks. Statistical significance was considered if * p
Figure Legend Snippet: The effect of SiNPs on apoptosis ( A , B , D , E ) and necrosis ( A , C , D , F ) of LBC3 ( A – C ) and LN-18 ( D – F ) cell lines evaluated by annexin V assay. The cells were incubated for 24 and 48 h in DMEM with 50 and 100 μg/mL of 5–15 nm SiNPs. The cells were double-stained with FITC-Annexin V and PI. Representative dot plots for Annexin V-FITC/propidium iodide (PI) staining are shown ( A , D ). Following acquisition of sample, the cells were gated through the forward scatter FSC and side scatter SSC and analyzed for fluorescence intensity of FITC-Annexin V and PI. The cells were divided into four subpopulations: live cells—Q3 (annexin V-FITC−/PI−), early apoptotic cells—Q4 (annexin V FITC+/PI−), late apoptotic cells—Q2 (annexin V-FITC+/PI+), and necrotic cells—Q1 (annexin V FITC−/ PI+). Percentage of apoptotic cells was the sum of percentage early apoptotic (Q4) and late apoptotic cells (Q2). Mean values of the percentage of apoptotic and necrotic cells, from three independent experiments ± SD are presented. Significant alterations are expressed relative to controls and marked with asterisks. Statistical significance was considered if * p

Techniques Used: Annexin V Assay, Incubation, Staining, Fluorescence, IF-P

28) Product Images from "REST Controls Self-Renewal and Tumorigenic Competence of Human Glioblastoma Cells"

Article Title: REST Controls Self-Renewal and Tumorigenic Competence of Human Glioblastoma Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0038486

REST knockdown reduces self-renewal potential of human tumorigenic-competent GBM cells and induces neuronal differentiation and cell death programs. (A) Representative live image of GB7 cells maintained in self-renewal conditions, 4 and 7 days after REST shRNA knockdown (shREST) and relative controls (CTRL: mock infected cells; NT shRNA: non targeting shRNA). REST knockdown causes reduction of proliferation and dramatic morphological changes, with appearance of a large proportion of flat and differentiated cells (arrows). (B) Colorimetric MTT-based cell viability assay performed on controls and shREST GB cell cultures. (C) Immunofluorescent analyses of controls and shREST self-renewing GB7 cells (7 days post infection). There is an evident reduction in the expression of neural progenitor (Nestin) and proliferation (P- HisH3) markers in shREST cells with respect to controls, with a parallel increase of neuronal differentiation (β3-tubulin) and apoptosis (Activated Caspase 3) markers. (D) Relative quantification of the numbers of immunoreactive cells in (C). CTRL: light gray bars; NT shRNA: dark gray bars; shREST: black bars. At least 700 cells per group were scored. (E) Detection of apoptosis on REST depleted GB cells. Seven days after siRNA transfection, the percentages of apoptotic GB7 cells in the cultures were analysed by FACS Annexin V–PE assay. (F) Luminometric detection of apoptosis on REST depleted GB cells. 48 and 72 hours after siRNA transfection, apoptosis in controls and siREST treated GB7 cells was analysed by Caspase-Glo 3/7 Assay. Data were analyzed by one-way ANOVA comparison of the controls groups and of the REST knockdown cultures. Results shown are relative to three independent experiments. Data are means ± s.d. ns not significant, * P
Figure Legend Snippet: REST knockdown reduces self-renewal potential of human tumorigenic-competent GBM cells and induces neuronal differentiation and cell death programs. (A) Representative live image of GB7 cells maintained in self-renewal conditions, 4 and 7 days after REST shRNA knockdown (shREST) and relative controls (CTRL: mock infected cells; NT shRNA: non targeting shRNA). REST knockdown causes reduction of proliferation and dramatic morphological changes, with appearance of a large proportion of flat and differentiated cells (arrows). (B) Colorimetric MTT-based cell viability assay performed on controls and shREST GB cell cultures. (C) Immunofluorescent analyses of controls and shREST self-renewing GB7 cells (7 days post infection). There is an evident reduction in the expression of neural progenitor (Nestin) and proliferation (P- HisH3) markers in shREST cells with respect to controls, with a parallel increase of neuronal differentiation (β3-tubulin) and apoptosis (Activated Caspase 3) markers. (D) Relative quantification of the numbers of immunoreactive cells in (C). CTRL: light gray bars; NT shRNA: dark gray bars; shREST: black bars. At least 700 cells per group were scored. (E) Detection of apoptosis on REST depleted GB cells. Seven days after siRNA transfection, the percentages of apoptotic GB7 cells in the cultures were analysed by FACS Annexin V–PE assay. (F) Luminometric detection of apoptosis on REST depleted GB cells. 48 and 72 hours after siRNA transfection, apoptosis in controls and siREST treated GB7 cells was analysed by Caspase-Glo 3/7 Assay. Data were analyzed by one-way ANOVA comparison of the controls groups and of the REST knockdown cultures. Results shown are relative to three independent experiments. Data are means ± s.d. ns not significant, * P

Techniques Used: shRNA, Infection, MTT Assay, Viability Assay, Expressing, Transfection, FACS, Caspase-Glo Assay

29) Product Images from "Dendritic Cells Crosspresent Antigens from Live B16 Cells More Efficiently than from Apoptotic Cells and Protect from Melanoma in a Therapeutic Model"

Article Title: Dendritic Cells Crosspresent Antigens from Live B16 Cells More Efficiently than from Apoptotic Cells and Protect from Melanoma in a Therapeutic Model

Journal: PLoS ONE

doi: 10.1371/journal.pone.0019104

DC cultured with live B16 cells induced strong protection against tumor in a therapeutic setting. A, time schedule outlining the different stages as used in the experiments. Five C57BL/6 mice per group were inoculated i.v. with 1.10 6 B16 cells at day 0. At days 3 and 10, mice were immunized with 0,5.10 6 of purified and irradiated DC that were previously cultured for 16 h in the presence of LPS and IFNγ, with either culture medium (DC), or gp100 25–33 and TRP2 181–188 peptides (DC-peptides), or live (zVAD treated) B16 cells (DC-B16 zVAD) or apoptotic (γ-irradiated) B16 cells (DC-B16γ). B, B16 cell apoptosis was evaluated by annexin-V staining (solid lines) compared to no staining (dotted lines) and the mean number ± SEM of the Annexin-positive events is shown. At day 15, mice were sacrificed, lung tumors were counted (C) and T cell responses were evaluated in the spleen. D, Splenocytes were restimulated with gp100 25–33 or TRP2 181–188 peptides or B16 cells or culture medium and tested in an IFNγ ELISPOT assay. E, After B16 restimulation, splenocytes were also stimulated with PMA and ionomycin and then labeled with anti-CD3, anti-CD8 and intracellularly with anti-IFNγ antibodies. Events were gated on CD3 + CD8 + splenocytes and IFNγ production was mesured by flow cytometry. The mean ± SEM of data from 5 mice per group are shown for an experiment representative of two independent experiments. The significance of differences between series was assessed by unpaired t test.
Figure Legend Snippet: DC cultured with live B16 cells induced strong protection against tumor in a therapeutic setting. A, time schedule outlining the different stages as used in the experiments. Five C57BL/6 mice per group were inoculated i.v. with 1.10 6 B16 cells at day 0. At days 3 and 10, mice were immunized with 0,5.10 6 of purified and irradiated DC that were previously cultured for 16 h in the presence of LPS and IFNγ, with either culture medium (DC), or gp100 25–33 and TRP2 181–188 peptides (DC-peptides), or live (zVAD treated) B16 cells (DC-B16 zVAD) or apoptotic (γ-irradiated) B16 cells (DC-B16γ). B, B16 cell apoptosis was evaluated by annexin-V staining (solid lines) compared to no staining (dotted lines) and the mean number ± SEM of the Annexin-positive events is shown. At day 15, mice were sacrificed, lung tumors were counted (C) and T cell responses were evaluated in the spleen. D, Splenocytes were restimulated with gp100 25–33 or TRP2 181–188 peptides or B16 cells or culture medium and tested in an IFNγ ELISPOT assay. E, After B16 restimulation, splenocytes were also stimulated with PMA and ionomycin and then labeled with anti-CD3, anti-CD8 and intracellularly with anti-IFNγ antibodies. Events were gated on CD3 + CD8 + splenocytes and IFNγ production was mesured by flow cytometry. The mean ± SEM of data from 5 mice per group are shown for an experiment representative of two independent experiments. The significance of differences between series was assessed by unpaired t test.

Techniques Used: Cell Culture, Mouse Assay, Purification, Irradiation, Staining, Enzyme-linked Immunospot, Labeling, Flow Cytometry, Cytometry

30) Product Images from "Loss of Perp in T Cells Promotes Resistance to Apoptosis of T Helper 17 Cells and Exacerbates the Development of Experimental Autoimmune Encephalomyelitis in Mice"

Article Title: Loss of Perp in T Cells Promotes Resistance to Apoptosis of T Helper 17 Cells and Exacerbates the Development of Experimental Autoimmune Encephalomyelitis in Mice

Journal: Frontiers in Immunology

doi: 10.3389/fimmu.2018.00842

Perp −/− in T cells promotes the resistance to apoptosis in T helper 17 (Th17) cells. CD4 + T cells were isolated from littermate control, Lck-Cre × Perp fl/+ and Lck-Cre × Perp fl/fl mice by negative selection and stimulated with anti-CD3/anti-CD28 in the presence of cocktail of cytokines and antibodies to induce Th1, Th17, or Treg cell differentiation for 3 days. The cells were stained with anti-CD4 and/or anti-CD25, fixed, permeabilized, and intracellularly stained with anti-IFN-γ, anti-IL-17A, or anti-Foxp3. The frequency of Th1, Th17, or Treg cells was determined by FACS. Furthermore, following in vitro differentiation for 3 days, the cells were re-stimulated with anti-CD3/anti-CD28 in the presence or absence of anti-Fas for 72 h. The percentages of apoptotic Th1, Th17, or Treg cells were determined by FACS using Annexin V-PE and 7-AAD staining. Data are representative images or expressed as the mean ± SEM of each group from three separate experiments. (A,B) Th1 cell differentiation. (C,D) Th17 cell differentiation. (E,F) Treg cell differentiation. (G,J) Th1 cell apoptosis. (H,K) Th17 cell apoptosis. (I,L) Th17 cell apoptosis (* P
Figure Legend Snippet: Perp −/− in T cells promotes the resistance to apoptosis in T helper 17 (Th17) cells. CD4 + T cells were isolated from littermate control, Lck-Cre × Perp fl/+ and Lck-Cre × Perp fl/fl mice by negative selection and stimulated with anti-CD3/anti-CD28 in the presence of cocktail of cytokines and antibodies to induce Th1, Th17, or Treg cell differentiation for 3 days. The cells were stained with anti-CD4 and/or anti-CD25, fixed, permeabilized, and intracellularly stained with anti-IFN-γ, anti-IL-17A, or anti-Foxp3. The frequency of Th1, Th17, or Treg cells was determined by FACS. Furthermore, following in vitro differentiation for 3 days, the cells were re-stimulated with anti-CD3/anti-CD28 in the presence or absence of anti-Fas for 72 h. The percentages of apoptotic Th1, Th17, or Treg cells were determined by FACS using Annexin V-PE and 7-AAD staining. Data are representative images or expressed as the mean ± SEM of each group from three separate experiments. (A,B) Th1 cell differentiation. (C,D) Th17 cell differentiation. (E,F) Treg cell differentiation. (G,J) Th1 cell apoptosis. (H,K) Th17 cell apoptosis. (I,L) Th17 cell apoptosis (* P

Techniques Used: Isolation, Mouse Assay, Selection, Cell Differentiation, Staining, FACS, In Vitro

31) Product Images from "GZD824 suppresses the growth of human B cell precursor acute lymphoblastic leukemia cells by inhibiting the SRC kinase and PI3K/AKT pathways"

Article Title: GZD824 suppresses the growth of human B cell precursor acute lymphoblastic leukemia cells by inhibiting the SRC kinase and PI3K/AKT pathways

Journal: Oncotarget

doi: 10.18632/oncotarget.10881

GZD824 induces apoptosis of primary pre-B ALL cells from patients with no toxicity to normal B cells A. GZD824 cytotoxicity in primary pre-B ALL cells: primary pre-B ALL cells of P#1, P#2, and P#3 were from Ph- pre-B ALL, and primary pre-B ALL cells of P#4 and P#5 were from Ph+ pre-B ALL. Up: Representative flow cytometric analysis of primary pre-B ALL cells treated with DMSO or 1μM GZD824. Bottom: Statistical analysis of Annexin V-positive cells in GZD824 treated primary pre-B ALL cells. (P#1, P#2, P#3, P#4 and P#5 are short for patient #1, patient #2, patient #3, patient #4, and patient #5) B. Left: Representative flow cytometric analysis of normal B cells treated with DMSO or 1μM GZD824. Right: Statistical analysis of AnnexinV-positive cells in GZD824 treated primary B cells. Data are shown as the mean ± SEM (error bars) from three independent experiments. Significance values: * P
Figure Legend Snippet: GZD824 induces apoptosis of primary pre-B ALL cells from patients with no toxicity to normal B cells A. GZD824 cytotoxicity in primary pre-B ALL cells: primary pre-B ALL cells of P#1, P#2, and P#3 were from Ph- pre-B ALL, and primary pre-B ALL cells of P#4 and P#5 were from Ph+ pre-B ALL. Up: Representative flow cytometric analysis of primary pre-B ALL cells treated with DMSO or 1μM GZD824. Bottom: Statistical analysis of Annexin V-positive cells in GZD824 treated primary pre-B ALL cells. (P#1, P#2, P#3, P#4 and P#5 are short for patient #1, patient #2, patient #3, patient #4, and patient #5) B. Left: Representative flow cytometric analysis of normal B cells treated with DMSO or 1μM GZD824. Right: Statistical analysis of AnnexinV-positive cells in GZD824 treated primary B cells. Data are shown as the mean ± SEM (error bars) from three independent experiments. Significance values: * P

Techniques Used: Flow Cytometry

32) Product Images from "Loss of Perp in T Cells Promotes Resistance to Apoptosis of T Helper 17 Cells and Exacerbates the Development of Experimental Autoimmune Encephalomyelitis in Mice"

Article Title: Loss of Perp in T Cells Promotes Resistance to Apoptosis of T Helper 17 Cells and Exacerbates the Development of Experimental Autoimmune Encephalomyelitis in Mice

Journal: Frontiers in Immunology

doi: 10.3389/fimmu.2018.00842

Perp −/− in T cells promotes the resistance to apoptosis in T helper 17 (Th17) cells. CD4 + T cells were isolated from littermate control, Lck-Cre × Perp fl/+ and Lck-Cre × Perp fl/fl mice by negative selection and stimulated with anti-CD3/anti-CD28 in the presence of cocktail of cytokines and antibodies to induce Th1, Th17, or Treg cell differentiation for 3 days. The cells were stained with anti-CD4 and/or anti-CD25, fixed, permeabilized, and intracellularly stained with anti-IFN-γ, anti-IL-17A, or anti-Foxp3. The frequency of Th1, Th17, or Treg cells was determined by FACS. Furthermore, following in vitro differentiation for 3 days, the cells were re-stimulated with anti-CD3/anti-CD28 in the presence or absence of anti-Fas for 72 h. The percentages of apoptotic Th1, Th17, or Treg cells were determined by FACS using Annexin V-PE and 7-AAD staining. Data are representative images or expressed as the mean ± SEM of each group from three separate experiments. (A,B) Th1 cell differentiation. (C,D) Th17 cell differentiation. (E,F) Treg cell differentiation. (G,J) Th1 cell apoptosis. (H,K) Th17 cell apoptosis. (I,L) Th17 cell apoptosis (* P
Figure Legend Snippet: Perp −/− in T cells promotes the resistance to apoptosis in T helper 17 (Th17) cells. CD4 + T cells were isolated from littermate control, Lck-Cre × Perp fl/+ and Lck-Cre × Perp fl/fl mice by negative selection and stimulated with anti-CD3/anti-CD28 in the presence of cocktail of cytokines and antibodies to induce Th1, Th17, or Treg cell differentiation for 3 days. The cells were stained with anti-CD4 and/or anti-CD25, fixed, permeabilized, and intracellularly stained with anti-IFN-γ, anti-IL-17A, or anti-Foxp3. The frequency of Th1, Th17, or Treg cells was determined by FACS. Furthermore, following in vitro differentiation for 3 days, the cells were re-stimulated with anti-CD3/anti-CD28 in the presence or absence of anti-Fas for 72 h. The percentages of apoptotic Th1, Th17, or Treg cells were determined by FACS using Annexin V-PE and 7-AAD staining. Data are representative images or expressed as the mean ± SEM of each group from three separate experiments. (A,B) Th1 cell differentiation. (C,D) Th17 cell differentiation. (E,F) Treg cell differentiation. (G,J) Th1 cell apoptosis. (H,K) Th17 cell apoptosis. (I,L) Th17 cell apoptosis (* P

Techniques Used: Isolation, Mouse Assay, Selection, Cell Differentiation, Staining, FACS, In Vitro

33) Product Images from "The nuclear receptor corepressor NCoR1 regulates hematopoiesis and leukemogenesis in vivo"

Article Title: The nuclear receptor corepressor NCoR1 regulates hematopoiesis and leukemogenesis in vivo

Journal: Blood Advances

doi: 10.1182/bloodadvances.2018022756

NCoR1 deficiency promotes HSC proliferation. (A) Apoptosis analysis of LT-HSCs and LSK cells in control and Vav-iCre + , NCoR1 f/f mice (n = 6 per genotype). Representative FACS profiles (left panel) and the frequency of Annexin V + cells (right panel) are shown. (B) Cell cycle analysis of LT-HSCs and LSK cells in control and Vav-iCre + , NCoR1 f/f mice (n = 5 per genotype). Representative FACS profiles (left panel) and the frequency of BrdU + cells (right panel) are shown. (C) Cell cycle analysis of LT-HSCs and LSK cells in control and Vav-iCre + , NCoR1 f/f mice (n = 5 per genotype). Representative FACS profiles (left panel) and the frequency of cell cycle distribution (left and middle panels) are shown. (D) Real-time qPCR analysis of the expression of cycling-dependent kinase inhibitors genes (n = 3). mRNA levels were normalized to Gapdh expression. Data are mean ± standard error of the mean from 3 independent experiments. ** P
Figure Legend Snippet: NCoR1 deficiency promotes HSC proliferation. (A) Apoptosis analysis of LT-HSCs and LSK cells in control and Vav-iCre + , NCoR1 f/f mice (n = 6 per genotype). Representative FACS profiles (left panel) and the frequency of Annexin V + cells (right panel) are shown. (B) Cell cycle analysis of LT-HSCs and LSK cells in control and Vav-iCre + , NCoR1 f/f mice (n = 5 per genotype). Representative FACS profiles (left panel) and the frequency of BrdU + cells (right panel) are shown. (C) Cell cycle analysis of LT-HSCs and LSK cells in control and Vav-iCre + , NCoR1 f/f mice (n = 5 per genotype). Representative FACS profiles (left panel) and the frequency of cell cycle distribution (left and middle panels) are shown. (D) Real-time qPCR analysis of the expression of cycling-dependent kinase inhibitors genes (n = 3). mRNA levels were normalized to Gapdh expression. Data are mean ± standard error of the mean from 3 independent experiments. ** P

Techniques Used: Mouse Assay, FACS, Cell Cycle Assay, Real-time Polymerase Chain Reaction, Expressing

34) Product Images from "The PLAG1-GDH1 axis promotes anoikis resistance and tumor metastasis through CamKK2-AMPK signaling in LKB1-deficient lung cancer"

Article Title: The PLAG1-GDH1 axis promotes anoikis resistance and tumor metastasis through CamKK2-AMPK signaling in LKB1-deficient lung cancer

Journal: Molecular cell

doi: 10.1016/j.molcel.2017.11.025

Loss of GDH1 and its product α-KG sensitizes LKB1-deficient lung cancer cells to anoikis induction and attenuates tumor metastasis in vivo (A) Effect of GDH1 knockdown on detachment-induced apoptosis in a panel of LKB1 null lung cancer cells. Cells were cultured on 1% agar followed by annexin V staining ( top ) and caspase 3/7 activity assay ( bottom ). (B) Rescue effect of α-KG on anoikis resistance in A549 cells with GDH1 knockdown. Cells were cultured under detachment conditions in the presence and absence of methyl-αKG and intracellular α-KG level and anoikis were determined. (C) Effect of α-KG, succinate, malate, and fumarate on anoikis resistance of GDH1 knockdown cells. Detached cells were cultured in the presence or absence of cell permeable metabolites, followed by annexin V staining. (D) Effect of shRNA-resistant GDH1 wild-type (WT) or enzyme-dead mutant GDH1 R443S expression on anoikis resistance in GDH1 knockdown cells. (E) Effect of α-KG producing enzymes GPT2, GOT2, and IDH2 on α-KG and anoikis in GDH1 knockdown cells. (F–G) Effect of GDH1 knockdown on tumor metastasis potential in A549 and H460 xenograft mice in an experimental metastasis model. Western blot analysis of GDH1 in injected A549-GFP-luciferase or H460 cells ( top left panels ). Nude mice were injected with A549-GFP-luciferase cells with or without GDH1 knockdown and average photonic flux and bioluminescence images of each group at week 7 are shown (F). NSG mice were injected with H460 cells harboring GDH1 shRNA or control vector and number of metastatic nodule in livers and representative liver and lung images of each group at day 18 are shown (G). Bars represent 5 mm for morphology and 1 mm for H E staining. Data are mean ± SD of three technical replicates and are representative of three (A, C), four (B) or two (D and E) independent biological experiments. For (F-G), data are mean ± SEM and reflect a single cohort experiment (n=8). p values were determined by a two-tailed Student’s t .
Figure Legend Snippet: Loss of GDH1 and its product α-KG sensitizes LKB1-deficient lung cancer cells to anoikis induction and attenuates tumor metastasis in vivo (A) Effect of GDH1 knockdown on detachment-induced apoptosis in a panel of LKB1 null lung cancer cells. Cells were cultured on 1% agar followed by annexin V staining ( top ) and caspase 3/7 activity assay ( bottom ). (B) Rescue effect of α-KG on anoikis resistance in A549 cells with GDH1 knockdown. Cells were cultured under detachment conditions in the presence and absence of methyl-αKG and intracellular α-KG level and anoikis were determined. (C) Effect of α-KG, succinate, malate, and fumarate on anoikis resistance of GDH1 knockdown cells. Detached cells were cultured in the presence or absence of cell permeable metabolites, followed by annexin V staining. (D) Effect of shRNA-resistant GDH1 wild-type (WT) or enzyme-dead mutant GDH1 R443S expression on anoikis resistance in GDH1 knockdown cells. (E) Effect of α-KG producing enzymes GPT2, GOT2, and IDH2 on α-KG and anoikis in GDH1 knockdown cells. (F–G) Effect of GDH1 knockdown on tumor metastasis potential in A549 and H460 xenograft mice in an experimental metastasis model. Western blot analysis of GDH1 in injected A549-GFP-luciferase or H460 cells ( top left panels ). Nude mice were injected with A549-GFP-luciferase cells with or without GDH1 knockdown and average photonic flux and bioluminescence images of each group at week 7 are shown (F). NSG mice were injected with H460 cells harboring GDH1 shRNA or control vector and number of metastatic nodule in livers and representative liver and lung images of each group at day 18 are shown (G). Bars represent 5 mm for morphology and 1 mm for H E staining. Data are mean ± SD of three technical replicates and are representative of three (A, C), four (B) or two (D and E) independent biological experiments. For (F-G), data are mean ± SEM and reflect a single cohort experiment (n=8). p values were determined by a two-tailed Student’s t .

Techniques Used: In Vivo, Cell Culture, Staining, Activity Assay, shRNA, Mutagenesis, Expressing, Mouse Assay, Western Blot, Injection, Luciferase, Plasmid Preparation, Two Tailed Test

35) Product Images from "Dengue Virus Type 3 Isolated from a Fatal Case with Visceral Complications Induces Enhanced Proinflammatory Responses and Apoptosis of Human Dendritic Cells ▿Dengue Virus Type 3 Isolated from a Fatal Case with Visceral Complications Induces Enhanced Proinflammatory Responses and Apoptosis of Human Dendritic Cells ▿ †"

Article Title: Dengue Virus Type 3 Isolated from a Fatal Case with Visceral Complications Induces Enhanced Proinflammatory Responses and Apoptosis of Human Dendritic Cells ▿Dengue Virus Type 3 Isolated from a Fatal Case with Visceral Complications Induces Enhanced Proinflammatory Responses and Apoptosis of Human Dendritic Cells ▿ †

Journal: Journal of Virology

doi: 10.1128/JVI.01915-10

DENV3/5532 displays higher infectivity and increased mdDC apoptosis compared to DENV3/290. (A) Percentages of infected mdDCs exposed to DENV3/290 or DENV3/5532 or mock infected. (B) Viral progeny in mdDC culture supernatants. For viral progeny, results are expressed in log 10 focus-forming units (ffu) in C6/36 cells/ml. (C) Percentages of apoptotic cells. mdDCs were infected as described for panel A and assessed for annexin V-positive events by flow cytometry. Data were analyzed using two-way ANOVA followed by a Bonferroni test; values represent means ± SD of the results of four different experiments. (D) Percentages of mdDCs costained for CD11c + fluorescein isothiocyanate (FITC) and 4G2 + anti-flavivirus antibody plus anti-mouse PE conjugated by flow cytometry after 72 hpi. Data were analyzed using one-way ANOVA followed by a Bonferroni test; values represent means ± SD of the results of three different experiments. (E) Dot plot analyzes of one representative mdDC culture costained for CD11c + and 4G2 + . *, P
Figure Legend Snippet: DENV3/5532 displays higher infectivity and increased mdDC apoptosis compared to DENV3/290. (A) Percentages of infected mdDCs exposed to DENV3/290 or DENV3/5532 or mock infected. (B) Viral progeny in mdDC culture supernatants. For viral progeny, results are expressed in log 10 focus-forming units (ffu) in C6/36 cells/ml. (C) Percentages of apoptotic cells. mdDCs were infected as described for panel A and assessed for annexin V-positive events by flow cytometry. Data were analyzed using two-way ANOVA followed by a Bonferroni test; values represent means ± SD of the results of four different experiments. (D) Percentages of mdDCs costained for CD11c + fluorescein isothiocyanate (FITC) and 4G2 + anti-flavivirus antibody plus anti-mouse PE conjugated by flow cytometry after 72 hpi. Data were analyzed using one-way ANOVA followed by a Bonferroni test; values represent means ± SD of the results of three different experiments. (E) Dot plot analyzes of one representative mdDC culture costained for CD11c + and 4G2 + . *, P

Techniques Used: Infection, Flow Cytometry, Cytometry

36) Product Images from "Enhanced anti-colorectal cancer effects of carfilzomib combined with CPT-11 via downregulation of nuclear factor-κB in vitro and in vivo"

Article Title: Enhanced anti-colorectal cancer effects of carfilzomib combined with CPT-11 via downregulation of nuclear factor-κB in vitro and in vivo

Journal: International Journal of Oncology

doi: 10.3892/ijo.2014.2513

CFZ ± CPT-11 induces apoptosis by caspase 3 and CD95 upregulation, as well as p-p38 and ATF3 activation in SW620 cells. (A) The effect of CFZ ± CPT-11 on the expression of Annexin V/PI in SW620 cells. SW620 cells were treated with 20 and 50 nM CFZ alone or in combination with 50 and 100 μM CPT-11 for 48 h. The results show the percentages of apoptosis (Annexin V + /PI − ) and necrosis (Annexin V + /PI + ). (B) SW620 cells were treated with the indicated concentration of CFZ ± CPT-11 for 48 h, and cell death was determined by Annexin V/PI (part of the data from Fig. 5A; bars, SD, n=3). * P
Figure Legend Snippet: CFZ ± CPT-11 induces apoptosis by caspase 3 and CD95 upregulation, as well as p-p38 and ATF3 activation in SW620 cells. (A) The effect of CFZ ± CPT-11 on the expression of Annexin V/PI in SW620 cells. SW620 cells were treated with 20 and 50 nM CFZ alone or in combination with 50 and 100 μM CPT-11 for 48 h. The results show the percentages of apoptosis (Annexin V + /PI − ) and necrosis (Annexin V + /PI + ). (B) SW620 cells were treated with the indicated concentration of CFZ ± CPT-11 for 48 h, and cell death was determined by Annexin V/PI (part of the data from Fig. 5A; bars, SD, n=3). * P

Techniques Used: Cycling Probe Technology, Activation Assay, Expressing, Concentration Assay

37) Product Images from "Acetylsalicylic acid rescues the immunomodulation of inflamed gingiva-derived mesenchymal stem cells via upregulating FasL in mice"

Article Title: Acetylsalicylic acid rescues the immunomodulation of inflamed gingiva-derived mesenchymal stem cells via upregulating FasL in mice

Journal: Stem Cell Research & Therapy

doi: 10.1186/s13287-019-1485-5

The immunomodulatory properties of iGMSCs are impaired. a In vitro co-culture system showed a significantly decreased capacity of iGMSCs to induce Annexin V + /7AAD + T cell apoptosis when compared with the GMSC group. b Schema showing iGMSC and GMSC transplantation for treating dextran sodium sulfate (DSS)-induced experimental colitis mice. c – e iGMSCs showed impaired immunomodulation capacity compared with GMSCs, as assessed by c amelioration of losing body weight, d decreased disease activity index (DAI), and e alleviation of colitis histologic activity index (HAI). f – h FACS analysis showed that the levels of Th1 and Th17 were significantly elevated while the levels of Tregs were significantly reduced in colitis mice compared with the control mice. iGMSC infusion exhibited compromised reduction of Th1 and Th17 cells and upregulation of Treg levels in colitis in mice than did GMSCs. i ELISA analysis showed that the levels of tumor necrosis factor α (TNF-α) and IL-17 in serum were markedly increased in colitis mice compared with the control mice at 10 days post-DSS induction. iGMSC infusion exhibited compromised ability to downregulate serum levels of TNF-α and IL-17 compared with GMSCs. n = 5 in each group. Scale bar = 200 μm. * P
Figure Legend Snippet: The immunomodulatory properties of iGMSCs are impaired. a In vitro co-culture system showed a significantly decreased capacity of iGMSCs to induce Annexin V + /7AAD + T cell apoptosis when compared with the GMSC group. b Schema showing iGMSC and GMSC transplantation for treating dextran sodium sulfate (DSS)-induced experimental colitis mice. c – e iGMSCs showed impaired immunomodulation capacity compared with GMSCs, as assessed by c amelioration of losing body weight, d decreased disease activity index (DAI), and e alleviation of colitis histologic activity index (HAI). f – h FACS analysis showed that the levels of Th1 and Th17 were significantly elevated while the levels of Tregs were significantly reduced in colitis mice compared with the control mice. iGMSC infusion exhibited compromised reduction of Th1 and Th17 cells and upregulation of Treg levels in colitis in mice than did GMSCs. i ELISA analysis showed that the levels of tumor necrosis factor α (TNF-α) and IL-17 in serum were markedly increased in colitis mice compared with the control mice at 10 days post-DSS induction. iGMSC infusion exhibited compromised ability to downregulate serum levels of TNF-α and IL-17 compared with GMSCs. n = 5 in each group. Scale bar = 200 μm. * P

Techniques Used: In Vitro, Co-Culture Assay, Transplantation Assay, Mouse Assay, Activity Assay, FACS, Enzyme-linked Immunosorbent Assay

ASA treatment rescues the impaired immunomodulation capacity of iGMSCs via upregulation of FasL. a Western blot analysis showed that FasL siRNA downregulated the expression level of FasL in ASA-iGMSCs. b , c In vitro co-culture system showed that FasL siRNA-treated ASA-iGMSCs was not able to sufficiently induce Annexin V + /7AAD + T cell apoptosis compared with untreated ASA-iGMSCs. d – f FasL siRNA-treated ASA-iGMSCs showed decreased immunomodulation capacity compared with untreated ASA-iGMSCs, as assessed by d amelioration of losing body weight, e decreased disease activity index (DAI), and f alleviation of colitis histologic activity index (HAI). g FasL siRNA treatment inhibits ASA-iGMSCs’ ability to reduce the levels of Th1 and Th17 cells and elevate the levels of Treg cells in colitis mice. h FasL siRNA treatment inhibits ASA-iGMSCs’ ability to downregulate the serum levels of TNF-α and IL-17 in colitis mice. n = 5 in each group. Scale bar = 200 μm. * P
Figure Legend Snippet: ASA treatment rescues the impaired immunomodulation capacity of iGMSCs via upregulation of FasL. a Western blot analysis showed that FasL siRNA downregulated the expression level of FasL in ASA-iGMSCs. b , c In vitro co-culture system showed that FasL siRNA-treated ASA-iGMSCs was not able to sufficiently induce Annexin V + /7AAD + T cell apoptosis compared with untreated ASA-iGMSCs. d – f FasL siRNA-treated ASA-iGMSCs showed decreased immunomodulation capacity compared with untreated ASA-iGMSCs, as assessed by d amelioration of losing body weight, e decreased disease activity index (DAI), and f alleviation of colitis histologic activity index (HAI). g FasL siRNA treatment inhibits ASA-iGMSCs’ ability to reduce the levels of Th1 and Th17 cells and elevate the levels of Treg cells in colitis mice. h FasL siRNA treatment inhibits ASA-iGMSCs’ ability to downregulate the serum levels of TNF-α and IL-17 in colitis mice. n = 5 in each group. Scale bar = 200 μm. * P

Techniques Used: Western Blot, Expressing, In Vitro, Co-Culture Assay, Activity Assay, Mouse Assay

ASA treatment rescues the impaired immunomodulatory properties of iGMSCs. a In vitro co-culture system showed a significantly increased capacity of ASA-treated iGMSCs to induce Annexin V + /7AAD + T cell apoptosis when compared with the iGMSC group. b Western blot analysis showed that ASA treatment elevated the expression of FasL in iGMSCs. c – e ASA-treated iGMSCs showed restored immunomodulation capacity compared with untreated iGMSCs, as assessed by c amelioration of losing body weight, d decreased disease activity index (DAI), and e alleviation of colitis histologic activity index (HAI). f FACS analysis showed that ASA-treated iGMSC infusion rescued the iGMSCs’ ability to reduce Th1 and Th17 cells and elevate Treg cells in colitis mice. g ELISA analysis showed that ASA-treated iGMSC infusion rescued the iGMSCs’ ability to downregulate the levels of TNF-α and IL-17 in colitis mice. n = 5 in each group. Scale bar = 200 μm. * P
Figure Legend Snippet: ASA treatment rescues the impaired immunomodulatory properties of iGMSCs. a In vitro co-culture system showed a significantly increased capacity of ASA-treated iGMSCs to induce Annexin V + /7AAD + T cell apoptosis when compared with the iGMSC group. b Western blot analysis showed that ASA treatment elevated the expression of FasL in iGMSCs. c – e ASA-treated iGMSCs showed restored immunomodulation capacity compared with untreated iGMSCs, as assessed by c amelioration of losing body weight, d decreased disease activity index (DAI), and e alleviation of colitis histologic activity index (HAI). f FACS analysis showed that ASA-treated iGMSC infusion rescued the iGMSCs’ ability to reduce Th1 and Th17 cells and elevate Treg cells in colitis mice. g ELISA analysis showed that ASA-treated iGMSC infusion rescued the iGMSCs’ ability to downregulate the levels of TNF-α and IL-17 in colitis mice. n = 5 in each group. Scale bar = 200 μm. * P

Techniques Used: In Vitro, Co-Culture Assay, Western Blot, Expressing, Activity Assay, FACS, Mouse Assay, Enzyme-linked Immunosorbent Assay

38) Product Images from "Obesity Impairs γδ T Cell Homeostasis and Antiviral Function in Humans"

Article Title: Obesity Impairs γδ T Cell Homeostasis and Antiviral Function in Humans

Journal: PLoS ONE

doi: 10.1371/journal.pone.0120918

Vγ9Vδ2 T cells are more sensitive to apoptosis and have a reduced ability to produce IFN-γ in response to nonobese APCs. PBMC from non-obese (NOD) or obese (OD) donors were co-cultured with a monocytic cell line U937 to determine if the dysfunction was T cell intrinsic. (A, B) Representative plots of IFN-γ production by Vδ2 + CD3 + gated cells. PBMC were either cultured alone (uninfected), with U937 cells (U937 ctrl) or with influenza infected U937 cells (U937 infected). (C) Percentage of Vδ2 + T cells producing IFN-γ in NOD and OD in response to infected U937 cells (corrected for controls, n = 8). Horizontal lines represent the mean. (D) Change in MFI of CD69 expression on Vδ2 + CD3 + gated cells following 8 hour incubation with uninfected or infected U937 cells. Data represents five separate donor pairs and their standard deviation. P-values calculated from Pearson’s unpaired Student’s t -test. (E) Data represents the percentage of Vδ2 + T cells staining positive for Annexin-V but not PI (dark bars) or positive for both Annexin-V and PI (white bars) by flow cytometry (n = 5) with standard deviation.
Figure Legend Snippet: Vγ9Vδ2 T cells are more sensitive to apoptosis and have a reduced ability to produce IFN-γ in response to nonobese APCs. PBMC from non-obese (NOD) or obese (OD) donors were co-cultured with a monocytic cell line U937 to determine if the dysfunction was T cell intrinsic. (A, B) Representative plots of IFN-γ production by Vδ2 + CD3 + gated cells. PBMC were either cultured alone (uninfected), with U937 cells (U937 ctrl) or with influenza infected U937 cells (U937 infected). (C) Percentage of Vδ2 + T cells producing IFN-γ in NOD and OD in response to infected U937 cells (corrected for controls, n = 8). Horizontal lines represent the mean. (D) Change in MFI of CD69 expression on Vδ2 + CD3 + gated cells following 8 hour incubation with uninfected or infected U937 cells. Data represents five separate donor pairs and their standard deviation. P-values calculated from Pearson’s unpaired Student’s t -test. (E) Data represents the percentage of Vδ2 + T cells staining positive for Annexin-V but not PI (dark bars) or positive for both Annexin-V and PI (white bars) by flow cytometry (n = 5) with standard deviation.

Techniques Used: Cell Culture, Infection, Expressing, Incubation, Standard Deviation, Staining, Flow Cytometry, Cytometry

39) Product Images from "Impact of Salinomycin on human cholangiocarcinoma: induction of apoptosis and impairment of tumor cell proliferation in vitro"

Article Title: Impact of Salinomycin on human cholangiocarcinoma: induction of apoptosis and impairment of tumor cell proliferation in vitro

Journal: BMC Cancer

doi: 10.1186/1471-2407-12-466

Salinomycin induces apoptosis in human CC cells. 1 x 10 6 Mz-ChA-1, TFK-1 and EGI-1 cells were cultured until confluence, followed by subsequent incubation of cells in the absence or presence of 1 mM Gemcitabine, 1 μM, 2 μM, 5 μM or 10 μM Salinomycin for 24 hours. Cells were trypsinized and stained with Annexin V-FITC and analyzed by flow cytometry. ( A ) As expected, treatment with Gemcitabine (dotted line in overlay) led to very weak increase of apoptotic cells in all tested cell lines in comparison to untreated cells (solid bright line). In contrast, Salinomycin induced strongly augmented number of apoptotic cells for Mz-ChA-1 and TFK-1, whereas EGI-1 cells revealed a pronounced resistance towards Salinomycin-induced apoptosis (solid dark line). Results are shown as representative scatter-grams of Annexin V + cells or summarizing 3 independent experiments as mean ± SD ( B ).
Figure Legend Snippet: Salinomycin induces apoptosis in human CC cells. 1 x 10 6 Mz-ChA-1, TFK-1 and EGI-1 cells were cultured until confluence, followed by subsequent incubation of cells in the absence or presence of 1 mM Gemcitabine, 1 μM, 2 μM, 5 μM or 10 μM Salinomycin for 24 hours. Cells were trypsinized and stained with Annexin V-FITC and analyzed by flow cytometry. ( A ) As expected, treatment with Gemcitabine (dotted line in overlay) led to very weak increase of apoptotic cells in all tested cell lines in comparison to untreated cells (solid bright line). In contrast, Salinomycin induced strongly augmented number of apoptotic cells for Mz-ChA-1 and TFK-1, whereas EGI-1 cells revealed a pronounced resistance towards Salinomycin-induced apoptosis (solid dark line). Results are shown as representative scatter-grams of Annexin V + cells or summarizing 3 independent experiments as mean ± SD ( B ).

Techniques Used: Cell Culture, Incubation, Staining, Flow Cytometry, Cytometry

40) Product Images from "Ruxolitinib synergistically enhances the anti-tumor activity of paclitaxel in human ovarian cancer"

Article Title: Ruxolitinib synergistically enhances the anti-tumor activity of paclitaxel in human ovarian cancer

Journal: Oncotarget

doi: 10.18632/oncotarget.24368

Dose dependent induction of apoptosis (A) and (B) OVCAR-8 and MDAH 2774 cells were incubated with various concentrations of ruxolitinib for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A) or using cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (B). * P
Figure Legend Snippet: Dose dependent induction of apoptosis (A) and (B) OVCAR-8 and MDAH 2774 cells were incubated with various concentrations of ruxolitinib for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A) or using cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (B). * P

Techniques Used: Incubation, Flow Cytometry, Cytometry, Staining, Western Blot

Ruxolitinib enhanced paclitaxel-induced apoptosis in human ovarian cancer cells (A) OVCAR-8 and (B) MDAH2774 cells were treated with ruxolitinib (30μM), paclitaxel (10nM) either alone or together, for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A B) or by cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (C) . ns: not significant; # P
Figure Legend Snippet: Ruxolitinib enhanced paclitaxel-induced apoptosis in human ovarian cancer cells (A) OVCAR-8 and (B) MDAH2774 cells were treated with ruxolitinib (30μM), paclitaxel (10nM) either alone or together, for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining (A B) or by cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (C) . ns: not significant; # P

Techniques Used: Flow Cytometry, Cytometry, Staining, Western Blot

41) Product Images from "Structure of human immunoproteasome with a reversible and noncompetitive inhibitor that selectively inhibits activated lymphocytes"

Article Title: Structure of human immunoproteasome with a reversible and noncompetitive inhibitor that selectively inhibits activated lymphocytes

Journal: Nature Communications

doi: 10.1038/s41467-017-01760-5

PKS21221 induced apoptosis and cell death in differentiated antibody-secreting cells (ASCs) and CD19 + B cells. PBMCs were cultured with or without IL-2/R848 for 5 days, followed by 12-h incubation with PKS21221 at different concentrations. a Representative flow cytometry plots of cells treated with IL-2 and R848 in the presence and absence of PKS21221. IL-2 and R848 differentiated B cells into CD27 + CD38 + antibody-secreting cells (ASCs). PKS21221 reduced the percentage of ASCs. b Viability of total PBMCs using 7-aminoactinomycin D (7-AAD). 12-h treatment with PKS21221 did not affect overall viability of PBMCs. c A representative plot of apoptosis and viability assay using Annexin V and 7-AAD. Annexin V + 7-AAD − cells were referred as “Early apoptosis” population, and Annexin V + 7-AAD + cells were referred as “Dead” population. d Percentage of early apoptotic ASCs (left) and dead ASCs (right), after 12-h incubation with PKS21221. PKS21221 treatment induced apoptotic cell death in a dose-dependent manner. e Early apoptotic (left) and dead (right) populations in CD19 − non-B cells and CD19 + non-ASCs. Experiments were repeated on PBMCs from 5 donors in 5 separate experiments, each data points in b and e , were the mean + SEM of three technical replicates. * p
Figure Legend Snippet: PKS21221 induced apoptosis and cell death in differentiated antibody-secreting cells (ASCs) and CD19 + B cells. PBMCs were cultured with or without IL-2/R848 for 5 days, followed by 12-h incubation with PKS21221 at different concentrations. a Representative flow cytometry plots of cells treated with IL-2 and R848 in the presence and absence of PKS21221. IL-2 and R848 differentiated B cells into CD27 + CD38 + antibody-secreting cells (ASCs). PKS21221 reduced the percentage of ASCs. b Viability of total PBMCs using 7-aminoactinomycin D (7-AAD). 12-h treatment with PKS21221 did not affect overall viability of PBMCs. c A representative plot of apoptosis and viability assay using Annexin V and 7-AAD. Annexin V + 7-AAD − cells were referred as “Early apoptosis” population, and Annexin V + 7-AAD + cells were referred as “Dead” population. d Percentage of early apoptotic ASCs (left) and dead ASCs (right), after 12-h incubation with PKS21221. PKS21221 treatment induced apoptotic cell death in a dose-dependent manner. e Early apoptotic (left) and dead (right) populations in CD19 − non-B cells and CD19 + non-ASCs. Experiments were repeated on PBMCs from 5 donors in 5 separate experiments, each data points in b and e , were the mean + SEM of three technical replicates. * p

Techniques Used: Cell Culture, Incubation, Flow Cytometry, Cytometry, Viability Assay

42) Product Images from "Structure of human immunoproteasome with a reversible and noncompetitive inhibitor that selectively inhibits activated lymphocytes"

Article Title: Structure of human immunoproteasome with a reversible and noncompetitive inhibitor that selectively inhibits activated lymphocytes

Journal: Nature Communications

doi: 10.1038/s41467-017-01760-5

PKS21221 induced apoptosis and cell death in differentiated antibody-secreting cells (ASCs) and CD19 + B cells. PBMCs were cultured with or without IL-2/R848 for 5 days, followed by 12-h incubation with PKS21221 at different concentrations. a Representative flow cytometry plots of cells treated with IL-2 and R848 in the presence and absence of PKS21221. IL-2 and R848 differentiated B cells into CD27 + CD38 + antibody-secreting cells (ASCs). PKS21221 reduced the percentage of ASCs. b Viability of total PBMCs using 7-aminoactinomycin D (7-AAD). 12-h treatment with PKS21221 did not affect overall viability of PBMCs. c A representative plot of apoptosis and viability assay using Annexin V and 7-AAD. Annexin V + 7-AAD − cells were referred as “Early apoptosis” population, and Annexin V + 7-AAD + cells were referred as “Dead” population. d Percentage of early apoptotic ASCs (left) and dead ASCs (right), after 12-h incubation with PKS21221. PKS21221 treatment induced apoptotic cell death in a dose-dependent manner. e Early apoptotic (left) and dead (right) populations in CD19 − non-B cells and CD19 + non-ASCs. Experiments were repeated on PBMCs from 5 donors in 5 separate experiments, each data points in b and e , were the mean + SEM of three technical replicates. * p
Figure Legend Snippet: PKS21221 induced apoptosis and cell death in differentiated antibody-secreting cells (ASCs) and CD19 + B cells. PBMCs were cultured with or without IL-2/R848 for 5 days, followed by 12-h incubation with PKS21221 at different concentrations. a Representative flow cytometry plots of cells treated with IL-2 and R848 in the presence and absence of PKS21221. IL-2 and R848 differentiated B cells into CD27 + CD38 + antibody-secreting cells (ASCs). PKS21221 reduced the percentage of ASCs. b Viability of total PBMCs using 7-aminoactinomycin D (7-AAD). 12-h treatment with PKS21221 did not affect overall viability of PBMCs. c A representative plot of apoptosis and viability assay using Annexin V and 7-AAD. Annexin V + 7-AAD − cells were referred as “Early apoptosis” population, and Annexin V + 7-AAD + cells were referred as “Dead” population. d Percentage of early apoptotic ASCs (left) and dead ASCs (right), after 12-h incubation with PKS21221. PKS21221 treatment induced apoptotic cell death in a dose-dependent manner. e Early apoptotic (left) and dead (right) populations in CD19 − non-B cells and CD19 + non-ASCs. Experiments were repeated on PBMCs from 5 donors in 5 separate experiments, each data points in b and e , were the mean + SEM of three technical replicates. * p

Techniques Used: Cell Culture, Incubation, Flow Cytometry, Cytometry, Viability Assay

43) Product Images from "Transcriptome Kinetics of Circulating Neutrophils during Human Experimental Endotoxemia"

Article Title: Transcriptome Kinetics of Circulating Neutrophils during Human Experimental Endotoxemia

Journal: PLoS ONE

doi: 10.1371/journal.pone.0038255

Ex vivo neutrophil stimulation. A Gene expression in in vitro stimulated neutrophils. Cells were stimulated with 10 ng LPS, 10 ng rTNFα, 50 ng rG-CSF or 50 ng rGM-CSF. At t = 2 h after stimulation RNA was isolated and q-pcr was performed with taqman probes for specific genes. Fold change relative to unstimulated. Error bars represent SEM (N = 4). B Survival after stimulation with 10 ng LPS, 10 ng rTNFα, 50 ng rG-CSF or 50 ng rGM-CSF. Cell viability was determined in ≥1×10 5 cells with Annexin V apoptosis detection kit at 7 hours after stimulation. Dots represent the percentage of viable (Annexin V negative and 7 AAD negative) cells. N = 4 *p
Figure Legend Snippet: Ex vivo neutrophil stimulation. A Gene expression in in vitro stimulated neutrophils. Cells were stimulated with 10 ng LPS, 10 ng rTNFα, 50 ng rG-CSF or 50 ng rGM-CSF. At t = 2 h after stimulation RNA was isolated and q-pcr was performed with taqman probes for specific genes. Fold change relative to unstimulated. Error bars represent SEM (N = 4). B Survival after stimulation with 10 ng LPS, 10 ng rTNFα, 50 ng rG-CSF or 50 ng rGM-CSF. Cell viability was determined in ≥1×10 5 cells with Annexin V apoptosis detection kit at 7 hours after stimulation. Dots represent the percentage of viable (Annexin V negative and 7 AAD negative) cells. N = 4 *p

Techniques Used: Ex Vivo, Expressing, In Vitro, Isolation, Polymerase Chain Reaction

44) Product Images from "Knockdown delta-5-desaturase promotes the formation of a novel free radical byproduct from COX-catalyzed ω-6 peroxidation to induce apoptosis and sensitize pancreatic cancer cells to chemotherapy drugs"

Article Title: Knockdown delta-5-desaturase promotes the formation of a novel free radical byproduct from COX-catalyzed ω-6 peroxidation to induce apoptosis and sensitize pancreatic cancer cells to chemotherapy drugs

Journal: Free radical biology & medicine

doi: 10.1016/j.freeradbiomed.2016.06.028

8-HOA’s growth inhibitory effects on BxPC-3 cells. (A) Clonogenic assay of BxPC-3 cells at 10 days after 48 h of 8-HOA treatment (1.0 μM), gemcitabine (0.1 μM), and 8-HOA+gemcitabine. The BxPC-3 cells treated with vehicle only were used as control; (B) Cell apoptosis analysis of BxPC-3 cells. After 48 h treatment of vehicle (control), 8-HOA (1.0 μM), gemcitabine (0.1 μM) and gemcitabine +8-HOA, cells were double stained with FITC-Annexin V/PI and subjected to flow cytometry. (*: significant difference vs. control with p
Figure Legend Snippet: 8-HOA’s growth inhibitory effects on BxPC-3 cells. (A) Clonogenic assay of BxPC-3 cells at 10 days after 48 h of 8-HOA treatment (1.0 μM), gemcitabine (0.1 μM), and 8-HOA+gemcitabine. The BxPC-3 cells treated with vehicle only were used as control; (B) Cell apoptosis analysis of BxPC-3 cells. After 48 h treatment of vehicle (control), 8-HOA (1.0 μM), gemcitabine (0.1 μM) and gemcitabine +8-HOA, cells were double stained with FITC-Annexin V/PI and subjected to flow cytometry. (*: significant difference vs. control with p

Techniques Used: Clonogenic Assay, Staining, Flow Cytometry, Cytometry

45) Product Images from "Pterostilbene Suppresses Ovarian Cancer Growth via Induction of Apoptosis and Blockade of Cell Cycle Progression Involving Inhibition of the STAT3 Pathway"

Article Title: Pterostilbene Suppresses Ovarian Cancer Growth via Induction of Apoptosis and Blockade of Cell Cycle Progression Involving Inhibition of the STAT3 Pathway

Journal: International Journal of Molecular Sciences

doi: 10.3390/ijms19071983

Pterostilbene induces cell apoptosis. OVCAR-8 and Caov-3 cells were treated with vehicle and PTE (25–300 μm) for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining ( A , B ) or by Western blot for the expression of cleaved poly-ADP ribose polymerase (PARP) ( C ). Results are representative of 3 or more preparations. *, p
Figure Legend Snippet: Pterostilbene induces cell apoptosis. OVCAR-8 and Caov-3 cells were treated with vehicle and PTE (25–300 μm) for 48 h. Apoptosis was determined by flow cytometry using annexin V and PI staining ( A , B ) or by Western blot for the expression of cleaved poly-ADP ribose polymerase (PARP) ( C ). Results are representative of 3 or more preparations. *, p

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

46) Product Images from "Peroxisome proliferator-activated receptor-γ enhances human pulmonary artery smooth muscle cell apoptosis through microRNA-21 and programmed cell death 4"

Article Title: Peroxisome proliferator-activated receptor-γ enhances human pulmonary artery smooth muscle cell apoptosis through microRNA-21 and programmed cell death 4

Journal: American Journal of Physiology - Lung Cellular and Molecular Physiology

doi: 10.1152/ajplung.00532.2016

PPARγ modulates PDCD4 protein, HPASMC proliferation, and apoptosis in response to hypoxia. HPASMC monolayers were exposed to normoxic (21% O 2 ) or hypoxic (1% O 2 ) conditions for 72 h. At the onset of the study, selected HPASMCs were transfected with AdGFP (10 MOI) or AdPPARγ (10 MOI) for 6 h. RSG (10 μM) was added to the culture medium after 24 h of incubation. Upon the conclusion of the study period, total protein was extracted from HPASMC monolayer lysates, and proliferation and apoptosis assays were conducted. A : demonstration of PDCD4 protein expression in HPASMCs with representative immunoblot. B : proliferation was detected in HPASMCs using automated cell counting. C : demonstration of caspase-3 activation as an indicator of apoptosis in normoxia- or hypoxia-exposed HPASMCs treated with AdPPARγ or AdGFP + RSG. D : HPASMCs exposed to normoxia or hypoxia for 72 h were treated with RSG (10–25 μM) or an equal volume of DMSO at the onset of the study period. With the use of FACS analysis, apoptosis was examined by detecting annexin V staining in HPASMCs. Staurosporine (0.1 μM) was applied to selected normoxia-exposed HPASMCs as a positive control to enhance apoptosis. Bars represent means ± SE for PDCD4 protein ( A ), proliferation ( B ), caspase-3 activation ( C ), or annexin V ( D ) detection by median fluorescence. A : n = 12; **** P
Figure Legend Snippet: PPARγ modulates PDCD4 protein, HPASMC proliferation, and apoptosis in response to hypoxia. HPASMC monolayers were exposed to normoxic (21% O 2 ) or hypoxic (1% O 2 ) conditions for 72 h. At the onset of the study, selected HPASMCs were transfected with AdGFP (10 MOI) or AdPPARγ (10 MOI) for 6 h. RSG (10 μM) was added to the culture medium after 24 h of incubation. Upon the conclusion of the study period, total protein was extracted from HPASMC monolayer lysates, and proliferation and apoptosis assays were conducted. A : demonstration of PDCD4 protein expression in HPASMCs with representative immunoblot. B : proliferation was detected in HPASMCs using automated cell counting. C : demonstration of caspase-3 activation as an indicator of apoptosis in normoxia- or hypoxia-exposed HPASMCs treated with AdPPARγ or AdGFP + RSG. D : HPASMCs exposed to normoxia or hypoxia for 72 h were treated with RSG (10–25 μM) or an equal volume of DMSO at the onset of the study period. With the use of FACS analysis, apoptosis was examined by detecting annexin V staining in HPASMCs. Staurosporine (0.1 μM) was applied to selected normoxia-exposed HPASMCs as a positive control to enhance apoptosis. Bars represent means ± SE for PDCD4 protein ( A ), proliferation ( B ), caspase-3 activation ( C ), or annexin V ( D ) detection by median fluorescence. A : n = 12; **** P

Techniques Used: Transfection, Incubation, Expressing, Cell Counting, Activation Assay, FACS, Staining, Positive Control, Fluorescence

47) Product Images from "Increased Expression of the Pro-apoptotic Protein BIM, a Mechanism for cAMP/Protein Kinase A (PKA)-induced Apoptosis of Immature T Cells *"

Article Title: Increased Expression of the Pro-apoptotic Protein BIM, a Mechanism for cAMP/Protein Kinase A (PKA)-induced Apoptosis of Immature T Cells *

Journal: The Journal of Biological Chemistry

doi: 10.1074/jbc.M111.268979

Bim is required for CPT-cAMP-mediated apoptosis of CD4+/CD8+ thymocytes. T cells were isolated from Bim +/+ (WT) and Bim −/− mice, incubated with CPT-cAMP, and then assessed for apoptosis, as determined by the expression of annexin V. A
Figure Legend Snippet: Bim is required for CPT-cAMP-mediated apoptosis of CD4+/CD8+ thymocytes. T cells were isolated from Bim +/+ (WT) and Bim −/− mice, incubated with CPT-cAMP, and then assessed for apoptosis, as determined by the expression of annexin V. A

Techniques Used: Cycling Probe Technology, Isolation, Mouse Assay, Incubation, Expressing

48) Product Images from "SNAI1-Mediated Epithelial-Mesenchymal Transition Confers Chemoresistance and Cellular Plasticity by Regulating Genes Involved in Cell Death and Stem Cell Maintenance"

Article Title: SNAI1-Mediated Epithelial-Mesenchymal Transition Confers Chemoresistance and Cellular Plasticity by Regulating Genes Involved in Cell Death and Stem Cell Maintenance

Journal: PLoS ONE

doi: 10.1371/journal.pone.0066558

EMT generates cells with increased resistance to cell death. (A) MCF10A cells stably expressing SNAI1 (MCF10A-SNAI1) induced an EMT as shown by fibroblast-like appearance. Bar = 10 µm. Western blot analysis showed down-regulation of the epithelial marker, E-cadherin (CDH1) in SNAI1-overexpressing cells. (B) MCF10A-SNAI1 cells were more resistant to camptothecin (CPT) - and doxorubicin-induced cell death, as determined by MTT analysis. (C) MCF10A-SNAI1 cells were more resistant to the CPT-induced apoptosis as revealed by Hoechst staining and immunofluorescence using anti-active caspase-3 antibody. Positive cells for Hoechst or active caspase-3 staining were counted in the table. Bar = 100 µm. (D) Using Annexin V apoptosis detection kit, apoptotic cells were quantified upon CPT treatment (24 h). Viable cells are Annexin V-PE and 7-AAD-PerCP negative, and cells that are in early apoptosis are Annexin V-PE positive and 7-AAD-PerCP negative. Cells that are in late apoptosis or already dead are both Annexin V-PE and 7-AAD-PerCP positive. (E) To investigate the cellular response to CPT-induced DNA damage, western blot analysis was performed using antibody against γ-H2A.X, which is an indicator for DNA repair response. No difference in the accumulation of γ-H2A.X was observed in two cell lines for the indicated times, whereas MCF10A-SNAI1 cells showed higher AKT activation 6, 18 and 24 h after CPT treatment. (F) Serum-depletion for 10–15 days induced more cell death in MCF10A cells than MCF10A-SNAI1 cells as determined by a trypan blue assay. * P
Figure Legend Snippet: EMT generates cells with increased resistance to cell death. (A) MCF10A cells stably expressing SNAI1 (MCF10A-SNAI1) induced an EMT as shown by fibroblast-like appearance. Bar = 10 µm. Western blot analysis showed down-regulation of the epithelial marker, E-cadherin (CDH1) in SNAI1-overexpressing cells. (B) MCF10A-SNAI1 cells were more resistant to camptothecin (CPT) - and doxorubicin-induced cell death, as determined by MTT analysis. (C) MCF10A-SNAI1 cells were more resistant to the CPT-induced apoptosis as revealed by Hoechst staining and immunofluorescence using anti-active caspase-3 antibody. Positive cells for Hoechst or active caspase-3 staining were counted in the table. Bar = 100 µm. (D) Using Annexin V apoptosis detection kit, apoptotic cells were quantified upon CPT treatment (24 h). Viable cells are Annexin V-PE and 7-AAD-PerCP negative, and cells that are in early apoptosis are Annexin V-PE positive and 7-AAD-PerCP negative. Cells that are in late apoptosis or already dead are both Annexin V-PE and 7-AAD-PerCP positive. (E) To investigate the cellular response to CPT-induced DNA damage, western blot analysis was performed using antibody against γ-H2A.X, which is an indicator for DNA repair response. No difference in the accumulation of γ-H2A.X was observed in two cell lines for the indicated times, whereas MCF10A-SNAI1 cells showed higher AKT activation 6, 18 and 24 h after CPT treatment. (F) Serum-depletion for 10–15 days induced more cell death in MCF10A cells than MCF10A-SNAI1 cells as determined by a trypan blue assay. * P

Techniques Used: Stable Transfection, Expressing, Western Blot, Marker, Cycling Probe Technology, MTT Assay, Staining, Immunofluorescence, Activation Assay, Serum Depletion

49) Product Images from "Synergistic anti-tumor effect of combined inhibition of EGFR and JAK/STAT3 pathways in human ovarian cancer"

Article Title: Synergistic anti-tumor effect of combined inhibition of EGFR and JAK/STAT3 pathways in human ovarian cancer

Journal: Molecular Cancer

doi: 10.1186/s12943-015-0366-5

Suppressing JAK/STAT3 signaling enhanced gefitinib-induced apoptosis in human ovarian cancer cells. SKOV3 (A B) and MDAH2774 (C D) cells were treated with gefitinib, JAKi, either alone or together, for 48 h. Apoptosis was determined by flow cytometry using Annexin V and PI staining (A C) or for cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (B D) . **, P
Figure Legend Snippet: Suppressing JAK/STAT3 signaling enhanced gefitinib-induced apoptosis in human ovarian cancer cells. SKOV3 (A B) and MDAH2774 (C D) cells were treated with gefitinib, JAKi, either alone or together, for 48 h. Apoptosis was determined by flow cytometry using Annexin V and PI staining (A C) or for cleaved poly-ADP ribose polymerase (PARP) and cleaved caspase-3 by Western blot (B D) . **, P

Techniques Used: Flow Cytometry, Cytometry, Staining, Western Blot

50) Product Images from "Iguratimod Inhibits the Aggressiveness of Rheumatoid Fibroblast-Like Synoviocytes"

Article Title: Iguratimod Inhibits the Aggressiveness of Rheumatoid Fibroblast-Like Synoviocytes

Journal: Journal of Immunology Research

doi: 10.1155/2019/6929286

Iguratimod promoted apoptosis of RA-FLSs. (a) RA-FLSs were treated with TNF- α (25 ng/ml) and different concentrations of iguratimod for 24 h. Propidium iodide (PI) and Annexin V (AV) staining were determined by flow cytometry. Typical flow plots were shown. (b, c) The summary data of early apoptotic cells (PI − AV + ) and late apoptotic or dead cells (PI + AV + ) were shown. The data were described as the mean ± SD for three independent experiments (totally 8 RA-FLSs lines). (d, e) Caspase 3 and cFLIP mRNA expression were measured by qPCR in RA-FLSs. (f) For inhibitor experiments, cells were pretreated with the pan-caspase inhibitor z-VAD-fmk (Sigma-Aldrich, 10 μ M) or N-acetyl-l-cysteine (NAC, Sigma-Aldrich, 5 mM) for 1 h before the treatment of iguratimod. RA-FLSs were treated with TNF- α (25 ng/ml) and iguratimod (0.5 μ g/ml) for 24 h. Apoptosis was determined by flow cytometry. Typical flow plots were shown. The data were shown as the mean ± SD for three independent experiments. The data were analyzed using one-way ANOVA followed by Turkey's test. ∗ P
Figure Legend Snippet: Iguratimod promoted apoptosis of RA-FLSs. (a) RA-FLSs were treated with TNF- α (25 ng/ml) and different concentrations of iguratimod for 24 h. Propidium iodide (PI) and Annexin V (AV) staining were determined by flow cytometry. Typical flow plots were shown. (b, c) The summary data of early apoptotic cells (PI − AV + ) and late apoptotic or dead cells (PI + AV + ) were shown. The data were described as the mean ± SD for three independent experiments (totally 8 RA-FLSs lines). (d, e) Caspase 3 and cFLIP mRNA expression were measured by qPCR in RA-FLSs. (f) For inhibitor experiments, cells were pretreated with the pan-caspase inhibitor z-VAD-fmk (Sigma-Aldrich, 10 μ M) or N-acetyl-l-cysteine (NAC, Sigma-Aldrich, 5 mM) for 1 h before the treatment of iguratimod. RA-FLSs were treated with TNF- α (25 ng/ml) and iguratimod (0.5 μ g/ml) for 24 h. Apoptosis was determined by flow cytometry. Typical flow plots were shown. The data were shown as the mean ± SD for three independent experiments. The data were analyzed using one-way ANOVA followed by Turkey's test. ∗ P

Techniques Used: Staining, Flow Cytometry, Cytometry, Expressing, Real-time Polymerase Chain Reaction

51) Product Images from "Reactive Oxygen Species Is Essential for Cycloheximide to Sensitize Lexatumumab-Induced Apoptosis in Hepatocellular Carcinoma Cells"

Article Title: Reactive Oxygen Species Is Essential for Cycloheximide to Sensitize Lexatumumab-Induced Apoptosis in Hepatocellular Carcinoma Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0016966

Lexa and CHX combination treatment induces apoptosis in HCC cells. A, Huh7 cells were treated with TNF-α (10 ng/ml) or Lexa (1 µg/ml) for indicated times. Cell lysates were prepared and subjected to Western blotting to detect the expression of IκB-α and phospho-IκB-α. β-actin protein levels were used as an equal protein loading control (Lexa, lexatumumab). B , Huh7 cells were treated with DMSO (Con), CHX (10 µg/ml), Lexa (1 µg/ml), or a combination of Lexa (1 µg/ml) and CHX (10 µg/ml). Apoptosis was measured by nuclear dye Hoechst 33258 staining to label DNA fragmentation (nuclear morphology changes). (Apoptotic cells were marked with arrows). C , Huh7 cells were treated with DMSO (Con), CHX (10 µg/ml), Lexa (1 µg/ml), or combination treatment of Lexa (1 µg/ml) and CHX (10 µg/ml) and apoptosis was evaluated through Annexin V and PI double staining based FACS analysis using the Annexin-V assay kit (see ' materials and methods '). D , The percentage of apoptotic cells were characterized as those that stained with Annexin-V. Data represent the mean values of three independent experiments (*p
Figure Legend Snippet: Lexa and CHX combination treatment induces apoptosis in HCC cells. A, Huh7 cells were treated with TNF-α (10 ng/ml) or Lexa (1 µg/ml) for indicated times. Cell lysates were prepared and subjected to Western blotting to detect the expression of IκB-α and phospho-IκB-α. β-actin protein levels were used as an equal protein loading control (Lexa, lexatumumab). B , Huh7 cells were treated with DMSO (Con), CHX (10 µg/ml), Lexa (1 µg/ml), or a combination of Lexa (1 µg/ml) and CHX (10 µg/ml). Apoptosis was measured by nuclear dye Hoechst 33258 staining to label DNA fragmentation (nuclear morphology changes). (Apoptotic cells were marked with arrows). C , Huh7 cells were treated with DMSO (Con), CHX (10 µg/ml), Lexa (1 µg/ml), or combination treatment of Lexa (1 µg/ml) and CHX (10 µg/ml) and apoptosis was evaluated through Annexin V and PI double staining based FACS analysis using the Annexin-V assay kit (see ' materials and methods '). D , The percentage of apoptotic cells were characterized as those that stained with Annexin-V. Data represent the mean values of three independent experiments (*p

Techniques Used: Western Blot, Expressing, Staining, Double Staining, FACS, Annexin V Assay

52) Product Images from "Downregulation of A20 increases the cytotoxicity of IFN-γ in hepatocellular carcinoma cells"

Article Title: Downregulation of A20 increases the cytotoxicity of IFN-γ in hepatocellular carcinoma cells

Journal: Drug Design, Development and Therapy

doi: 10.2147/DDDT.S135993

A20 knockdown increases the apoptosis induced by IFN-γ in HepG2 cells. Notes: ( A ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were then subjected to flow cytometry analysis using Annexin-V/propidium iodide staining. Quantification of apoptosis was derived from three independent experiments (right). ( B ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cellular lysates were subjected to Western blot analysis with the indicated antibodies. Graphs represent quantification of target protein bands relative to GAPDH. ( C ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were lysed and assayed for caspase-3, -8 and -9 activity. Graphs represent the mean ± SD of three independent experiments. ( D ) sh-A20 cells were incubated with 100 ng/mL of IFN-γ for 24 h after 1 h pretreatment with z-DEVD-fmk, z-LEHD-fmk and z-IETD-fmk, respectively. Cell viability (left) was determined by MTT assay and apoptosis (right) was determined by flow cytometry. The data are represented as the mean ± SD of three independent experiments. The significance was determined by the Student’s t -test (** P
Figure Legend Snippet: A20 knockdown increases the apoptosis induced by IFN-γ in HepG2 cells. Notes: ( A ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were then subjected to flow cytometry analysis using Annexin-V/propidium iodide staining. Quantification of apoptosis was derived from three independent experiments (right). ( B ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cellular lysates were subjected to Western blot analysis with the indicated antibodies. Graphs represent quantification of target protein bands relative to GAPDH. ( C ) sh-NC and sh-A20 cells were treated with or without 100 ng/mL of IFN-γ for 24 h. Cells were lysed and assayed for caspase-3, -8 and -9 activity. Graphs represent the mean ± SD of three independent experiments. ( D ) sh-A20 cells were incubated with 100 ng/mL of IFN-γ for 24 h after 1 h pretreatment with z-DEVD-fmk, z-LEHD-fmk and z-IETD-fmk, respectively. Cell viability (left) was determined by MTT assay and apoptosis (right) was determined by flow cytometry. The data are represented as the mean ± SD of three independent experiments. The significance was determined by the Student’s t -test (** P

Techniques Used: Flow Cytometry, Cytometry, Staining, Derivative Assay, Western Blot, Activity Assay, Incubation, MTT Assay

53) Product Images from "Vorinostat and quinacrine have synergistic effects in T-cell acute lymphoblastic leukemia through reactive oxygen species increase and mitophagy inhibition"

Article Title: Vorinostat and quinacrine have synergistic effects in T-cell acute lymphoblastic leukemia through reactive oxygen species increase and mitophagy inhibition

Journal: Cell Death & Disease

doi: 10.1038/s41419-018-0679-6

Effects of vorinostat and QC on the apoptosis of Jurkat and Molt-4 cells. Jurkat cells were treated with vorinostat (1 μM) and/or QC (5 μM) for 48 h and Molt-4 cells were treated with vorinostat (1 μM) and/or QC (2.5 μM) for 24 h. a , b Cell cycle distribution of Jurkat and Molt-4 cells was analyzed by flow cytometry. c , d Apoptosis rate of Jurkat and Molt-4 cells was established by flow cytometry analysis of Annexin-V-PI dual staining. e The indicated proteins were examined by western blots. * p
Figure Legend Snippet: Effects of vorinostat and QC on the apoptosis of Jurkat and Molt-4 cells. Jurkat cells were treated with vorinostat (1 μM) and/or QC (5 μM) for 48 h and Molt-4 cells were treated with vorinostat (1 μM) and/or QC (2.5 μM) for 24 h. a , b Cell cycle distribution of Jurkat and Molt-4 cells was analyzed by flow cytometry. c , d Apoptosis rate of Jurkat and Molt-4 cells was established by flow cytometry analysis of Annexin-V-PI dual staining. e The indicated proteins were examined by western blots. * p

Techniques Used: Flow Cytometry, Cytometry, Staining, Western Blot

54) Product Images from "RhoA GTPase controls cytokinesis and programmed necrosis of hematopoietic progenitors"

Article Title: RhoA GTPase controls cytokinesis and programmed necrosis of hematopoietic progenitors

Journal: The Journal of Experimental Medicine

doi: 10.1084/jem.20122348

RhoA deficiency causes HPC necrosis but not apoptosis. (A–C) Congenic transplantation recipients described in Fig. 1 G were used, and BM cells were isolated 3 d after poly I:C induction. Cells were gated on the LK and LSK populations and analyzed by flow cytometry for Annexin V/7-AAD staining (A), apoptotic cells (B), and dead cells (C). (D) Cleavage of caspase 3 (CCIII) in Lin − cells of RhoA fl/fl ; Mx-cre + or Mx-cre − mice. Lin − cells were isolated 3 d after poly I:C injections. Staurosporine (STS)-treated WT Lin − cells were used as a positive apoptosis control. (E) CCIII staining of isolated LK cells. (F) p53, Bcl-xL, Bcl-2, and Survivin expression in Lin − cells of RhoA fl/fl ; Mx-cre + or Mx-cre − mice. Protein levels of apoptotic-related proteins were determined at 3 d after induction. (G) Levels of LC3B isoforms in Lin − cells of RhoA fl/fl ; Mx-cre + or Mx-cre − mice. Lin − cells were isolated 3 d after poly I:C injections. 100 nM Rapamycin (RAPA)–treated WT Lin − cells were used a positive autophagy control. (D, F, and G) Molecular masses (kilodaltons) are indicated to the right of the blots. (H) Representative electron micrographs of Lin − cells in native mice. 16 mM H 2 O 2 –treated Lin − cells were used as a positive necrosis control. Red arrowheads indicate features of membrane integrity loss. Green arrowheads indicate enlarged organelles. (I) Expression of TNF–RIP-related genes in LK cells 3 d after induction. (J) Association between cytokinesis failure (4N) and increased necrosis (7-AAD + ) in RhoA -deficient HPCs. BM Lin − cells were isolated at 3 d after induction. (K) Necrosis analysis of LK cells from recipients transplanted with LSK cells reconstituted with exogenous RhoA as described in Fig. 4 F . Numbers of samples analyzed: three (B, C, and J) or four (I and K) per group. (A–K) The results from a representative experiment of two independent experiments are shown. Error bars indicate SEM. *, P
Figure Legend Snippet: RhoA deficiency causes HPC necrosis but not apoptosis. (A–C) Congenic transplantation recipients described in Fig. 1 G were used, and BM cells were isolated 3 d after poly I:C induction. Cells were gated on the LK and LSK populations and analyzed by flow cytometry for Annexin V/7-AAD staining (A), apoptotic cells (B), and dead cells (C). (D) Cleavage of caspase 3 (CCIII) in Lin − cells of RhoA fl/fl ; Mx-cre + or Mx-cre − mice. Lin − cells were isolated 3 d after poly I:C injections. Staurosporine (STS)-treated WT Lin − cells were used as a positive apoptosis control. (E) CCIII staining of isolated LK cells. (F) p53, Bcl-xL, Bcl-2, and Survivin expression in Lin − cells of RhoA fl/fl ; Mx-cre + or Mx-cre − mice. Protein levels of apoptotic-related proteins were determined at 3 d after induction. (G) Levels of LC3B isoforms in Lin − cells of RhoA fl/fl ; Mx-cre + or Mx-cre − mice. Lin − cells were isolated 3 d after poly I:C injections. 100 nM Rapamycin (RAPA)–treated WT Lin − cells were used a positive autophagy control. (D, F, and G) Molecular masses (kilodaltons) are indicated to the right of the blots. (H) Representative electron micrographs of Lin − cells in native mice. 16 mM H 2 O 2 –treated Lin − cells were used as a positive necrosis control. Red arrowheads indicate features of membrane integrity loss. Green arrowheads indicate enlarged organelles. (I) Expression of TNF–RIP-related genes in LK cells 3 d after induction. (J) Association between cytokinesis failure (4N) and increased necrosis (7-AAD + ) in RhoA -deficient HPCs. BM Lin − cells were isolated at 3 d after induction. (K) Necrosis analysis of LK cells from recipients transplanted with LSK cells reconstituted with exogenous RhoA as described in Fig. 4 F . Numbers of samples analyzed: three (B, C, and J) or four (I and K) per group. (A–K) The results from a representative experiment of two independent experiments are shown. Error bars indicate SEM. *, P

Techniques Used: Transplantation Assay, Isolation, Flow Cytometry, Cytometry, Staining, Mouse Assay, Expressing

55) Product Images from "Inhibition of Proliferation and Induction of Apoptosis in Multiple Myeloma Cell Lines by CD137 Ligand Signaling"

Article Title: Inhibition of Proliferation and Induction of Apoptosis in Multiple Myeloma Cell Lines by CD137 Ligand Signaling

Journal: PLoS ONE

doi: 10.1371/journal.pone.0010845

CD137 induces apoptosis in the MM cell lines. (A) SGH-MM5 cells at a density of 10 6 cells/ml were cultured on plate-bound Fc or CD137-Fc protein or on uncoated plates (PBS). After 24 h the cells were stained with Annexin V and 7-AAD. Similar results were obtained for the other MM cell lines. (B) Cells from (A) were stained with Acridine Orange (green) and Ethidium Bromide (red). Photographs were taken at a magnification of 40×. (C) CD137-Fc treated SGH-MM5 cells of B at a magnification of 200×. (D) Caspase 3 activity was determined 6 h after exposure of SGH-MM5 and RPMI 8226 cells to immobilized Fc or CD137-Fc protein. These experiments are representative of three independent experiments with similar results.
Figure Legend Snippet: CD137 induces apoptosis in the MM cell lines. (A) SGH-MM5 cells at a density of 10 6 cells/ml were cultured on plate-bound Fc or CD137-Fc protein or on uncoated plates (PBS). After 24 h the cells were stained with Annexin V and 7-AAD. Similar results were obtained for the other MM cell lines. (B) Cells from (A) were stained with Acridine Orange (green) and Ethidium Bromide (red). Photographs were taken at a magnification of 40×. (C) CD137-Fc treated SGH-MM5 cells of B at a magnification of 200×. (D) Caspase 3 activity was determined 6 h after exposure of SGH-MM5 and RPMI 8226 cells to immobilized Fc or CD137-Fc protein. These experiments are representative of three independent experiments with similar results.

Techniques Used: Cell Culture, Staining, Activity Assay

Requirement of immobilization of CD137 ligand agonists. SGH-MM5 (A and B) or SGH-MM6 (C and D) cells at a density of 10 6 cells/ml were cultured on uncoated plates (PBS), or on plate-bound Fc, CD137-Fc, mouse IgG (MOPC21) or anti-CD137 ligand antibody (clones 5F4 and C65-485) or on uncoated plates (PBS), or to which Fc or CD137-Fc proteins were added soluble at 10 µg/ml. (A) Percentage of dead cells (left panel) and number of total live cells (right panel) were determined after 24 h via trypan blue staining. (B) Extent of apoptosis of cells in (A) was determined by Annexin V and 7-AAD staining. (C) Percentages of dead cells were determined at indicated times via trypan blue staining. (D) IL-8 concentrations in 24 h cell supernatants as determined by ELISA. Depicted are means ± standard deviations of triplicate measurements. * p
Figure Legend Snippet: Requirement of immobilization of CD137 ligand agonists. SGH-MM5 (A and B) or SGH-MM6 (C and D) cells at a density of 10 6 cells/ml were cultured on uncoated plates (PBS), or on plate-bound Fc, CD137-Fc, mouse IgG (MOPC21) or anti-CD137 ligand antibody (clones 5F4 and C65-485) or on uncoated plates (PBS), or to which Fc or CD137-Fc proteins were added soluble at 10 µg/ml. (A) Percentage of dead cells (left panel) and number of total live cells (right panel) were determined after 24 h via trypan blue staining. (B) Extent of apoptosis of cells in (A) was determined by Annexin V and 7-AAD staining. (C) Percentages of dead cells were determined at indicated times via trypan blue staining. (D) IL-8 concentrations in 24 h cell supernatants as determined by ELISA. Depicted are means ± standard deviations of triplicate measurements. * p

Techniques Used: Cell Culture, Staining, Enzyme-linked Immunosorbent Assay

56) Product Images from "Combined HDAC and Bromodomain Protein Inhibition Reprograms Tumor Cell Metabolism and elicits Synthetic Lethality in Glioblastoma"

Article Title: Combined HDAC and Bromodomain Protein Inhibition Reprograms Tumor Cell Metabolism and elicits Synthetic Lethality in Glioblastoma

Journal: Clinical cancer research : an official journal of the American Association for Cancer Research

doi: 10.1158/1078-0432.CCR-18-0260

Dual inhibition of HDAC (Pb) and BRD (OTX) causes synergistic activation of apoptosis in glioblastoma cells A, B, LN229 GBM cells were treated with indicated drugs for 24h, stained with Annexin V/PI and analyzed by flow cytometry. n=3 biological replicates. Shown are means and SD. C, Microarray was performed with NCH644 stem-like GBM cells treated with the combination of OTX+Pb and vehicle DMSO control. Shown is GSEA enrichment plot with FDR q-values (false discovery rate), NES (normalized enrichment score) and p-values. n=2 biological replicates for each condition. D, E, LN229 GBM cells were treated with indicated drugs and stained with tetramethylrhodamine, ethyl ester (TMRE) and analyzed by flow cytometry for the change in mitochondrial membrane potential. n=3 biological replicates. Shown are means and SD. F, Western blotting analysis of LN229 GBM cells treated with indicated drugs. TF: Total form, CF: cleaved form. All concentrations are in μM. G, H, LN229, U87, NCH644 or GBM14 cells were treated with indicated drugs and analyzed for the levels of the indicated proteins by conventional western blotting or capillary electrophoresis. All concentrations are in μM. Blots or capillary electrophoresis were quantified for the levels of Mcl-1 and Noxa normalized with its related loading control. I, J, LN229 or NCH644 GBM cells were treated with indicated drugs and analyzed by real-time PCR for the indicated makers. Shown are means and SD (n=3), and statistical analysis was performed.
Figure Legend Snippet: Dual inhibition of HDAC (Pb) and BRD (OTX) causes synergistic activation of apoptosis in glioblastoma cells A, B, LN229 GBM cells were treated with indicated drugs for 24h, stained with Annexin V/PI and analyzed by flow cytometry. n=3 biological replicates. Shown are means and SD. C, Microarray was performed with NCH644 stem-like GBM cells treated with the combination of OTX+Pb and vehicle DMSO control. Shown is GSEA enrichment plot with FDR q-values (false discovery rate), NES (normalized enrichment score) and p-values. n=2 biological replicates for each condition. D, E, LN229 GBM cells were treated with indicated drugs and stained with tetramethylrhodamine, ethyl ester (TMRE) and analyzed by flow cytometry for the change in mitochondrial membrane potential. n=3 biological replicates. Shown are means and SD. F, Western blotting analysis of LN229 GBM cells treated with indicated drugs. TF: Total form, CF: cleaved form. All concentrations are in μM. G, H, LN229, U87, NCH644 or GBM14 cells were treated with indicated drugs and analyzed for the levels of the indicated proteins by conventional western blotting or capillary electrophoresis. All concentrations are in μM. Blots or capillary electrophoresis were quantified for the levels of Mcl-1 and Noxa normalized with its related loading control. I, J, LN229 or NCH644 GBM cells were treated with indicated drugs and analyzed by real-time PCR for the indicated makers. Shown are means and SD (n=3), and statistical analysis was performed.

Techniques Used: Inhibition, Activation Assay, Staining, Flow Cytometry, Cytometry, Microarray, Western Blot, Electrophoresis, Real-time Polymerase Chain Reaction

Molecular requirements of apoptosis induction by combined inhibition of HDAC and BRD A, LN229 GBM cells were transfected with non-targeting (siNT), BAK, Noxa siRNA. Indicated protein levels were shown. B, C, The same transfected LN229 GBM cells from A were subjected to the combination treatment, OTX+Pb, stained with propidium iodide for flow cytometric analysis. D, T98G GBM cells were treated with vehicle or the OTX+Pb combination for 16h. Thereafter, protein lysates were immunoprecipitated with a species control IgG or Mcl-1 specific antibody and detected by conventional western blotting. The left portion shows the immunoprecipitated lysates, whereas on the right side 1% input lysates were loaded. All concentrations are in μM. E, LN229 GBM cells were transfected with indicated siRNA for 3 days, then treated with indicated drugs for 24h and analyzed for the expression of the indicated proteins by either standard western blotting or capillary electrophoresis. Arrows indicate total and cleavage forms of PARP. All concentrations are in μM. F, LN229 cells treated as in E were stained with propidium iodide and analyzed by flow cytometry for DNA – fragmentation. n=3 biological replicates. Shown are means and SD. G, H, LN229 or NCH644 GBM cells were treated with indicated drugs for 24h and 48h and analyzed for the indicated proteins by capillary electrophoresis. All concentrations are in μM. Quantifications are provided for Mcl-1 normalized with vinculin. J, LN229 GBM cells were treated with indicated drugs and stained with Annexin V/PI and analyzed by flow cytometry. Displayed are representative flow plots and quantification of the results. n=3 biological replicates. Shown are means and SD.
Figure Legend Snippet: Molecular requirements of apoptosis induction by combined inhibition of HDAC and BRD A, LN229 GBM cells were transfected with non-targeting (siNT), BAK, Noxa siRNA. Indicated protein levels were shown. B, C, The same transfected LN229 GBM cells from A were subjected to the combination treatment, OTX+Pb, stained with propidium iodide for flow cytometric analysis. D, T98G GBM cells were treated with vehicle or the OTX+Pb combination for 16h. Thereafter, protein lysates were immunoprecipitated with a species control IgG or Mcl-1 specific antibody and detected by conventional western blotting. The left portion shows the immunoprecipitated lysates, whereas on the right side 1% input lysates were loaded. All concentrations are in μM. E, LN229 GBM cells were transfected with indicated siRNA for 3 days, then treated with indicated drugs for 24h and analyzed for the expression of the indicated proteins by either standard western blotting or capillary electrophoresis. Arrows indicate total and cleavage forms of PARP. All concentrations are in μM. F, LN229 cells treated as in E were stained with propidium iodide and analyzed by flow cytometry for DNA – fragmentation. n=3 biological replicates. Shown are means and SD. G, H, LN229 or NCH644 GBM cells were treated with indicated drugs for 24h and 48h and analyzed for the indicated proteins by capillary electrophoresis. All concentrations are in μM. Quantifications are provided for Mcl-1 normalized with vinculin. J, LN229 GBM cells were treated with indicated drugs and stained with Annexin V/PI and analyzed by flow cytometry. Displayed are representative flow plots and quantification of the results. n=3 biological replicates. Shown are means and SD.

Techniques Used: Inhibition, Transfection, Staining, Flow Cytometry, Immunoprecipitation, Western Blot, Expressing, Electrophoresis, Cytometry

57) Product Images from "Targeting Poly (ADP-Ribose) Polymerase Partially Contributes to Bufalin-Induced Cell Death in Multiple Myeloma Cells"

Article Title: Targeting Poly (ADP-Ribose) Polymerase Partially Contributes to Bufalin-Induced Cell Death in Multiple Myeloma Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0066130

The role of PARP1 in bufalin-mediated apoptosis in MM cells. (A–B) H929 cells were stablely transfected with the control empty vector (pMSCV) or a PARP ovexpression plasmid (ovPARP1), or with the control nonspecific shRNA (pSIREN) or the shRNA against PARP1 (shPARP1), PARP1 expression was measured by western blot. (C) Indicated cells were treated with 20 nM bufalin or 20 µM olaparib for 48 hours. Annexin-V-positive cells were counted using flow cytometry. All values represent means ± S.D. of three independent experiments, each performed in triplicate ( * P
Figure Legend Snippet: The role of PARP1 in bufalin-mediated apoptosis in MM cells. (A–B) H929 cells were stablely transfected with the control empty vector (pMSCV) or a PARP ovexpression plasmid (ovPARP1), or with the control nonspecific shRNA (pSIREN) or the shRNA against PARP1 (shPARP1), PARP1 expression was measured by western blot. (C) Indicated cells were treated with 20 nM bufalin or 20 µM olaparib for 48 hours. Annexin-V-positive cells were counted using flow cytometry. All values represent means ± S.D. of three independent experiments, each performed in triplicate ( * P

Techniques Used: Transfection, Plasmid Preparation, shRNA, Expressing, Western Blot, Flow Cytometry, Cytometry

Bufalin induces apoptosis and cell cycle arrest in MM cells. H929 (A, C, E, G) and U266 (B, D, F, H) cells were treated with or without 20 nM bufalin for 48 h, and cell morphology (A–B) was monitored by phase contrast microscope (40×, left) or Wright's staining (100×, right). Arrowheads indicated apoptotic cells. Annexin-V-positive cells (C–D) were quantified by flow cytometry analysis. Indicated proteins were analyzed by western blot analysis (E–F). Cell cycle distribution was determined by flow cytometry analysis of DNA content (G–H). Each experiment was performed in triplicate and repeated at least three times.
Figure Legend Snippet: Bufalin induces apoptosis and cell cycle arrest in MM cells. H929 (A, C, E, G) and U266 (B, D, F, H) cells were treated with or without 20 nM bufalin for 48 h, and cell morphology (A–B) was monitored by phase contrast microscope (40×, left) or Wright's staining (100×, right). Arrowheads indicated apoptotic cells. Annexin-V-positive cells (C–D) were quantified by flow cytometry analysis. Indicated proteins were analyzed by western blot analysis (E–F). Cell cycle distribution was determined by flow cytometry analysis of DNA content (G–H). Each experiment was performed in triplicate and repeated at least three times.

Techniques Used: Microscopy, Staining, Flow Cytometry, Cytometry, Western Blot

58) Product Images from "Liposomal C6 Ceramide Activates Protein Phosphatase 1 to Inhibit Melanoma Cells"

Article Title: Liposomal C6 Ceramide Activates Protein Phosphatase 1 to Inhibit Melanoma Cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0159849

Activation of PP1 is required for liposomal C6-induced anti-melanoma cell activity in vitro . Stable WM-115 cells expressing the pan PP1 shRNA, constitutively-activate mutant Akt1 (“CA-Akt1”), or empty vector (“pSuper-puro”) were treated with or without applied concentration of liposomal C6 ceramide (“Lipo C6”), expression of listed proteins was tested by Western blots (A); Relative protein phosphatase activity was shown (B); Cell survival (C), and cell apoptosis (D) were tested by MTT assay and Annexin V assay, respectively. Experiments were repeated three times, and similar results were obtained. Data were presented as mean ± SD. *p
Figure Legend Snippet: Activation of PP1 is required for liposomal C6-induced anti-melanoma cell activity in vitro . Stable WM-115 cells expressing the pan PP1 shRNA, constitutively-activate mutant Akt1 (“CA-Akt1”), or empty vector (“pSuper-puro”) were treated with or without applied concentration of liposomal C6 ceramide (“Lipo C6”), expression of listed proteins was tested by Western blots (A); Relative protein phosphatase activity was shown (B); Cell survival (C), and cell apoptosis (D) were tested by MTT assay and Annexin V assay, respectively. Experiments were repeated three times, and similar results were obtained. Data were presented as mean ± SD. *p

Techniques Used: Activation Assay, Activity Assay, In Vitro, Expressing, shRNA, Mutagenesis, Plasmid Preparation, Concentration Assay, Western Blot, MTT Assay, Annexin V Assay

59) Product Images from "Attenuated DNA damage responses and increased apoptosis characterize human hematopoietic stem cells exposed to irradiation"

Article Title: Attenuated DNA damage responses and increased apoptosis characterize human hematopoietic stem cells exposed to irradiation

Journal: Scientific Reports

doi: 10.1038/s41598-018-24440-w

Analysis of IR-induced apoptosis in different fractions of primitive human hematopoietic cells. (A) Flow cytometry gating strategy for HSPCs and CPs. (B,D) Example of FACS data identifying cPARP + (B) and AnnexinV + (D) fractions in HSPCs and CPs gated as described in (A) 6 h after 3 Gy IR-treatment. (C,E) CD34 + cells were pre-treated with Z-vad-FMK (100 μM), Q-VD-OPh (20 μM) or DMSO for 1 h, irradiated with 3 Gy and analyzed for cPARP+ or Annexin V+ cells in HSPCs and CPs 6 h later, n ≥ 3. ( F ) Survival of clonogenic cells from sorted HSPC (CD34 + CD38 −/low CD90 + 45RA − ), CMP (CD34 + CD38 + CD45RA − CD135 + ), GMP (CD34 + CD38 + CD45RA + CD135 + ) and MEP (CD34 + CD38 + CD45RA − CD135 − ) fractions following IR with 3 Gy. CFC surviving fraction was calculated by dividing number of colonies counted in the irradiated plates by the number of colonies from the same fraction scored in the non-irradiated plates, n = 3. Represented are mean values ± SD; *p
Figure Legend Snippet: Analysis of IR-induced apoptosis in different fractions of primitive human hematopoietic cells. (A) Flow cytometry gating strategy for HSPCs and CPs. (B,D) Example of FACS data identifying cPARP + (B) and AnnexinV + (D) fractions in HSPCs and CPs gated as described in (A) 6 h after 3 Gy IR-treatment. (C,E) CD34 + cells were pre-treated with Z-vad-FMK (100 μM), Q-VD-OPh (20 μM) or DMSO for 1 h, irradiated with 3 Gy and analyzed for cPARP+ or Annexin V+ cells in HSPCs and CPs 6 h later, n ≥ 3. ( F ) Survival of clonogenic cells from sorted HSPC (CD34 + CD38 −/low CD90 + 45RA − ), CMP (CD34 + CD38 + CD45RA − CD135 + ), GMP (CD34 + CD38 + CD45RA + CD135 + ) and MEP (CD34 + CD38 + CD45RA − CD135 − ) fractions following IR with 3 Gy. CFC surviving fraction was calculated by dividing number of colonies counted in the irradiated plates by the number of colonies from the same fraction scored in the non-irradiated plates, n = 3. Represented are mean values ± SD; *p

Techniques Used: Flow Cytometry, Cytometry, FACS, Irradiation

60) Product Images from "Emetine induces chemosensitivity and reduces clonogenicity of acute myeloid leukemia cells"

Article Title: Emetine induces chemosensitivity and reduces clonogenicity of acute myeloid leukemia cells

Journal: Oncotarget

doi: 10.18632/oncotarget.8096

Emetine treatment reduced cell viability, induced apoptosis, prompted AML cells towards differentiation and downregulated HIF-1α A. HL-60, KG-1, MonoMac-1 (MM), Kasumi-1 (K-1) and THP-1 AML cell lines were treated for 24 h at different concentrations of emetine (μM). Each point represents the mean value of a biological triplicate and error bars represent SEM. Y-axis: number of live cells relative to vehicle-treated control as assessed by flow cytometry (correct FSC-SSC profile and 7-AAD − Hoechst low ). The EC50 is indicated in the nM range. B. MonoMac-1 and Kasumi-1 cell lines were treated with 150 nM emetine (red) or with vehicle control (light grey) for 24 h. Annexin-V staining was measured by flow cytometry. Data from both cell lines are presented combined ( n = 6 for each line) (left panel). Y-axis: relative frequency of early and late apoptotic cells. Early apoptotic: Annexin V + , 7-AAD − ; late apoptotic: Annexin V + , 7-AAD + . Representative flow cytometry plot of vehicle-treated or emetine-treated MM (right panel). C. HL-60, KG-1, MonoMac-1 and Kasumi-1 cells were treated with 150 nM emetine (red) for 24 h or vehicle control (light grey) and cell cycle was analyzed by flow cytometry. Relative frequency of G0/G1, S and G2/M phases is presented (left panel). Bars represent the mean value of all AML cell lines ( n = 6 for each cell line) and error bars represent SEM. Representative DNA content flow profile of control and emetine-treated KG-1 cells (right panel). Green represents G0/G1 phase; yellow, S-phase; blue, G2/M. HL-60, KG-1 and Kasumi-1 cells were treated with 150 nM emetine (E, red) for 24 h or with vehicle control (C, light grey). D. and E. C/EBPα (K-1) and PU.1 (HL-60, KG-1) mRNA levels are represented in K-1 and HL-60 and KG-1 cell lines upon treatment with emetine (E) compared to control (C). F . HL-60 and KG-1 cells were treated for 24 h with 50 μM CoCl 2 (C), 50 μM CoCl 2 plus 150 nM emetine (C+E) and vehicle control (Ø) [ 34 ]. HIF-1α was detected by Western Blot. Left panel represents HIF-1α protein level normalized to β-actin and refer to CoCl 2 control as assessed by band densitometry. Data from all 5 cell lines are presented combined. Error bar represents SEM. Right panel shows a representative Western blot of HL-60 samples. * p
Figure Legend Snippet: Emetine treatment reduced cell viability, induced apoptosis, prompted AML cells towards differentiation and downregulated HIF-1α A. HL-60, KG-1, MonoMac-1 (MM), Kasumi-1 (K-1) and THP-1 AML cell lines were treated for 24 h at different concentrations of emetine (μM). Each point represents the mean value of a biological triplicate and error bars represent SEM. Y-axis: number of live cells relative to vehicle-treated control as assessed by flow cytometry (correct FSC-SSC profile and 7-AAD − Hoechst low ). The EC50 is indicated in the nM range. B. MonoMac-1 and Kasumi-1 cell lines were treated with 150 nM emetine (red) or with vehicle control (light grey) for 24 h. Annexin-V staining was measured by flow cytometry. Data from both cell lines are presented combined ( n = 6 for each line) (left panel). Y-axis: relative frequency of early and late apoptotic cells. Early apoptotic: Annexin V + , 7-AAD − ; late apoptotic: Annexin V + , 7-AAD + . Representative flow cytometry plot of vehicle-treated or emetine-treated MM (right panel). C. HL-60, KG-1, MonoMac-1 and Kasumi-1 cells were treated with 150 nM emetine (red) for 24 h or vehicle control (light grey) and cell cycle was analyzed by flow cytometry. Relative frequency of G0/G1, S and G2/M phases is presented (left panel). Bars represent the mean value of all AML cell lines ( n = 6 for each cell line) and error bars represent SEM. Representative DNA content flow profile of control and emetine-treated KG-1 cells (right panel). Green represents G0/G1 phase; yellow, S-phase; blue, G2/M. HL-60, KG-1 and Kasumi-1 cells were treated with 150 nM emetine (E, red) for 24 h or with vehicle control (C, light grey). D. and E. C/EBPα (K-1) and PU.1 (HL-60, KG-1) mRNA levels are represented in K-1 and HL-60 and KG-1 cell lines upon treatment with emetine (E) compared to control (C). F . HL-60 and KG-1 cells were treated for 24 h with 50 μM CoCl 2 (C), 50 μM CoCl 2 plus 150 nM emetine (C+E) and vehicle control (Ø) [ 34 ]. HIF-1α was detected by Western Blot. Left panel represents HIF-1α protein level normalized to β-actin and refer to CoCl 2 control as assessed by band densitometry. Data from all 5 cell lines are presented combined. Error bar represents SEM. Right panel shows a representative Western blot of HL-60 samples. * p

Techniques Used: Flow Cytometry, Cytometry, Staining, Western Blot

61) Product Images from "A protease-resistant PML-RAR? has increased leukemogenic potential in a murine model of acute promyelocytic leukemia"

Article Title: A protease-resistant PML-RAR? has increased leukemogenic potential in a murine model of acute promyelocytic leukemia

Journal: Blood

doi: 10.1182/blood-2008-11-189282

Expression of PR 2VR in myeloid cell lines . (A) Staining for PML oncogenic domains in K562 cell lines transfected with GFP-PR WT or GFP-PR 2VR (B) Analysis of apoptosis by annexin V staining in U937 cells at 24 hours after transfection with GFP-PR WT or GFP-PR
Figure Legend Snippet: Expression of PR 2VR in myeloid cell lines . (A) Staining for PML oncogenic domains in K562 cell lines transfected with GFP-PR WT or GFP-PR 2VR (B) Analysis of apoptosis by annexin V staining in U937 cells at 24 hours after transfection with GFP-PR WT or GFP-PR

Techniques Used: Expressing, Staining, Transfection

62) Product Images from "Inhibition of Bcl-2/Bcl-xL and c-MET causes synthetic lethality in model systems of glioblastoma"

Article Title: Inhibition of Bcl-2/Bcl-xL and c-MET causes synthetic lethality in model systems of glioblastoma

Journal: Scientific Reports

doi: 10.1038/s41598-018-25802-0

Enhanced apoptosis induction and dissipation of mitochondrial membrane potential by c-MET and Bcl-2/Bcl-xL inhibition. ( A) LN229 GBM cells were treated with ABT263 (ABT), Crizotinib (Crizo) or the combination of both and analyzed for mitochondrial membrane potential by TMRE staining with subsequent flow cytometry. ( B ) LN229 GBM cells were treated as in A, stained with Annexin V/propidium iodide and analyzed by multi-parametric flow cytometry. ( C–E ) Quantifications of Annexin and TMRE staining flow cytometric results in LN229, U87 and stem-like GBM cells. Shown are means and SD. n = 3. ( F ) LN229 and U87 GBM cells were treated with ABT263, Crizotinib (Crizo) and the combination treatment (1d). Whole cell protein lysates were prepared and analyzed by conventional Western blotting for the expression of PARP, cCP9 (cleaved caspase 9), cCP3 (cleaved Caspase-3). Actin serves as loading control. Arrows highlight the cleavage fragments of the respective proteins. Concentrations are in μM.
Figure Legend Snippet: Enhanced apoptosis induction and dissipation of mitochondrial membrane potential by c-MET and Bcl-2/Bcl-xL inhibition. ( A) LN229 GBM cells were treated with ABT263 (ABT), Crizotinib (Crizo) or the combination of both and analyzed for mitochondrial membrane potential by TMRE staining with subsequent flow cytometry. ( B ) LN229 GBM cells were treated as in A, stained with Annexin V/propidium iodide and analyzed by multi-parametric flow cytometry. ( C–E ) Quantifications of Annexin and TMRE staining flow cytometric results in LN229, U87 and stem-like GBM cells. Shown are means and SD. n = 3. ( F ) LN229 and U87 GBM cells were treated with ABT263, Crizotinib (Crizo) and the combination treatment (1d). Whole cell protein lysates were prepared and analyzed by conventional Western blotting for the expression of PARP, cCP9 (cleaved caspase 9), cCP3 (cleaved Caspase-3). Actin serves as loading control. Arrows highlight the cleavage fragments of the respective proteins. Concentrations are in μM.

Techniques Used: Inhibition, Staining, Flow Cytometry, Cytometry, Western Blot, Expressing

63) Product Images from "Mechanisms of the anti-tumor activity of Methyl 2-(-5-fluoro-2-hydroxyphenyl)-1 H-benzo[d]imidazole-5-carboxylate against breast cancer in vitro and in vivo"

Article Title: Mechanisms of the anti-tumor activity of Methyl 2-(-5-fluoro-2-hydroxyphenyl)-1 H-benzo[d]imidazole-5-carboxylate against breast cancer in vitro and in vivo

Journal: Oncotarget

doi: 10.18632/oncotarget.16263

MBIC induces caspase-dependent apoptosis ( A ) Two-dimensional forward and side scatter plots of FITC-conjugated Annexin V vs PI were generated using FACS technology when cells were treated with various concentrations (0.7, 1.5 and 3 μM against MCF-7; 20, 40 and 80 μM against MDA-MB-231) of MBIC for 24 h. Representative figure shows viable cells accumulation (Q3) vs early apoptotic (Q4), late apoptotic (Q2) and necrotic cells (Q1). ( B ) MCF-7 and MDA-MB-231 cells were treated with various concentrations of MBIC (0.7, 1.5 and 3 μM against MCF-7; 20, 40 and 80 μM against MDA-MB-231 cells) before measuring protein level of Bax and cleaved caspases-3/7/9 (Cl-C-3/7/9) with Western blot analysis. β-actin was used as loading control. ( C ) The relative intensity of each protein was normalized with β-actin. Data were results of three independent experiments with mean ± SD. All the treatment groups were compared with control.”*” indicates statistically significant at P
Figure Legend Snippet: MBIC induces caspase-dependent apoptosis ( A ) Two-dimensional forward and side scatter plots of FITC-conjugated Annexin V vs PI were generated using FACS technology when cells were treated with various concentrations (0.7, 1.5 and 3 μM against MCF-7; 20, 40 and 80 μM against MDA-MB-231) of MBIC for 24 h. Representative figure shows viable cells accumulation (Q3) vs early apoptotic (Q4), late apoptotic (Q2) and necrotic cells (Q1). ( B ) MCF-7 and MDA-MB-231 cells were treated with various concentrations of MBIC (0.7, 1.5 and 3 μM against MCF-7; 20, 40 and 80 μM against MDA-MB-231 cells) before measuring protein level of Bax and cleaved caspases-3/7/9 (Cl-C-3/7/9) with Western blot analysis. β-actin was used as loading control. ( C ) The relative intensity of each protein was normalized with β-actin. Data were results of three independent experiments with mean ± SD. All the treatment groups were compared with control.”*” indicates statistically significant at P

Techniques Used: Generated, FACS, Multiple Displacement Amplification, Western Blot

64) Product Images from "Thy-1 interaction with Fas in lipid rafts regulates fibroblast apoptosis and lung injury resolution"

Article Title: Thy-1 interaction with Fas in lipid rafts regulates fibroblast apoptosis and lung injury resolution

Journal: Laboratory investigation; a journal of technical methods and pathology

doi: 10.1038/labinvest.2016.145

Inhibition of caspase-8 decreases cleaved caspase 3 and abolishes the induction of apoptosis by rhFasL in Thy-1 (+) cells A. Caspase-8 inhibitor was added to a final concentrations indicated for 30 min and then Thy-1 (+) RFL-6 were treated with 50ng/mL rhFasL for 16h. Immunoblotting with anti-cleaved caspase-3 identifies caspase-3 cleavage products in Thy-1 (+) RFL-6. B. Thy-1 (+) cells were pretreated with 20μM of caspase 8 inhibitor negative control or caspase 8 specific inhibitor for 30 min followed by 50ng/mL rhFasL for 16h, and then cells were collected and stained with Annexin V and PI followed by flow cytometry as described in Methods. Bar graph shows average of three independent experiments of FACS analysis with FITC-Annexin V and PI. * p
Figure Legend Snippet: Inhibition of caspase-8 decreases cleaved caspase 3 and abolishes the induction of apoptosis by rhFasL in Thy-1 (+) cells A. Caspase-8 inhibitor was added to a final concentrations indicated for 30 min and then Thy-1 (+) RFL-6 were treated with 50ng/mL rhFasL for 16h. Immunoblotting with anti-cleaved caspase-3 identifies caspase-3 cleavage products in Thy-1 (+) RFL-6. B. Thy-1 (+) cells were pretreated with 20μM of caspase 8 inhibitor negative control or caspase 8 specific inhibitor for 30 min followed by 50ng/mL rhFasL for 16h, and then cells were collected and stained with Annexin V and PI followed by flow cytometry as described in Methods. Bar graph shows average of three independent experiments of FACS analysis with FITC-Annexin V and PI. * p

Techniques Used: Inhibition, Negative Control, Staining, Flow Cytometry, Cytometry, FACS

Thy-1 expression promotes fibroblast susceptibility to apoptosis A. RFL-6 Thy-1 (−) and Thy-1 (+) fibroblasts were treated with the indicated concentrations of rhFasL for 16h, and then cells were harvested and stained with Annexin V and PI and apoptosis quantified by flow cytometry. Results are representative of three independent experiments. Upper panels: Dot plots of FACS analysis with FITC-Annexin V and PI. The lower-left quadrants of the dot plot panels represent live cells (Annexin V−/PI−); the lower-right quadrants of the panels quadrants represent early apoptotic cells (Annexin V+/PI−); the upper-right quadrants of the panels quadrants represent later apoptotic cells (Annexin V+/PI+). Lower panel shows a quantitative representation of the flow cytograms. Each bar represents the mean±SEM of early + late apoptosis for at least three experiments. * denotes p
Figure Legend Snippet: Thy-1 expression promotes fibroblast susceptibility to apoptosis A. RFL-6 Thy-1 (−) and Thy-1 (+) fibroblasts were treated with the indicated concentrations of rhFasL for 16h, and then cells were harvested and stained with Annexin V and PI and apoptosis quantified by flow cytometry. Results are representative of three independent experiments. Upper panels: Dot plots of FACS analysis with FITC-Annexin V and PI. The lower-left quadrants of the dot plot panels represent live cells (Annexin V−/PI−); the lower-right quadrants of the panels quadrants represent early apoptotic cells (Annexin V+/PI−); the upper-right quadrants of the panels quadrants represent later apoptotic cells (Annexin V+/PI+). Lower panel shows a quantitative representation of the flow cytograms. Each bar represents the mean±SEM of early + late apoptosis for at least three experiments. * denotes p

Techniques Used: Expressing, Staining, Flow Cytometry, Cytometry, FACS

65) Product Images from "Inhibition of Bcl-2/Bcl-xL and c-MET causes synthetic lethality in model systems of glioblastoma"

Article Title: Inhibition of Bcl-2/Bcl-xL and c-MET causes synthetic lethality in model systems of glioblastoma

Journal: Scientific Reports

doi: 10.1038/s41598-018-25802-0

Enhanced apoptosis induction and dissipation of mitochondrial membrane potential by c-MET and Bcl-2/Bcl-xL inhibition. ( A) LN229 GBM cells were treated with ABT263 (ABT), Crizotinib (Crizo) or the combination of both and analyzed for mitochondrial membrane potential by TMRE staining with subsequent flow cytometry. ( B ) LN229 GBM cells were treated as in A, stained with Annexin V/propidium iodide and analyzed by multi-parametric flow cytometry. ( C–E ) Quantifications of Annexin and TMRE staining flow cytometric results in LN229, U87 and stem-like GBM cells. Shown are means and SD. n = 3. ( F ) LN229 and U87 GBM cells were treated with ABT263, Crizotinib (Crizo) and the combination treatment (1d). Whole cell protein lysates were prepared and analyzed by conventional Western blotting for the expression of PARP, cCP9 (cleaved caspase 9), cCP3 (cleaved Caspase-3). Actin serves as loading control. Arrows highlight the cleavage fragments of the respective proteins. Concentrations are in μM.
Figure Legend Snippet: Enhanced apoptosis induction and dissipation of mitochondrial membrane potential by c-MET and Bcl-2/Bcl-xL inhibition. ( A) LN229 GBM cells were treated with ABT263 (ABT), Crizotinib (Crizo) or the combination of both and analyzed for mitochondrial membrane potential by TMRE staining with subsequent flow cytometry. ( B ) LN229 GBM cells were treated as in A, stained with Annexin V/propidium iodide and analyzed by multi-parametric flow cytometry. ( C–E ) Quantifications of Annexin and TMRE staining flow cytometric results in LN229, U87 and stem-like GBM cells. Shown are means and SD. n = 3. ( F ) LN229 and U87 GBM cells were treated with ABT263, Crizotinib (Crizo) and the combination treatment (1d). Whole cell protein lysates were prepared and analyzed by conventional Western blotting for the expression of PARP, cCP9 (cleaved caspase 9), cCP3 (cleaved Caspase-3). Actin serves as loading control. Arrows highlight the cleavage fragments of the respective proteins. Concentrations are in μM.

Techniques Used: Inhibition, Staining, Flow Cytometry, Cytometry, Western Blot, Expressing

66) Product Images from "Elucidating the mechanism of action of domatinostat (4SC-202) in cutaneous T cell lymphoma cells"

Article Title: Elucidating the mechanism of action of domatinostat (4SC-202) in cutaneous T cell lymphoma cells

Journal: Journal of Hematology & Oncology

doi: 10.1186/s13045-019-0719-4

4SC-202 induces G2/M arrest and apoptosis in CTCL cell lines at concentrations hardly affecting acetylation and methylation of histone proteins. CRL-2105 and HTB-176 cells were treated with incremental doses of 4SC-202 or FK228. a After 24 and 48 h, cellular DNA content was analyzed by PI staining of fixed cells, and apoptosis was assessed by annexin V/7-AAD double staining. Early and late apoptotic cells were identified as 7-AAD − /Annexin V + and 7-AAD + /Annexin V + , respectively. Mean values (± standard deviation (SD)) of three independent experiments are depicted. b Total cell lysates harvested after 48 h of drug treatment were subjected to immunoblot analysis using antibodies recognizing the indicated targets (H3K4me2: histone H3 dimethyl lysine 4; H3K9ac: histone H3 acetyl lysine 9)
Figure Legend Snippet: 4SC-202 induces G2/M arrest and apoptosis in CTCL cell lines at concentrations hardly affecting acetylation and methylation of histone proteins. CRL-2105 and HTB-176 cells were treated with incremental doses of 4SC-202 or FK228. a After 24 and 48 h, cellular DNA content was analyzed by PI staining of fixed cells, and apoptosis was assessed by annexin V/7-AAD double staining. Early and late apoptotic cells were identified as 7-AAD − /Annexin V + and 7-AAD + /Annexin V + , respectively. Mean values (± standard deviation (SD)) of three independent experiments are depicted. b Total cell lysates harvested after 48 h of drug treatment were subjected to immunoblot analysis using antibodies recognizing the indicated targets (H3K4me2: histone H3 dimethyl lysine 4; H3K9ac: histone H3 acetyl lysine 9)

Techniques Used: Methylation, Staining, Double Staining, Standard Deviation

67) Product Images from "Bufalin Enhances the Cytotoxity of Human Multiple Myeloma Cells H929 to AKT Inhibitor MK2206: The Role of Protein AKT Phosphorylation"

Article Title: Bufalin Enhances the Cytotoxity of Human Multiple Myeloma Cells H929 to AKT Inhibitor MK2206: The Role of Protein AKT Phosphorylation

Journal: Indian Journal of Hematology & Blood Transfusion

doi: 10.1007/s12288-017-0883-z

Effect of bufalin and MK2206 on apoptosis in H929 cells. Values indicating Annexin V+/PI− and Annexin V+/PI+ cells detected by flow cytometry were expressed as mean ± SD of three independent experiments
Figure Legend Snippet: Effect of bufalin and MK2206 on apoptosis in H929 cells. Values indicating Annexin V+/PI− and Annexin V+/PI+ cells detected by flow cytometry were expressed as mean ± SD of three independent experiments

Techniques Used: Flow Cytometry, Cytometry

68) Product Images from "Attenuated DNA damage responses and increased apoptosis characterize human hematopoietic stem cells exposed to irradiation"

Article Title: Attenuated DNA damage responses and increased apoptosis characterize human hematopoietic stem cells exposed to irradiation

Journal: Scientific Reports

doi: 10.1038/s41598-018-24440-w

Analysis of IR-induced apoptosis in different fractions of primitive human hematopoietic cells. (A) Flow cytometry gating strategy for HSPCs and CPs. (B,D) Example of FACS data identifying cPARP + (B) and AnnexinV + (D) fractions in HSPCs and CPs gated as described in (A) 6 h after 3 Gy IR-treatment. (C,E) CD34 + cells were pre-treated with Z-vad-FMK (100 μM), Q-VD-OPh (20 μM) or DMSO for 1 h, irradiated with 3 Gy and analyzed for cPARP+ or Annexin V+ cells in HSPCs and CPs 6 h later, n ≥ 3. ( F ) Survival of clonogenic cells from sorted HSPC (CD34 + CD38 −/low CD90 + 45RA − ), CMP (CD34 + CD38 + CD45RA − CD135 + ), GMP (CD34 + CD38 + CD45RA + CD135 + ) and MEP (CD34 + CD38 + CD45RA − CD135 − ) fractions following IR with 3 Gy. CFC surviving fraction was calculated by dividing number of colonies counted in the irradiated plates by the number of colonies from the same fraction scored in the non-irradiated plates, n = 3. Represented are mean values ± SD; *p
Figure Legend Snippet: Analysis of IR-induced apoptosis in different fractions of primitive human hematopoietic cells. (A) Flow cytometry gating strategy for HSPCs and CPs. (B,D) Example of FACS data identifying cPARP + (B) and AnnexinV + (D) fractions in HSPCs and CPs gated as described in (A) 6 h after 3 Gy IR-treatment. (C,E) CD34 + cells were pre-treated with Z-vad-FMK (100 μM), Q-VD-OPh (20 μM) or DMSO for 1 h, irradiated with 3 Gy and analyzed for cPARP+ or Annexin V+ cells in HSPCs and CPs 6 h later, n ≥ 3. ( F ) Survival of clonogenic cells from sorted HSPC (CD34 + CD38 −/low CD90 + 45RA − ), CMP (CD34 + CD38 + CD45RA − CD135 + ), GMP (CD34 + CD38 + CD45RA + CD135 + ) and MEP (CD34 + CD38 + CD45RA − CD135 − ) fractions following IR with 3 Gy. CFC surviving fraction was calculated by dividing number of colonies counted in the irradiated plates by the number of colonies from the same fraction scored in the non-irradiated plates, n = 3. Represented are mean values ± SD; *p

Techniques Used: Flow Cytometry, Cytometry, FACS, Irradiation

69) Product Images from "PKK deficiency in B cells prevents lupus development in Sle lupus mice"

Article Title: PKK deficiency in B cells prevents lupus development in Sle lupus mice

Journal: Immunology letters

doi: 10.1016/j.imlet.2017.03.002

Impaired proliferation and survival of PKK-deficient Sle B cells A. Decreased viability of the B cells from the PKK-deficient Sle mice in response to anti-IgM or lipopolysaccharide (LPS) stimulation. Purified spleen B cells were treated as indicated. The viable cells were determined by trypan blue exclusion assay and are depicted as the percentage of the initial input. All experiments were performed in triplicate. B. Increased apoptosis of PKK-deficient Sle B cells in response to anti-IgM treatment. Splenocytes isolated from each mouse strain were treated with anti-IgM antibody for 30 hours. Cells were stained with Annexin V and 7AAD, and flow cytometric analysis was performed on gated lymphoid and B220 + cells. Shown are representative flow cytometry profiles from two independent experiments. C. Defective proliferation of PKK-deficient Sle B cells in response to anti-IgM and LPS stimulation. Splenocytes were labeled with CFSE and were either left in the medium without any added stimuli (shaded area) or stimulated with F(ab′) 2 anti-IgM or LPS for 30 hours (black line). Analysis was carried out on B220 gated cells. D . Flow cytometric analysis of CD69 (left panels) and CD86 (right panels) expression in B220 gated B cells after anti-IgM and LPS stimulation. Overlays of CD histograms (black line) on unstained controls (shaded) are shown. E. Decreased calcium signaling in PKK-deficient Sle B cells. Splenocytes from indicated mice strains were loaded with 1 μM Fura Red to evaluate Ca 2+ flux in response to anti-IgM stimulation. B220 + cells were gated for the analysis. Data in panels B–E are representative of two independent experiments. 8-month-old mice were used in the experiments described in this figure.
Figure Legend Snippet: Impaired proliferation and survival of PKK-deficient Sle B cells A. Decreased viability of the B cells from the PKK-deficient Sle mice in response to anti-IgM or lipopolysaccharide (LPS) stimulation. Purified spleen B cells were treated as indicated. The viable cells were determined by trypan blue exclusion assay and are depicted as the percentage of the initial input. All experiments were performed in triplicate. B. Increased apoptosis of PKK-deficient Sle B cells in response to anti-IgM treatment. Splenocytes isolated from each mouse strain were treated with anti-IgM antibody for 30 hours. Cells were stained with Annexin V and 7AAD, and flow cytometric analysis was performed on gated lymphoid and B220 + cells. Shown are representative flow cytometry profiles from two independent experiments. C. Defective proliferation of PKK-deficient Sle B cells in response to anti-IgM and LPS stimulation. Splenocytes were labeled with CFSE and were either left in the medium without any added stimuli (shaded area) or stimulated with F(ab′) 2 anti-IgM or LPS for 30 hours (black line). Analysis was carried out on B220 gated cells. D . Flow cytometric analysis of CD69 (left panels) and CD86 (right panels) expression in B220 gated B cells after anti-IgM and LPS stimulation. Overlays of CD histograms (black line) on unstained controls (shaded) are shown. E. Decreased calcium signaling in PKK-deficient Sle B cells. Splenocytes from indicated mice strains were loaded with 1 μM Fura Red to evaluate Ca 2+ flux in response to anti-IgM stimulation. B220 + cells were gated for the analysis. Data in panels B–E are representative of two independent experiments. 8-month-old mice were used in the experiments described in this figure.

Techniques Used: Mouse Assay, Purification, Trypan Blue Exclusion Assay, Isolation, Staining, Flow Cytometry, Cytometry, Labeling, Expressing

70) Product Images from "MK2206 enhances the cytocidal effects of bufalin in multiple myeloma by inhibiting the AKT/mTOR pathway"

Article Title: MK2206 enhances the cytocidal effects of bufalin in multiple myeloma by inhibiting the AKT/mTOR pathway

Journal: Cell Death & Disease

doi: 10.1038/cddis.2017.188

MK2206 enhanced the induction of apoptosis by bufalin in primary myeloma cells. ( a ) Patients’ mononuclear cells were separated by Ficoll–Hipaque density sedimentation and CD138-positive cells were isolated and treated with 12 nM of bufalin alone and/or in addition of 6 μ M of MK2206 for 48 h. The survival rates were assessed by Annexin V/PI staining. ( b ) Freshly isolated PBMCs from 3 healthy donors were cultured with 12 nM of bufalin and 6 μ M of MK2206 for 48 h. The viability was assessed by the tryphan blue assay. Each bar represented the mean±S.E. of triplicate experiments (* P
Figure Legend Snippet: MK2206 enhanced the induction of apoptosis by bufalin in primary myeloma cells. ( a ) Patients’ mononuclear cells were separated by Ficoll–Hipaque density sedimentation and CD138-positive cells were isolated and treated with 12 nM of bufalin alone and/or in addition of 6 μ M of MK2206 for 48 h. The survival rates were assessed by Annexin V/PI staining. ( b ) Freshly isolated PBMCs from 3 healthy donors were cultured with 12 nM of bufalin and 6 μ M of MK2206 for 48 h. The viability was assessed by the tryphan blue assay. Each bar represented the mean±S.E. of triplicate experiments (* P

Techniques Used: Sedimentation, Isolation, Staining, Cell Culture

71) Product Images from "Protopanaxadiol, an Active Ginseng Metabolite, Significantly Enhances the Effects of Fluorouracil on Colon Cancer"

Article Title: Protopanaxadiol, an Active Ginseng Metabolite, Significantly Enhances the Effects of Fluorouracil on Colon Cancer

Journal: Nutrients

doi: 10.3390/nu7020799

Effects of protopanaxadiol (PPD) on 5-FU-induced apoptosis. HCT-116 human colorectal cancer cells were treated with PPD at various concentrations (10 and 20 µM) in the absence or presence of 5-FU (30 µM) for 48 h. Apoptosis was quantified using annexin V/PI staining followed by flow cytometric analysis. ( A ) Representative scatter plots of PI ( y -axis) vs. annexin V ( x -axis) in each experimental group; ( B ) the percentage of apoptotic cells. Data obtained from triplicate experiments. + p
Figure Legend Snippet: Effects of protopanaxadiol (PPD) on 5-FU-induced apoptosis. HCT-116 human colorectal cancer cells were treated with PPD at various concentrations (10 and 20 µM) in the absence or presence of 5-FU (30 µM) for 48 h. Apoptosis was quantified using annexin V/PI staining followed by flow cytometric analysis. ( A ) Representative scatter plots of PI ( y -axis) vs. annexin V ( x -axis) in each experimental group; ( B ) the percentage of apoptotic cells. Data obtained from triplicate experiments. + p

Techniques Used: Staining, Flow Cytometry

72) Product Images from "Ubiquitin-specific protease 4 promotes glioblastoma multiforme via activating ERK pathway"

Article Title: Ubiquitin-specific protease 4 promotes glioblastoma multiforme via activating ERK pathway

Journal: OncoTargets and therapy

doi: 10.2147/OTT.S176582

Silencing of USP4 promoted cell apoptosis in GBM cell lines. Notes: ( A ) Cell apoptosis was detected by flow cytometry using annexin-V/PI kit in shUSP4- or NC-transduced U87 and T98G cells. ( B , C ) Real-time PCR ( B ) and Western blot ( C ) were used to measure the expression level of PCNA, Bcl-2, Bax and cleaved caspase3 in shUSP4- or NC-transduced U87 and T98G cells. *** P
Figure Legend Snippet: Silencing of USP4 promoted cell apoptosis in GBM cell lines. Notes: ( A ) Cell apoptosis was detected by flow cytometry using annexin-V/PI kit in shUSP4- or NC-transduced U87 and T98G cells. ( B , C ) Real-time PCR ( B ) and Western blot ( C ) were used to measure the expression level of PCNA, Bcl-2, Bax and cleaved caspase3 in shUSP4- or NC-transduced U87 and T98G cells. *** P

Techniques Used: Flow Cytometry, Cytometry, Real-time Polymerase Chain Reaction, Western Blot, Expressing

73) Product Images from "Vorinostat and quinacrine have synergistic effects in T-cell acute lymphoblastic leukemia through reactive oxygen species increase and mitophagy inhibition"

Article Title: Vorinostat and quinacrine have synergistic effects in T-cell acute lymphoblastic leukemia through reactive oxygen species increase and mitophagy inhibition

Journal: Cell Death & Disease

doi: 10.1038/s41419-018-0679-6

Effects of vorinostat and QC on the apoptosis of Jurkat and Molt-4 cells. Jurkat cells were treated with vorinostat (1 μM) and/or QC (5 μM) for 48 h and Molt-4 cells were treated with vorinostat (1 μM) and/or QC (2.5 μM) for 24 h. a , b Cell cycle distribution of Jurkat and Molt-4 cells was analyzed by flow cytometry. c , d Apoptosis rate of Jurkat and Molt-4 cells was established by flow cytometry analysis of Annexin-V-PI dual staining. e The indicated proteins were examined by western blots. * p
Figure Legend Snippet: Effects of vorinostat and QC on the apoptosis of Jurkat and Molt-4 cells. Jurkat cells were treated with vorinostat (1 μM) and/or QC (5 μM) for 48 h and Molt-4 cells were treated with vorinostat (1 μM) and/or QC (2.5 μM) for 24 h. a , b Cell cycle distribution of Jurkat and Molt-4 cells was analyzed by flow cytometry. c , d Apoptosis rate of Jurkat and Molt-4 cells was established by flow cytometry analysis of Annexin-V-PI dual staining. e The indicated proteins were examined by western blots. * p

Techniques Used: Flow Cytometry, Cytometry, Staining, Western Blot

74) Product Images from "Knockdown delta-5-desaturase in breast cancer cells that overexpress COX-2 results in inhibition of growth, migration and invasion via a dihomo-γ-linolenic acid peroxidation dependent mechanism"

Article Title: Knockdown delta-5-desaturase in breast cancer cells that overexpress COX-2 results in inhibition of growth, migration and invasion via a dihomo-γ-linolenic acid peroxidation dependent mechanism

Journal: BMC Cancer

doi: 10.1186/s12885-018-4250-8

D5D- KD and DGLA supplementation enhance the efficacy of 5-FU on growth of 4 T1 cells. a Colony formation assay of D5D- KD 4 T1 cells or control siRNA transfected cells at 10 days with treatment of 5-FU (20 μM) or 5-FU + DGLA (100 μM) for 48 h. Note, D5D- KD and NC-si control cells without DGLA and 5-FU treatment (shown in Fig. 3 ) were used to calculate survival fractions; and b Cell apoptosis was examined via flow cytometry after D5D- KD 4 T1 cells were treated with DGLA (100 μM), 5-FU (20 μM), or 5-FU + DGLA for 48 h, followed by Annexin V-FITC/PI double staining Data represent as mean ± standard deviation (*: significant difference vs. control with p
Figure Legend Snippet: D5D- KD and DGLA supplementation enhance the efficacy of 5-FU on growth of 4 T1 cells. a Colony formation assay of D5D- KD 4 T1 cells or control siRNA transfected cells at 10 days with treatment of 5-FU (20 μM) or 5-FU + DGLA (100 μM) for 48 h. Note, D5D- KD and NC-si control cells without DGLA and 5-FU treatment (shown in Fig. 3 ) were used to calculate survival fractions; and b Cell apoptosis was examined via flow cytometry after D5D- KD 4 T1 cells were treated with DGLA (100 μM), 5-FU (20 μM), or 5-FU + DGLA for 48 h, followed by Annexin V-FITC/PI double staining Data represent as mean ± standard deviation (*: significant difference vs. control with p

Techniques Used: Colony Assay, Transfection, Flow Cytometry, Cytometry, Double Staining, Standard Deviation

D5D- KD and DGLA supplementation enhance the efficacy of 5-FU on growth of MDA-MB 231 cells. a Colony formation assay of D5D- KD MDA-MB 231 cells or control siRNA transfected cells at 10 days with treatment of 5-FU (10 μM) or 5-FU + DGLA (100 μM) for 48 h. Note, D5D- KD and NC-si control cells without DGLA and 5-FU treatment (shown in Fig. 2 ) were used to calculate survival fractions; and b Cell apoptosis was examined via flow cytometry after D5D- KD MDA-MB 231 cells were treated with DGLA (100 μM), 5-FU (10 μM), or 5-FU + DGLA for 48 h, followed by Annexin V-FITC/PI double staining. Data represent as mean ± standard deviation (*: significant difference vs. control with p
Figure Legend Snippet: D5D- KD and DGLA supplementation enhance the efficacy of 5-FU on growth of MDA-MB 231 cells. a Colony formation assay of D5D- KD MDA-MB 231 cells or control siRNA transfected cells at 10 days with treatment of 5-FU (10 μM) or 5-FU + DGLA (100 μM) for 48 h. Note, D5D- KD and NC-si control cells without DGLA and 5-FU treatment (shown in Fig. 2 ) were used to calculate survival fractions; and b Cell apoptosis was examined via flow cytometry after D5D- KD MDA-MB 231 cells were treated with DGLA (100 μM), 5-FU (10 μM), or 5-FU + DGLA for 48 h, followed by Annexin V-FITC/PI double staining. Data represent as mean ± standard deviation (*: significant difference vs. control with p

Techniques Used: Multiple Displacement Amplification, Colony Assay, Transfection, Flow Cytometry, Cytometry, Double Staining, Standard Deviation

75) Product Images from "Human Immunodeficiency Virus Type 1 Inhibits DNA Damage-Triggered Apoptosis by a Nef-Independent Mechanism"

Article Title: Human Immunodeficiency Virus Type 1 Inhibits DNA Damage-Triggered Apoptosis by a Nef-Independent Mechanism

Journal: Journal of Virology

doi: 10.1128/JVI.79.9.5489-5498.2005

Quantitation of apoptosis using bicistronic vectors coexpressing Nef and GFP. Flow cytometric analysis of uninduced (A to C) or 20-s UV light-treated (D and E) Jurkat cells transfected with constructs expressing GFP alone (B and D) or together with the NL4-3 Nef (C and E). Live cells were selected by forward and side scattering (panel 1). GFP expression allowed us to discriminate between transfected and untransfected cells (panel 2). GFP-positive cells and mock-transfected cells were evaluated for annexin V binding (panel 3), and triple staining for 7-AAD allowed us to distinguish between early and late apoptotic cells (panel 4). Numbers are the percentages of annexin V- and 7-AAD-positive cells. Similar results were obtained in three independent experiments.
Figure Legend Snippet: Quantitation of apoptosis using bicistronic vectors coexpressing Nef and GFP. Flow cytometric analysis of uninduced (A to C) or 20-s UV light-treated (D and E) Jurkat cells transfected with constructs expressing GFP alone (B and D) or together with the NL4-3 Nef (C and E). Live cells were selected by forward and side scattering (panel 1). GFP expression allowed us to discriminate between transfected and untransfected cells (panel 2). GFP-positive cells and mock-transfected cells were evaluated for annexin V binding (panel 3), and triple staining for 7-AAD allowed us to distinguish between early and late apoptotic cells (panel 4). Numbers are the percentages of annexin V- and 7-AAD-positive cells. Similar results were obtained in three independent experiments.

Techniques Used: Quantitation Assay, Flow Cytometry, Transfection, Construct, Expressing, Binding Assay, Staining

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Article Snippet: .. Assessment of apoptosis Apoptosis in cells was assessed using an Annexin V Apoptosis Detection Kit (BD Pharmingen, San Jose, CA, USA) analyzed by using a BD Accuri C6 flow cytometer according to the manufacturer's instructions. .. Cell viability, caspase assays, and crystal violet cell cytotoxicity assay Cell viability in cells was measured in cells cultured in 96-well plates using Prestoblue Cell Viability reagent (Life technologies, Grand Island, NY, USA) according to the manufacturer's protocol.

Article Title: The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia
Article Snippet: Paragraph title: Flow cytometry and cell sorting ... Cell cycle analysis and apoptosis analysis were performed using the BrdU Flow Kit (BD Biosciences) and the Annexin V Apoptosis Detection Kit (BD Biosciences), respectively, according to the manufacturer’s instructions.

Article Title: LINC00365-SCGB2A1 axis inhibits the viability of breast cancer through targeting NF-κB signaling
Article Snippet: Each group of cells was stained with Annexin-V FITC/propidium iodide (Annexin V Apoptosis Detection Kit II; BD Biosciences). .. Samples were analyzed using a BD Accuri C6 flow cytometer (Becton, Dickinson and Company).

Article Title: Plk2 Regulated by miR-128 Induces Ischemia-Reperfusion Injury in Cardiac Cells
Article Snippet: Paragraph title: Apoptosis Analysis by Flow Cytometry ... Cells were double stained with fluorescein isothiocyanate (FITC)-labeled Annexin V and PI using the Annexin V Apoptosis Detection Kit (BD Biosciences, Mississauga, ON, Canada) according to the manufacturer’s instructions.

Article Title: Molecular Mechanisms of Action of Herbal Antifungal Alkaloid Berberine, in Candida albicans
Article Snippet: .. Annexin V (Annexin V Apoptosis Detection Kit I, BD Biosciences) binds to PS which is in turn linked to FITC hence can be detected through Flow cytometry as explained in . .. However, there was no significant change in the population of cells showing FITC labeling on treatment with BER at MIC80 as seen in .

Article Title: Dual Specificity Phosphatase 5 Is Essential for T Cell Survival
Article Snippet: Samples were run on a BD Accuri C6 cytometer and analyzed using FCS Express. .. For apoptosis data, 1x105 cells were stained using the Annexin V Apoptosis Detection Kit (Product 556547, BD Pharmingen, San Diego CA) according to protocol.

Article Title: Immune-mediated bone marrow failure in C57BL/6 mice
Article Snippet: Paragraph title: Blood counts and flow cytometry ... To measure cell apoptosis, cells were first stained with an antibody mixture along with Annexin V in specific high calcium buffer using reagents from an Annexin V apoptosis detection kit from BD Biosciences, and were then added with 7AAD 10 minutes before cell acquisition.

Article Title: Downregulation of A20 increases the cytotoxicity of IFN-γ in hepatocellular carcinoma cells
Article Snippet: Apoptosis analysis Cellular apoptosis was measured by flow cytometry. .. The cell pellets were resuspended in 100 μL of binding buffer and stained with Annexin V apoptosis detection kit (BD Biosciences) according to the manufacturer’s guide.

Article Title: Small interfering RNA targeting S100A4 sensitizes non-small-cell lung cancer cells (A549) to radiation treatment
Article Snippet: The experiments were performed with an annexin V apoptosis detection kit (BD Biosciences). .. Then, the samples were analyzed by flow cytometry on a FACScan (Becton Dickinson, Mountain View, CA, USA) using the Cell Quest program.

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma
Article Snippet: Paragraph title: Flow cytometry ... Flow cytometric analysis of apoptotic cells was performed by staining the cells with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD) using the Annexin V Apoptosis Detection Kit (BD Pharmingen) for 15 min according to the manufacturer's protocol.

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit. .. With Annexin-V (AnV)/propidium iodide (PI) staining, GIMAP6 displayed an apparent anti-apoptotic effect, as indicated by the difference between the control and knockdown lines.

CTL Assay:

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: Various Jurkat-derived cell lines (Ctl, KD1, and KD2) were treated with 100 μ m H2 O2 for 4, 8, 24, and 30 h to induce cell apoptosis/death ( , B and C ). .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit.

Incubation:

Article Title: Immune-mediated bone marrow failure in C57BL/6 mice
Article Snippet: Peripheral blood leukocytes and BM cells were first incubated with ACK buffer twice for 10 minutes in order to lyse red blood cells (RBCs). .. To measure cell apoptosis, cells were first stained with an antibody mixture along with Annexin V in specific high calcium buffer using reagents from an Annexin V apoptosis detection kit from BD Biosciences, and were then added with 7AAD 10 minutes before cell acquisition.

Article Title: Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells
Article Snippet: Coculture was conducted in MLR media [DMEM plus 5% FBS with 10 mM HEPES (pH 7.4), 1% sodium pyruvate, 1% penicillin/streptomycin, 1% l -glutamine, 0.4% l -arginine HCl, 1% folic acid/l -asparagine, and 0.2% 2-ME] supplemented with recombinant Murine IFN-γ (20 ng/mL, Peprotech, Rocky Hill, NJ) and the specific killing of target cells was tested by using Alamar Blue (Invitrogen, Waltham, MA) which is added at 10% of the sample volume followed by 4–16 h incubation. .. In another experimental setup, Annexin V Apoptosis Detection Kit I was used for viability staining according to the manufacturer instructions (BD Pharmingen) and culture supernatant was collected for NO measurement.

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma
Article Snippet: Flow cytometric analysis of apoptotic cells was performed by staining the cells with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD) using the Annexin V Apoptosis Detection Kit (BD Pharmingen) for 15 min according to the manufacturer's protocol. .. The cells were then washed twice in PBS, stained with propidium iodide (50 μg/ml) in the presence of 50 μg/ml RNase A (Sigma-Aldrich), and incubated for 1 h at room temperature.

Cell Culture:

Article Title: Moscatilin Inhibits Lung Cancer Cell Motility and Invasion via Suppression of Endogenous Reactive Oxygen Species
Article Snippet: Cells and Reagents Human lung adenocarcinoma H23 cells were obtained from American Type Culture Collection (ATCC, Manassas, VA, USA), and cultured in RPMI-1640 medium containing 10% fetal bovine serum, 2 mM L-glutamine, 100 IU/mL penicillin, and 100 μ g/mL streptomycin (Gibco, MD, USA) in 37°C with 5% CO2 -humidified incubator. .. Annexin V Apoptosis Detection Kit was obtained from BD Biosciences (Woburn, MA, USA).

Article Title: Dual Specificity Phosphatase 5 Is Essential for T Cell Survival
Article Snippet: Proliferation and Apoptosis Assays T cells were cultured as described above. .. For apoptosis data, 1x105 cells were stained using the Annexin V Apoptosis Detection Kit (Product 556547, BD Pharmingen, San Diego CA) according to protocol.

Expressing:

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: After single colony selection, the relative -fold of GIMAP6 expression was determined by immunoblot analysis ( A ). .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit.

Derivative Assay:

Article Title: The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia
Article Snippet: Cell cycle analysis and apoptosis analysis were performed using the BrdU Flow Kit (BD Biosciences) and the Annexin V Apoptosis Detection Kit (BD Biosciences), respectively, according to the manufacturer’s instructions. .. Single-cell clones were derived by directly sorting transduced cells into 96-well plates.

Flow Cytometry:

Article Title: Ubiquitin-like (UBX)-domain-containing protein, UBXN2A, promotes cell death by interfering with the p53-Mortalin interactions in colon cancer cells
Article Snippet: .. Assessment of apoptosis Apoptosis in cells was assessed using an Annexin V Apoptosis Detection Kit (BD Pharmingen, San Jose, CA, USA) analyzed by using a BD Accuri C6 flow cytometer according to the manufacturer's instructions. .. Cell viability, caspase assays, and crystal violet cell cytotoxicity assay Cell viability in cells was measured in cells cultured in 96-well plates using Prestoblue Cell Viability reagent (Life technologies, Grand Island, NY, USA) according to the manufacturer's protocol.

Article Title: The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia
Article Snippet: .. Cell cycle analysis and apoptosis analysis were performed using the BrdU Flow Kit (BD Biosciences) and the Annexin V Apoptosis Detection Kit (BD Biosciences), respectively, according to the manufacturer’s instructions. .. Single-cell clones were derived by directly sorting transduced cells into 96-well plates.

Article Title: LINC00365-SCGB2A1 axis inhibits the viability of breast cancer through targeting NF-κB signaling
Article Snippet: Paragraph title: Flow cytometric analysis ... Each group of cells was stained with Annexin-V FITC/propidium iodide (Annexin V Apoptosis Detection Kit II; BD Biosciences).

Article Title: Plk2 Regulated by miR-128 Induces Ischemia-Reperfusion Injury in Cardiac Cells
Article Snippet: Paragraph title: Apoptosis Analysis by Flow Cytometry ... Cells were double stained with fluorescein isothiocyanate (FITC)-labeled Annexin V and PI using the Annexin V Apoptosis Detection Kit (BD Biosciences, Mississauga, ON, Canada) according to the manufacturer’s instructions.

Article Title: Expression and Regulation of Prostate Apoptosis Response-4 (Par-4) in Human Glioma Stem Cells in Drug-Induced Apoptosis
Article Snippet: Annexin V and PI staining HNGC-2 cells were treated with tamoxifen (10µg/ml) for 12 h and apoptosis was determined using Annexin-V apoptosis detection kit (BD Pharmingen) according to the manufacturer's protocol. .. Annexin V and PI staining HNGC-2 cells were treated with tamoxifen (10µg/ml) for 12 h and apoptosis was determined using Annexin-V apoptosis detection kit (BD Pharmingen) according to the manufacturer's protocol.

Article Title: Molecular Mechanisms of Action of Herbal Antifungal Alkaloid Berberine, in Candida albicans
Article Snippet: .. Annexin V (Annexin V Apoptosis Detection Kit I, BD Biosciences) binds to PS which is in turn linked to FITC hence can be detected through Flow cytometry as explained in . .. However, there was no significant change in the population of cells showing FITC labeling on treatment with BER at MIC80 as seen in .

Article Title: Dual Specificity Phosphatase 5 Is Essential for T Cell Survival
Article Snippet: For apoptosis data, 1x105 cells were stained using the Annexin V Apoptosis Detection Kit (Product 556547, BD Pharmingen, San Diego CA) according to protocol. .. Samples were run on an LSRII flow cytometer and analyzed using FlowJo (FlowJo LLC, Ashland, OR).

Article Title: Immune-mediated bone marrow failure in C57BL/6 mice
Article Snippet: Paragraph title: Blood counts and flow cytometry ... To measure cell apoptosis, cells were first stained with an antibody mixture along with Annexin V in specific high calcium buffer using reagents from an Annexin V apoptosis detection kit from BD Biosciences, and were then added with 7AAD 10 minutes before cell acquisition.

Article Title: Downregulation of A20 increases the cytotoxicity of IFN-γ in hepatocellular carcinoma cells
Article Snippet: Apoptosis analysis Cellular apoptosis was measured by flow cytometry. .. The cell pellets were resuspended in 100 μL of binding buffer and stained with Annexin V apoptosis detection kit (BD Biosciences) according to the manufacturer’s guide.

Article Title: Small interfering RNA targeting S100A4 sensitizes non-small-cell lung cancer cells (A549) to radiation treatment
Article Snippet: The experiments were performed with an annexin V apoptosis detection kit (BD Biosciences). .. Then, the samples were analyzed by flow cytometry on a FACScan (Becton Dickinson, Mountain View, CA, USA) using the Cell Quest program.

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma
Article Snippet: .. Flow cytometric analysis of apoptotic cells was performed by staining the cells with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD) using the Annexin V Apoptosis Detection Kit (BD Pharmingen) for 15 min according to the manufacturer's protocol. .. For cell-cycle analysis, infected cells were washed in phosphate-buffered saline (PBS) and fixed in 70% ice-cold ethanol for 10 min at 4°C.

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit. .. With Annexin-V (AnV)/propidium iodide (PI) staining, GIMAP6 displayed an apparent anti-apoptotic effect, as indicated by the difference between the control and knockdown lines.

Infection:

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma
Article Snippet: Flow cytometry To analyze apoptosis, cells were harvested 48 h after infection. .. Flow cytometric analysis of apoptotic cells was performed by staining the cells with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD) using the Annexin V Apoptosis Detection Kit (BD Pharmingen) for 15 min according to the manufacturer's protocol.

Generated:

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: Stable knockdown cells (KD1 and KD2) were generated using anti-GIMAP6 shRNA, whereas control cells (Ctl) were prepared using the pLKO.1 empty vector plasmid. .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit.

Recombinant:

Article Title: Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells
Article Snippet: Coculture was conducted in MLR media [DMEM plus 5% FBS with 10 mM HEPES (pH 7.4), 1% sodium pyruvate, 1% penicillin/streptomycin, 1% l -glutamine, 0.4% l -arginine HCl, 1% folic acid/l -asparagine, and 0.2% 2-ME] supplemented with recombinant Murine IFN-γ (20 ng/mL, Peprotech, Rocky Hill, NJ) and the specific killing of target cells was tested by using Alamar Blue (Invitrogen, Waltham, MA) which is added at 10% of the sample volume followed by 4–16 h incubation. .. In another experimental setup, Annexin V Apoptosis Detection Kit I was used for viability staining according to the manufacturer instructions (BD Pharmingen) and culture supernatant was collected for NO measurement.

MTT Assay:

Article Title: Moscatilin Inhibits Lung Cancer Cell Motility and Invasion via Suppression of Endogenous Reactive Oxygen Species
Article Snippet: The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), Hoechst33342, propidium iodide (PI), phalloidin tetramethylrhodamine B isothiocyanate, ribonuclease A, bovine serum albumin (BSA), and dimethylsulfoxide (DMSO) were purchased from Sigma Chemical, Inc. (St. Louis, MO, USA). .. Annexin V Apoptosis Detection Kit was obtained from BD Biosciences (Woburn, MA, USA).

Fluorescence:

Article Title: The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia
Article Snippet: Flow cytometry analyses of the transduced HSPCs, cell lines, and patient blasts were performed on a fluorescence-activated cell sorter (FACS) Canto flow cytometer (BD Biosciences) or a CytoFLEX B5-R3-V5 (Beckman Coulter). .. Cell cycle analysis and apoptosis analysis were performed using the BrdU Flow Kit (BD Biosciences) and the Annexin V Apoptosis Detection Kit (BD Biosciences), respectively, according to the manufacturer’s instructions.

Article Title: Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells
Article Snippet: The percentage of lysis was calculated using the formula; %Lysis = 100 × {[(AF of targets alone)] − [(AF of mix) − (AF of effectors alone)]}/{AF of targets alone} where AF is the mean fluorescence for the triplicate wells after the mean fluorescence of the wells containing medium alone was subtracted. .. In another experimental setup, Annexin V Apoptosis Detection Kit I was used for viability staining according to the manufacturer instructions (BD Pharmingen) and culture supernatant was collected for NO measurement.

Article Title: Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS
Article Snippet: For quantification of apoptosis, cells were harvested, washed twice with ice-cold PBS and stained in calcium-binding buffer with APC-conjugated Annexin V and 7-Aminoactinomycin D (7-AAD) using Annexin V apoptosis detection kit (BD Pharmingen, San Diego, CA), according to manufacturer’s recommendations. .. Samples were analyzed within 1 h. The stained cells were analyzed for both cell cycle and apoptosis by fluorescence-activated cell sorting using a FACSCantoII equipped with a FACSDiva 6.1 CellQuest software (Becton Dickinson Instrument, San Josè, CA).

Isolation:

Article Title: Moscatilin Inhibits Lung Cancer Cell Motility and Invasion via Suppression of Endogenous Reactive Oxygen Species
Article Snippet: Moscatilin was isolated from Thai orchid Dendrobium pulchellum as previously described [ ]. .. Annexin V Apoptosis Detection Kit was obtained from BD Biosciences (Woburn, MA, USA).

Transfection:

Article Title: LINC00365-SCGB2A1 axis inhibits the viability of breast cancer through targeting NF-κB signaling
Article Snippet: Flow cytometric analysis MCF-7 and T47D breast cancer cells were seeded in 6-well plates and transfected with empty, LINC00365- or SCGB2A1-overexpressing vectors for 48 h. Attaching cells and cells in the supernatant were collected. .. Each group of cells was stained with Annexin-V FITC/propidium iodide (Annexin V Apoptosis Detection Kit II; BD Biosciences).

Article Title: Plk2 Regulated by miR-128 Induces Ischemia-Reperfusion Injury in Cardiac Cells
Article Snippet: Apoptosis Analysis by Flow Cytometry H9c2 cells were plated in six-well plates and then subjected to transfection and simulated cold I/R injury as described above. .. Cells were double stained with fluorescein isothiocyanate (FITC)-labeled Annexin V and PI using the Annexin V Apoptosis Detection Kit (BD Biosciences, Mississauga, ON, Canada) according to the manufacturer’s instructions.

Article Title: Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS
Article Snippet: Cell cycle and apoptosis assays Cells were transfected 24 h after seeding (Day 0) with siRNAs and after 24 h transfected again. .. For quantification of apoptosis, cells were harvested, washed twice with ice-cold PBS and stained in calcium-binding buffer with APC-conjugated Annexin V and 7-Aminoactinomycin D (7-AAD) using Annexin V apoptosis detection kit (BD Pharmingen, San Diego, CA), according to manufacturer’s recommendations.

Labeling:

Article Title: Molecular Mechanisms of Action of Herbal Antifungal Alkaloid Berberine, in Candida albicans
Article Snippet: Annexin V (Annexin V Apoptosis Detection Kit I, BD Biosciences) binds to PS which is in turn linked to FITC hence can be detected through Flow cytometry as explained in . .. However, there was no significant change in the population of cells showing FITC labeling on treatment with BER at MIC80 as seen in .

Selection:

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: After single colony selection, the relative -fold of GIMAP6 expression was determined by immunoblot analysis ( A ). .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit.

Staining:

Article Title: The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia
Article Snippet: Staining and measurement were performed according to standard protocols as previously described for human cells using the antibodies CD163-PE (BD Biosciences), CD11b-PeCy7 (Beckman Coulter), CD14-APC (Beckman Coulter), CD45-V500 (BD Biosciences), CD163-APC-Cy7 (BioLegend), CD45-APC (BD Biosciences), CD15-BV605 (BD Biosciences), and CD66b-PE (BD Biosciences). .. Cell cycle analysis and apoptosis analysis were performed using the BrdU Flow Kit (BD Biosciences) and the Annexin V Apoptosis Detection Kit (BD Biosciences), respectively, according to the manufacturer’s instructions.

Article Title: LINC00365-SCGB2A1 axis inhibits the viability of breast cancer through targeting NF-κB signaling
Article Snippet: .. Each group of cells was stained with Annexin-V FITC/propidium iodide (Annexin V Apoptosis Detection Kit II; BD Biosciences). .. Samples were analyzed using a BD Accuri C6 flow cytometer (Becton, Dickinson and Company).

Article Title: Plk2 Regulated by miR-128 Induces Ischemia-Reperfusion Injury in Cardiac Cells
Article Snippet: .. Cells were double stained with fluorescein isothiocyanate (FITC)-labeled Annexin V and PI using the Annexin V Apoptosis Detection Kit (BD Biosciences, Mississauga, ON, Canada) according to the manufacturer’s instructions. .. Cells were then analyzed by flow cytometry using a CytoFLEX system (Beckman Coulter, Mississauga, ON, Canada) and the CytExpert software.

Article Title: Dual Specificity Phosphatase 5 Is Essential for T Cell Survival
Article Snippet: .. For apoptosis data, 1x105 cells were stained using the Annexin V Apoptosis Detection Kit (Product 556547, BD Pharmingen, San Diego CA) according to protocol. .. Samples were run on an LSRII flow cytometer and analyzed using FlowJo (FlowJo LLC, Ashland, OR).

Article Title: Immune-mediated bone marrow failure in C57BL/6 mice
Article Snippet: .. To measure cell apoptosis, cells were first stained with an antibody mixture along with Annexin V in specific high calcium buffer using reagents from an Annexin V apoptosis detection kit from BD Biosciences, and were then added with 7AAD 10 minutes before cell acquisition. .. Monoclonal antibodies for murine CD3 (clone 145-2C11), CD4 (clone GK 1.5), CD8 (clone 53-6.72), CD11a (clone 2D7), CD11b (clone M1/70), CD95 (Fas, clone Jo2), CD117 (c-Kit, clone 2B8), CD178.1 (FasL, clone MFL3), erythroid cells (clone Ter119), granulocytes (Gr1/Ly6-G, clone RB6-8C5), and stem cell antigen 1 (Sca-1, clone E13-161) were from BD-Biosciences (San Diego, CA).

Article Title: Downregulation of A20 increases the cytotoxicity of IFN-γ in hepatocellular carcinoma cells
Article Snippet: .. The cell pellets were resuspended in 100 μL of binding buffer and stained with Annexin V apoptosis detection kit (BD Biosciences) according to the manufacturer’s guide. .. Then the samples were analyzed by flow cytometry (Becton Dickinson).

Article Title: Small interfering RNA targeting S100A4 sensitizes non-small-cell lung cancer cells (A549) to radiation treatment
Article Snippet: The experiments were performed with an annexin V apoptosis detection kit (BD Biosciences). .. The cell pellets were resuspended in 100 mL of binding buffer and stained with 5 mL each of annexin V and PI staining solution for 15 minutes.

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma
Article Snippet: .. Flow cytometric analysis of apoptotic cells was performed by staining the cells with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD) using the Annexin V Apoptosis Detection Kit (BD Pharmingen) for 15 min according to the manufacturer's protocol. .. For cell-cycle analysis, infected cells were washed in phosphate-buffered saline (PBS) and fixed in 70% ice-cold ethanol for 10 min at 4°C.

Article Title: Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS
Article Snippet: .. For quantification of apoptosis, cells were harvested, washed twice with ice-cold PBS and stained in calcium-binding buffer with APC-conjugated Annexin V and 7-Aminoactinomycin D (7-AAD) using Annexin V apoptosis detection kit (BD Pharmingen, San Diego, CA), according to manufacturer’s recommendations. .. Samples were analyzed within 1 h. The stained cells were analyzed for both cell cycle and apoptosis by fluorescence-activated cell sorting using a FACSCantoII equipped with a FACSDiva 6.1 CellQuest software (Becton Dickinson Instrument, San Josè, CA).

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit. .. With Annexin-V (AnV)/propidium iodide (PI) staining, GIMAP6 displayed an apparent anti-apoptotic effect, as indicated by the difference between the control and knockdown lines.

Plasmid Preparation:

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: Stable knockdown cells (KD1 and KD2) were generated using anti-GIMAP6 shRNA, whereas control cells (Ctl) were prepared using the pLKO.1 empty vector plasmid. .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit.

Software:

Article Title: Plk2 Regulated by miR-128 Induces Ischemia-Reperfusion Injury in Cardiac Cells
Article Snippet: Cells were double stained with fluorescein isothiocyanate (FITC)-labeled Annexin V and PI using the Annexin V Apoptosis Detection Kit (BD Biosciences, Mississauga, ON, Canada) according to the manufacturer’s instructions. .. Cells were then analyzed by flow cytometry using a CytoFLEX system (Beckman Coulter, Mississauga, ON, Canada) and the CytExpert software.

Article Title: Expression and Regulation of Prostate Apoptosis Response-4 (Par-4) in Human Glioma Stem Cells in Drug-Induced Apoptosis
Article Snippet: Annexin V and PI staining HNGC-2 cells were treated with tamoxifen (10µg/ml) for 12 h and apoptosis was determined using Annexin-V apoptosis detection kit (BD Pharmingen) according to the manufacturer's protocol. .. Annexin V and PI staining HNGC-2 cells were treated with tamoxifen (10µg/ml) for 12 h and apoptosis was determined using Annexin-V apoptosis detection kit (BD Pharmingen) according to the manufacturer's protocol.

Article Title: Immune-mediated bone marrow failure in C57BL/6 mice
Article Snippet: Residual leukocytes were stained with various antibodies and analyzed on a LSR II or Canto II flow cytometer using the FACSDiva software (Becton Dickson, San Diego CA). .. To measure cell apoptosis, cells were first stained with an antibody mixture along with Annexin V in specific high calcium buffer using reagents from an Annexin V apoptosis detection kit from BD Biosciences, and were then added with 7AAD 10 minutes before cell acquisition.

Article Title: Axon guidance molecule semaphorin3A is a novel tumor suppressor in head and neck squamous cell carcinoma
Article Snippet: Flow cytometric analysis of apoptotic cells was performed by staining the cells with Annexin V-allophycocyanin (APC) and 7-aminoactinomycin D (7-AAD) using the Annexin V Apoptosis Detection Kit (BD Pharmingen) for 15 min according to the manufacturer's protocol. .. The detection of the proportion of apoptotic cells and cell-cycle analysis were performed using a FACSCalibur flow cytometer (BD Biosciences) and CellQuest Pro software (BD Biosciences).

Article Title: Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS
Article Snippet: For quantification of apoptosis, cells were harvested, washed twice with ice-cold PBS and stained in calcium-binding buffer with APC-conjugated Annexin V and 7-Aminoactinomycin D (7-AAD) using Annexin V apoptosis detection kit (BD Pharmingen, San Diego, CA), according to manufacturer’s recommendations. .. Samples were analyzed within 1 h. The stained cells were analyzed for both cell cycle and apoptosis by fluorescence-activated cell sorting using a FACSCantoII equipped with a FACSDiva 6.1 CellQuest software (Becton Dickinson Instrument, San Josè, CA).

shRNA:

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: Stable knockdown cells (KD1 and KD2) were generated using anti-GIMAP6 shRNA, whereas control cells (Ctl) were prepared using the pLKO.1 empty vector plasmid. .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit.

In Vitro:

Article Title: Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells
Article Snippet: Paragraph title: In vitro culture experiments ... In another experimental setup, Annexin V Apoptosis Detection Kit I was used for viability staining according to the manufacturer instructions (BD Pharmingen) and culture supernatant was collected for NO measurement.

Apoptosis Assay:

Article Title: Small interfering RNA targeting S100A4 sensitizes non-small-cell lung cancer cells (A549) to radiation treatment
Article Snippet: Paragraph title: Apoptosis assay ... The experiments were performed with an annexin V apoptosis detection kit (BD Biosciences).

Article Title: Functional and biochemical characterization of a T cell-associated anti-apoptotic protein, GIMAP6
Article Snippet: To reveal the extent to which cell apoptosis/death was delayed by GIMAP6, an H2 O2 -mediated, multiple time point apoptosis assay was performed. .. The apoptosis signal, phosphatidylserine exposure, was then measured by flow cytometry using a BD Annexin-V apoptosis detection kit.

Concentration Assay:

Article Title: Molecular Mechanisms of Action of Herbal Antifungal Alkaloid Berberine, in Candida albicans
Article Snippet: Wild type cells treated with a MIC50 concentration of BER (100 µg/ml) were allowed to incubate in presence of 10 µM of CM-H2 DCFDA. .. Annexin V (Annexin V Apoptosis Detection Kit I, BD Biosciences) binds to PS which is in turn linked to FITC hence can be detected through Flow cytometry as explained in .

Lysis:

Article Title: Colonization of xenograft tumors by oncolytic vaccinia virus (VACV) results in enhanced tumor killing due to the involvement of myeloid cells
Article Snippet: The percentage of lysis was calculated using the formula; %Lysis = 100 × {[(AF of targets alone)] − [(AF of mix) − (AF of effectors alone)]}/{AF of targets alone} where AF is the mean fluorescence for the triplicate wells after the mean fluorescence of the wells containing medium alone was subtracted. .. In another experimental setup, Annexin V Apoptosis Detection Kit I was used for viability staining according to the manufacturer instructions (BD Pharmingen) and culture supernatant was collected for NO measurement.

Marker:

Article Title: Molecular Mechanisms of Action of Herbal Antifungal Alkaloid Berberine, in Candida albicans
Article Snippet: To estimate whether generation of ROS leads to apoptosis on BER treatment, we checked for externalization of phosphatidyl serine (PS), a marker for apoptotic cells. .. Annexin V (Annexin V Apoptosis Detection Kit I, BD Biosciences) binds to PS which is in turn linked to FITC hence can be detected through Flow cytometry as explained in .

FACS:

Article Title: The stem cell–specific long noncoding RNA HOXA10-AS in the pathogenesis of KMT2A-rearranged leukemia
Article Snippet: Paragraph title: Flow cytometry and cell sorting ... Cell cycle analysis and apoptosis analysis were performed using the BrdU Flow Kit (BD Biosciences) and the Annexin V Apoptosis Detection Kit (BD Biosciences), respectively, according to the manufacturer’s instructions.

Article Title: Expression and Regulation of Prostate Apoptosis Response-4 (Par-4) in Human Glioma Stem Cells in Drug-Induced Apoptosis
Article Snippet: Annexin V and PI staining HNGC-2 cells were treated with tamoxifen (10µg/ml) for 12 h and apoptosis was determined using Annexin-V apoptosis detection kit (BD Pharmingen) according to the manufacturer's protocol. .. Annexin V and PI staining HNGC-2 cells were treated with tamoxifen (10µg/ml) for 12 h and apoptosis was determined using Annexin-V apoptosis detection kit (BD Pharmingen) according to the manufacturer's protocol.

Article Title: Pharmacological inhibition of EZH2 as a promising differentiation therapy in embryonal RMS
Article Snippet: For quantification of apoptosis, cells were harvested, washed twice with ice-cold PBS and stained in calcium-binding buffer with APC-conjugated Annexin V and 7-Aminoactinomycin D (7-AAD) using Annexin V apoptosis detection kit (BD Pharmingen, San Diego, CA), according to manufacturer’s recommendations. .. Samples were analyzed within 1 h. The stained cells were analyzed for both cell cycle and apoptosis by fluorescence-activated cell sorting using a FACSCantoII equipped with a FACSDiva 6.1 CellQuest software (Becton Dickinson Instrument, San Josè, CA).

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    Becton Dickinson annexin v pi apoptosis detecting kit
    Synthetic peptide N-DEVGEANN-C successfully inhibited Nef from interacting with and there by restore the functions of endogenous wild type ASK1 activity. ( A ) HEK-293 cells were transfected with Nef plasmid or vector backbone. 24 hrs post transfection, cells were stimulated to progress through ASK1 mediated apoptotic progression using classic ligand viz. TNF-α (100ng/ml) either in absence or presence of synthetic peptide N-DEVGEANN-C (10ng/ml). The extent of interaction of endogenous wild type ASK1 with Nef and ability of peptide N-DEVGEANN-C to impair the interaction was evaluated using In situ proximity ligation assay. ( B ) Synthetic peptide N-DEVGEANN-C prevented HIV-1Nef from offering protection against TNF -α induced, endogenous wild type ASK1 mediated <t>apoptosis</t> as evaluated using <t>Annexin-V/PI</t> assay. ( C ) Endogenous wild type ASK1 activity was rendered active by synthetic peptide N-DEVGEANN-C even in presence of HIV-1Nef as indicated by markedly decreased inhibitory Ser 967 phosphorylation of ASK1 with a corresponding increase in p54/46JNK1 activation profile as indicated by higher phospho-p54/46JNK1 levels in western blot analysis. All western blotting was done three times and integrated density was determined by densitometry analysis using imageJ software.
    Annexin V Pi Apoptosis Detecting Kit, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 78/100, based on 23 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/annexin v pi apoptosis detecting kit/product/Becton Dickinson
    Average 78 stars, based on 23 article reviews
    Price from $9.99 to $1999.99
    annexin v pi apoptosis detecting kit - by Bioz Stars, 2020-02
    78/100 stars
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    99
    Becton Dickinson annexin v fitc pi apoptosis detection kit
    Pancreatic cancer cells are susceptible to isoorientin-induced <t>apoptosis.</t> Notes: The effect of isoorientin on apoptotic cell death. Cells were treated in the absence or presence of isoorientin (20, 40, 80, and 160 μM) for 24 hours. Apoptosis was evaluated by <t>Annexin</t> V staining ( A , B , D , E ). The expression of BCL-2 and BAX was evaluated by Western blotting( C and F ). GAPDH was used as a loading control. The immunoblots shown are representative of three independent experiments. The apoptosis data are presented as the mean ± SEM from three independent experiments ( * P
    Annexin V Fitc Pi Apoptosis Detection Kit, supplied by Becton Dickinson, used in various techniques. Bioz Stars score: 99/100, based on 990 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/annexin v fitc pi apoptosis detection kit/product/Becton Dickinson
    Average 99 stars, based on 990 article reviews
    Price from $9.99 to $1999.99
    annexin v fitc pi apoptosis detection kit - by Bioz Stars, 2020-02
    99/100 stars
      Buy from Supplier

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    Synthetic peptide N-DEVGEANN-C successfully inhibited Nef from interacting with and there by restore the functions of endogenous wild type ASK1 activity. ( A ) HEK-293 cells were transfected with Nef plasmid or vector backbone. 24 hrs post transfection, cells were stimulated to progress through ASK1 mediated apoptotic progression using classic ligand viz. TNF-α (100ng/ml) either in absence or presence of synthetic peptide N-DEVGEANN-C (10ng/ml). The extent of interaction of endogenous wild type ASK1 with Nef and ability of peptide N-DEVGEANN-C to impair the interaction was evaluated using In situ proximity ligation assay. ( B ) Synthetic peptide N-DEVGEANN-C prevented HIV-1Nef from offering protection against TNF -α induced, endogenous wild type ASK1 mediated apoptosis as evaluated using Annexin-V/PI assay. ( C ) Endogenous wild type ASK1 activity was rendered active by synthetic peptide N-DEVGEANN-C even in presence of HIV-1Nef as indicated by markedly decreased inhibitory Ser 967 phosphorylation of ASK1 with a corresponding increase in p54/46JNK1 activation profile as indicated by higher phospho-p54/46JNK1 levels in western blot analysis. All western blotting was done three times and integrated density was determined by densitometry analysis using imageJ software.

    Journal: PLoS ONE

    Article Title: Dynamics of Physical Interaction between HIV-1 Nef and ASK1: Identifying the Interacting Motif(S)

    doi: 10.1371/journal.pone.0067586

    Figure Lengend Snippet: Synthetic peptide N-DEVGEANN-C successfully inhibited Nef from interacting with and there by restore the functions of endogenous wild type ASK1 activity. ( A ) HEK-293 cells were transfected with Nef plasmid or vector backbone. 24 hrs post transfection, cells were stimulated to progress through ASK1 mediated apoptotic progression using classic ligand viz. TNF-α (100ng/ml) either in absence or presence of synthetic peptide N-DEVGEANN-C (10ng/ml). The extent of interaction of endogenous wild type ASK1 with Nef and ability of peptide N-DEVGEANN-C to impair the interaction was evaluated using In situ proximity ligation assay. ( B ) Synthetic peptide N-DEVGEANN-C prevented HIV-1Nef from offering protection against TNF -α induced, endogenous wild type ASK1 mediated apoptosis as evaluated using Annexin-V/PI assay. ( C ) Endogenous wild type ASK1 activity was rendered active by synthetic peptide N-DEVGEANN-C even in presence of HIV-1Nef as indicated by markedly decreased inhibitory Ser 967 phosphorylation of ASK1 with a corresponding increase in p54/46JNK1 activation profile as indicated by higher phospho-p54/46JNK1 levels in western blot analysis. All western blotting was done three times and integrated density was determined by densitometry analysis using imageJ software.

    Article Snippet: Annexin–V and PI staining Apoptosis /necrosis index was evaluated using BD Annexin–V/PI apoptosis detecting kit following manufacturer’s protocol.

    Techniques: Activity Assay, Transfection, Plasmid Preparation, In Situ, Proximity Ligation Assay, Activation Assay, Western Blot, Software

    Effects of cyanidin 3-O-glucoside treatment on apoptosis in H. pylori -infected KATO Ⅲ cells. (A), The assay of Annexin V and 7-AAD (7-Amino-Actinomycin) binding staining was performed with the same batch cells on Figure 3 . Data analysed with FACS flow cytometry (Becton-Dickinson). (B) , C3G treatment showed reduced apoptosis in H. pylori -infected KATO III cells, whereas cell viability of infected cells ( HP + / C3G-) was not recovered compared to uninfected control ( HP - / C3G- or HP - / C3G+) and C3G-treated groups ( HP + / C3G+).

    Journal: International Journal of Medical Sciences

    Article Title: Cyanidin 3-O-Glucoside Reduces Helicobacter pylori VacA-Induced Cell Death of Gastric KATO III Cells through Inhibition of the SecA Pathway

    doi: 10.7150/ijms.7167

    Figure Lengend Snippet: Effects of cyanidin 3-O-glucoside treatment on apoptosis in H. pylori -infected KATO Ⅲ cells. (A), The assay of Annexin V and 7-AAD (7-Amino-Actinomycin) binding staining was performed with the same batch cells on Figure 3 . Data analysed with FACS flow cytometry (Becton-Dickinson). (B) , C3G treatment showed reduced apoptosis in H. pylori -infected KATO III cells, whereas cell viability of infected cells ( HP + / C3G-) was not recovered compared to uninfected control ( HP - / C3G- or HP - / C3G+) and C3G-treated groups ( HP + / C3G+).

    Article Snippet: Annexin V and 7-AAD (7-Amino-Actinomycin) binding staining The assay of Annexin V and 7-AAD (7-Amino-Actinomycin) binding staining was performed with an Annexin V-PE Apoptosis Detection Kit I according to the manufacturer's instructions (Becton-Dickinson).

    Techniques: Infection, Binding Assay, Staining, FACS, Flow Cytometry, Cytometry

    Pancreatic cancer cells are susceptible to isoorientin-induced apoptosis. Notes: The effect of isoorientin on apoptotic cell death. Cells were treated in the absence or presence of isoorientin (20, 40, 80, and 160 μM) for 24 hours. Apoptosis was evaluated by Annexin V staining ( A , B , D , E ). The expression of BCL-2 and BAX was evaluated by Western blotting( C and F ). GAPDH was used as a loading control. The immunoblots shown are representative of three independent experiments. The apoptosis data are presented as the mean ± SEM from three independent experiments ( * P

    Journal: OncoTargets and therapy

    Article Title: Isoorientin induces apoptosis, decreases invasiveness, and downregulates VEGF secretion by activating AMPK signaling in pancreatic cancer cells

    doi: 10.2147/OTT.S122653

    Figure Lengend Snippet: Pancreatic cancer cells are susceptible to isoorientin-induced apoptosis. Notes: The effect of isoorientin on apoptotic cell death. Cells were treated in the absence or presence of isoorientin (20, 40, 80, and 160 μM) for 24 hours. Apoptosis was evaluated by Annexin V staining ( A , B , D , E ). The expression of BCL-2 and BAX was evaluated by Western blotting( C and F ). GAPDH was used as a loading control. The immunoblots shown are representative of three independent experiments. The apoptosis data are presented as the mean ± SEM from three independent experiments ( * P

    Article Snippet: The cell apoptosis rate was measured using the Annexin V-FITC/PI Apoptosis Detection Kit (BD, Franklin Lakes, NJ, USA), according to the manufacturer’s instructions.

    Techniques: Staining, Expressing, Western Blot

    Silibinin induced cell apoptosis in AsPC-1, Panc-1 and BxPC-3 cells. ( A ) Cells were treated with 2‰ (v/v) DMSO (Control) and 200 μM Silibinin for 24 h. Cells were then fixed and stained with DAPI. Stained nuclei were observed under a fluorescent microscope using a blue filter; ( B ) Cells treated with 100, 200 μM of Silibinin for 48 h were assessed for apoptosis by staining with Annexin V-FITC and PI; ( C ) Cells treated with 100, 200 μM of Silibinin for 48 h were assessed for apoptosis by staining with Annexin V-FITC and PI. The results are shown from one of three experiments with similar results. Each point represents the mean ± SD of three independent experiments. Significance was determined using a Student’s t-test ( p

    Journal: International Journal of Molecular Sciences

    Article Title: Silibinin Causes Apoptosis and Cell Cycle Arrest in Some Human Pancreatic Cancer Cells

    doi: 10.3390/ijms12084861

    Figure Lengend Snippet: Silibinin induced cell apoptosis in AsPC-1, Panc-1 and BxPC-3 cells. ( A ) Cells were treated with 2‰ (v/v) DMSO (Control) and 200 μM Silibinin for 24 h. Cells were then fixed and stained with DAPI. Stained nuclei were observed under a fluorescent microscope using a blue filter; ( B ) Cells treated with 100, 200 μM of Silibinin for 48 h were assessed for apoptosis by staining with Annexin V-FITC and PI; ( C ) Cells treated with 100, 200 μM of Silibinin for 48 h were assessed for apoptosis by staining with Annexin V-FITC and PI. The results are shown from one of three experiments with similar results. Each point represents the mean ± SD of three independent experiments. Significance was determined using a Student’s t-test ( p

    Article Snippet: Annexin-V-FITC Apoptosis Detection Kit I was obtained from BD, PVDF membrane was purchased from Millipore, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), 4,6-diamidino-2-phenylindile (DAPI), propidium iodide (PI) were purchased from Sigma, Caspase activity assay kits were obtained from Bestbio., DMEM was purchased from Invitrogen Corp. and fetal bovine serum (FBS) was purchased from GIBCO–BRL.

    Techniques: Staining, Microscopy