β lapachone  (Millipore)


Bioz Verified Symbol Millipore is a verified supplier  
  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 99
    Name:
    beta Lapachone
    Description:

    Catalog Number:
    l2037
    Price:
    None
    Buy from Supplier


    Structured Review

    Millipore β lapachone
    beta Lapachone

    https://www.bioz.com/result/β lapachone/product/Millipore
    Average 99 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
    β lapachone - by Bioz Stars, 2020-07
    99/100 stars

    Images

    1) Product Images from "Inactivation of the SMN complex by Oxidative Stress"

    Article Title: Inactivation of the SMN complex by Oxidative Stress

    Journal: Molecular cell

    doi: 10.1016/j.molcel.2008.06.004

    ROS reagents inhibit the activity of the SMN complex in vitro and in cells (A) Effect of H 2 O 2 on the activity of the SMN complex in vitro . Magnetic beads snRNP assembly assay was carried out in the presence of increasing amounts of H 2 O 2 . IC 50 was calculated from the dose-response curve. Error bars represent SDs from triplicate measurements. (B) Effect of menadione on SMN complex activity in vitro . The same experimental procedure was carried out as in (A), except that menadione was used instead. (C) Dose-dependent effect of menadione on SMN complex activity in cells. HeLa cells were treated with menadione at the indicated concentrations or with DMSO control for 1 hour. SMN complex activity in extracts from various treated cells was measured in comparison to DMSO control cell extract (100% activity) by magnetic beads snRNP assembly assay. Error bars represent SDs from 3 independent measurements. (D) Extracts from (C) mixed with sample buffer without or with DTT were resolved by SDS-PAGE and analyzed by Western blot of the entire membrane with anti-SMN antibody 62E7. The molecular mass markers in kDa are indicated on the left. “redSMN” indicates monomer SMN migrating at normal molecular mass and “oxSMN” indicates disulfide-crosslinked SMN upon oxidation. (E) β-lapachone and menadione generate ROS in live cells. HeLa cells were incubated 30 minutes with ROS indicator dye H 2 DCFDA (10 μM) or without dye as a control, then treated with compounds at indicated concentrations or DMSO as control. Fluorescence images were acquired 30 minutes after treatment.
    Figure Legend Snippet: ROS reagents inhibit the activity of the SMN complex in vitro and in cells (A) Effect of H 2 O 2 on the activity of the SMN complex in vitro . Magnetic beads snRNP assembly assay was carried out in the presence of increasing amounts of H 2 O 2 . IC 50 was calculated from the dose-response curve. Error bars represent SDs from triplicate measurements. (B) Effect of menadione on SMN complex activity in vitro . The same experimental procedure was carried out as in (A), except that menadione was used instead. (C) Dose-dependent effect of menadione on SMN complex activity in cells. HeLa cells were treated with menadione at the indicated concentrations or with DMSO control for 1 hour. SMN complex activity in extracts from various treated cells was measured in comparison to DMSO control cell extract (100% activity) by magnetic beads snRNP assembly assay. Error bars represent SDs from 3 independent measurements. (D) Extracts from (C) mixed with sample buffer without or with DTT were resolved by SDS-PAGE and analyzed by Western blot of the entire membrane with anti-SMN antibody 62E7. The molecular mass markers in kDa are indicated on the left. “redSMN” indicates monomer SMN migrating at normal molecular mass and “oxSMN” indicates disulfide-crosslinked SMN upon oxidation. (E) β-lapachone and menadione generate ROS in live cells. HeLa cells were incubated 30 minutes with ROS indicator dye H 2 DCFDA (10 μM) or without dye as a control, then treated with compounds at indicated concentrations or DMSO as control. Fluorescence images were acquired 30 minutes after treatment.

    Techniques Used: Activity Assay, In Vitro, Magnetic Beads, SDS Page, Western Blot, Incubation, Fluorescence

    ROS mapping of disulfide-crosslinked cysteines in SMN (A) Sequence alignment of human ( Homo sapien s), mouse ( Mus musculus ), and frog ( Xenopus laevies ) SMN protein sequences. Conserved residues are shaded in gray. Cysteine residues are highlighted. Exons and their boundaries are indicated with opposite-directed arrows. The schematic diagram shows SMN protein domain organization and positions of cysteines. Two cysteines (C60 and C250) that form disulfide bridges are marked (-S-S-). (B) Human SMN protein (WT, wild type; ΔEx7, exon7 deletion mutant; no Cys, mutation of all 8 cysteines to alanines; C60, C98, C123, and C250, mutation of 7 cysteines to alanines except for cysteine at positions 60, 98, 123, and 250, respectively) were produced by in vitro transcription and translation in the presence of 35 S-Met and then treated with 40 μM β-lapachone or DMSO for 1 hour. Samples were mixed with sample buffer without DTT, and then analyzed by SDS-PAGE and autoradiography. Molecular mass markers in kDa are indicated on the left. Protein bands corresponding to monomer SMN (redSMN), disulfide-crosslinked SMN (oxSMN) and SMN dimer (oxSMN dimer) are indicated on the right. (C) SMN amino terminus deletion (Ex3-7) and carboxyl terminus deletion (Ex1-4) mutants were tested for crosslinking, as described in panel (B). Note that these mutants were constructed in pcDNA3-myc-pyruvate kinase (PK, ~60kD) vector to facilitate detection of otherwise small fragments.
    Figure Legend Snippet: ROS mapping of disulfide-crosslinked cysteines in SMN (A) Sequence alignment of human ( Homo sapien s), mouse ( Mus musculus ), and frog ( Xenopus laevies ) SMN protein sequences. Conserved residues are shaded in gray. Cysteine residues are highlighted. Exons and their boundaries are indicated with opposite-directed arrows. The schematic diagram shows SMN protein domain organization and positions of cysteines. Two cysteines (C60 and C250) that form disulfide bridges are marked (-S-S-). (B) Human SMN protein (WT, wild type; ΔEx7, exon7 deletion mutant; no Cys, mutation of all 8 cysteines to alanines; C60, C98, C123, and C250, mutation of 7 cysteines to alanines except for cysteine at positions 60, 98, 123, and 250, respectively) were produced by in vitro transcription and translation in the presence of 35 S-Met and then treated with 40 μM β-lapachone or DMSO for 1 hour. Samples were mixed with sample buffer without DTT, and then analyzed by SDS-PAGE and autoradiography. Molecular mass markers in kDa are indicated on the left. Protein bands corresponding to monomer SMN (redSMN), disulfide-crosslinked SMN (oxSMN) and SMN dimer (oxSMN dimer) are indicated on the right. (C) SMN amino terminus deletion (Ex3-7) and carboxyl terminus deletion (Ex1-4) mutants were tested for crosslinking, as described in panel (B). Note that these mutants were constructed in pcDNA3-myc-pyruvate kinase (PK, ~60kD) vector to facilitate detection of otherwise small fragments.

    Techniques Used: Sequencing, Mutagenesis, Produced, In Vitro, SDS Page, Autoradiography, Construct, Plasmid Preparation

    β-lapachone potently and selectively inhibits the SMN complex-mediated snRNP assembly in vitro and in cells (A) Chemical structure of β-lapachone. (B) Concentration-dependent inhibition of SMN complex activity by β-lapachone in cells. HeLa cells were treated with various concentrations of β-lapachone or with DMSO (control) for 1 hour. SMN complex activity in extracts from treated cells was measured using magnetic beads snRNP assembly assay and compared to DMSO-treated control cells (100% activity). IC 50 was calculated from the dose-reponse graph. Error bars represent SDs from 3 independent measurements. (C) β-lapachone selectively inhibits SMN complex-mediated Sm core assembly. Assembly reactions were performed using either cell extracts or purified native snRNP proteins lacking the SMN complex (Sm proteins). Both samples were adjusted to contain a similar amount of Sm proteins. Magnetic beads snRNP assembly assay was carried out with U4 or control U4ΔSm snRNA in the presence of either 20 or 100 μM β-lapachone or DMSO control. Sm core assembly on U4 snRNA in the presence of DMSO was considered 100% activity. The error bars represent SDs from 3 independent measurements.
    Figure Legend Snippet: β-lapachone potently and selectively inhibits the SMN complex-mediated snRNP assembly in vitro and in cells (A) Chemical structure of β-lapachone. (B) Concentration-dependent inhibition of SMN complex activity by β-lapachone in cells. HeLa cells were treated with various concentrations of β-lapachone or with DMSO (control) for 1 hour. SMN complex activity in extracts from treated cells was measured using magnetic beads snRNP assembly assay and compared to DMSO-treated control cells (100% activity). IC 50 was calculated from the dose-reponse graph. Error bars represent SDs from 3 independent measurements. (C) β-lapachone selectively inhibits SMN complex-mediated Sm core assembly. Assembly reactions were performed using either cell extracts or purified native snRNP proteins lacking the SMN complex (Sm proteins). Both samples were adjusted to contain a similar amount of Sm proteins. Magnetic beads snRNP assembly assay was carried out with U4 or control U4ΔSm snRNA in the presence of either 20 or 100 μM β-lapachone or DMSO control. Sm core assembly on U4 snRNA in the presence of DMSO was considered 100% activity. The error bars represent SDs from 3 independent measurements.

    Techniques Used: In Vitro, Concentration Assay, Inhibition, Activity Assay, Magnetic Beads, Purification

    SMN protein is oxidized to form intermolecular disulfide bonds upon β-lapachone treatment (A) Indirect immunofluorescence staining of SMN (2B1; green) and snRNPs (Y12; red) in HeLa PV cells treated 3 hours with 5 μM β-lapachone or DMSO control. (B) HeLa total cell extracts prepared from cells treated 3 hours with 5 μM β-lapachone or DMSO control were resolved by SDS-PAGE and analyzed by quantitative Western blotting, using JBP1 and Magoh as loading controls. Extracts were prepared and mixed with sample buffer without DTT. The membrane was cut into strips for probing of each protein at the corresponding molecular mass. ) were used for in vitro assembly reactions in the presence of either 100 μM β-lapachone or DMSO control. The SMN complex was isolated by anti-Flag immunoprecipitation, mixed with sample buffer without or with DTT and resolved by SDS-PAGE. Western blot analysis was performed on the entire membrane with anti-SMN antibody 62E7. Molecular mass markers in kDa are indicated on the left. “redSMN” indicates monomer SMN migrating at normal molecular mass and “oxSMN” indicates disulfide-crosslinked SMN upon oxidation.
    Figure Legend Snippet: SMN protein is oxidized to form intermolecular disulfide bonds upon β-lapachone treatment (A) Indirect immunofluorescence staining of SMN (2B1; green) and snRNPs (Y12; red) in HeLa PV cells treated 3 hours with 5 μM β-lapachone or DMSO control. (B) HeLa total cell extracts prepared from cells treated 3 hours with 5 μM β-lapachone or DMSO control were resolved by SDS-PAGE and analyzed by quantitative Western blotting, using JBP1 and Magoh as loading controls. Extracts were prepared and mixed with sample buffer without DTT. The membrane was cut into strips for probing of each protein at the corresponding molecular mass. ) were used for in vitro assembly reactions in the presence of either 100 μM β-lapachone or DMSO control. The SMN complex was isolated by anti-Flag immunoprecipitation, mixed with sample buffer without or with DTT and resolved by SDS-PAGE. Western blot analysis was performed on the entire membrane with anti-SMN antibody 62E7. Molecular mass markers in kDa are indicated on the left. “redSMN” indicates monomer SMN migrating at normal molecular mass and “oxSMN” indicates disulfide-crosslinked SMN upon oxidation.

    Techniques Used: Immunofluorescence, Staining, SDS Page, Western Blot, In Vitro, Isolation, Immunoprecipitation

    DTT prevents the inhibition of the activity of the SMN complex by β-lapachone Cell extracts treated with 20 μM β-lapachone, or 20 μM β-lapachone together with 20 mM DTT, or DMSO control were analyzed by non-reducing Western blot. The relative levels of monomer SMN (“redSMN”) were calculated as the percentage of that in DMSO control and shown by the blue bar. Assembly activities of the SMN complex were measured by magnetic beads snRNP assembly assay using the same set of treated extracts and shown by the red bar. Error bars represent SDs from 3 independent experiments.
    Figure Legend Snippet: DTT prevents the inhibition of the activity of the SMN complex by β-lapachone Cell extracts treated with 20 μM β-lapachone, or 20 μM β-lapachone together with 20 mM DTT, or DMSO control were analyzed by non-reducing Western blot. The relative levels of monomer SMN (“redSMN”) were calculated as the percentage of that in DMSO control and shown by the blue bar. Assembly activities of the SMN complex were measured by magnetic beads snRNP assembly assay using the same set of treated extracts and shown by the red bar. Error bars represent SDs from 3 independent experiments.

    Techniques Used: Inhibition, Activity Assay, Western Blot, Magnetic Beads

    2) Product Images from "β-Lapachone induces programmed necrosis through the RIP1-PARP-AIF-dependent pathway in human hepatocellular carcinoma SK-Hep1 cells"

    Article Title: β-Lapachone induces programmed necrosis through the RIP1-PARP-AIF-dependent pathway in human hepatocellular carcinoma SK-Hep1 cells

    Journal: Cell Death & Disease

    doi: 10.1038/cddis.2014.202

    β -Lapachone induces necroptosis via RIP1-PARP-1-AIF signaling. ( a ) SK-Hep1 cells were treated with the indicated concentrations of  β -lapachone and 300 ng/ml TRAIL (PC; positive control) for 18 h. ( b ) SK-Hep1 cells were treated with the indicated time of 4  μ M  β -lapachone. ( c ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of 30  μ M DPQ for 1 h. ( d ,  e ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of 30  μ M DPQ for 18 h. Cell death was analyzed with the LDH release ( d ). Cell viability was analyzed using XTT assay ( e ). ( f ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of 50  μ M DPQ for 14 h. Cytoplasmic fractions were analyzed for AIF cytosol translocation. ( g ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of necrostatin-1 (60  μ M) for 1 h. ( h ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of necrostatin-1 (60  μ M) for 14 h. Cytoplasmic fractions were analyzed for AIF cytosol translocation. Fractionation was performed and the translocation of AIF was determined by immunoblotting with an anti-AIF antibody. The PARP ( a ), PAR ( b ,  c ,  g ), and AIF ( f ,  h ) protein levels were determined by western blotting. The level of actin and ERK was used as a loading control. MnSOD was used as a mitochondria loading control. The values in  d ,  e  represent the mean±S.D. from three independent samples ( n  =3). * P
    Figure Legend Snippet: β -Lapachone induces necroptosis via RIP1-PARP-1-AIF signaling. ( a ) SK-Hep1 cells were treated with the indicated concentrations of β -lapachone and 300 ng/ml TRAIL (PC; positive control) for 18 h. ( b ) SK-Hep1 cells were treated with the indicated time of 4  μ M β -lapachone. ( c ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of 30  μ M DPQ for 1 h. ( d , e ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of 30  μ M DPQ for 18 h. Cell death was analyzed with the LDH release ( d ). Cell viability was analyzed using XTT assay ( e ). ( f ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of 50  μ M DPQ for 14 h. Cytoplasmic fractions were analyzed for AIF cytosol translocation. ( g ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of necrostatin-1 (60  μ M) for 1 h. ( h ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of necrostatin-1 (60  μ M) for 14 h. Cytoplasmic fractions were analyzed for AIF cytosol translocation. Fractionation was performed and the translocation of AIF was determined by immunoblotting with an anti-AIF antibody. The PARP ( a ), PAR ( b , c , g ), and AIF ( f , h ) protein levels were determined by western blotting. The level of actin and ERK was used as a loading control. MnSOD was used as a mitochondria loading control. The values in d , e represent the mean±S.D. from three independent samples ( n =3). * P

    Techniques Used: Positive Control, XTT Assay, Translocation Assay, Fractionation, Western Blot

    β -Lapachone induces programmed necrosis  via  ROS generation in SK-Hep1 cells. ( a ,  b ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of GSH (5 mM) or NAC (5 mM) for 1 h, and loaded with a fluorescent dye, H 2 DCF-DA. H 2 DCF-DA fluorescence intensity was measured by flow cytometry ( a ). MitoSOX Red intensity was analyzed with fluorescent microscope ( b ). ( c ,  d ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of GSH (5 mM) or NAC (5 mM) for 18 h ( c ) or 1 h ( d ). Cell death was analyzed via the LDH release ( c ). PAR accumulation was determined by western blotting ( d ). ( e ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of GSH (5 mM) or NAC (5 mM) for 14 h. Fractionation was performed and the translocation of AIF was determined by immunoblotting with an anti-AIF antibody. The level of ERK was used as a loading control and cytoplasmic fraction marker, and MnSOD was used as a loading control and mitochondria fraction marker. ( f ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of necrostatin-1 (60  μ M) for 1 h, and loaded with fluorescent dye, H 2 DCF-DA or MitoSOX Red. H 2 DCF-DA and MitoSOX Red fluorescence intensity was analyzed with fluorescent microscope. ( g ) RIP1+/+ MEF and RIP1−/− MEF cells were treated with or without 4  μ M  β -lapachone for 1 h, and loaded with H 2 DCF-DA or MitoSOX Red. H 2 DCF-DA and MitoSOX Red intensity was analyzed with fluorescent microscope. The values in  c  represent the mean±S.D. from three independent samples ( n  =3)
    Figure Legend Snippet: β -Lapachone induces programmed necrosis via ROS generation in SK-Hep1 cells. ( a , b ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of GSH (5 mM) or NAC (5 mM) for 1 h, and loaded with a fluorescent dye, H 2 DCF-DA. H 2 DCF-DA fluorescence intensity was measured by flow cytometry ( a ). MitoSOX Red intensity was analyzed with fluorescent microscope ( b ). ( c , d ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of GSH (5 mM) or NAC (5 mM) for 18 h ( c ) or 1 h ( d ). Cell death was analyzed via the LDH release ( c ). PAR accumulation was determined by western blotting ( d ). ( e ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of GSH (5 mM) or NAC (5 mM) for 14 h. Fractionation was performed and the translocation of AIF was determined by immunoblotting with an anti-AIF antibody. The level of ERK was used as a loading control and cytoplasmic fraction marker, and MnSOD was used as a loading control and mitochondria fraction marker. ( f ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of necrostatin-1 (60  μ M) for 1 h, and loaded with fluorescent dye, H 2 DCF-DA or MitoSOX Red. H 2 DCF-DA and MitoSOX Red fluorescence intensity was analyzed with fluorescent microscope. ( g ) RIP1+/+ MEF and RIP1−/− MEF cells were treated with or without 4  μ M β -lapachone for 1 h, and loaded with H 2 DCF-DA or MitoSOX Red. H 2 DCF-DA and MitoSOX Red intensity was analyzed with fluorescent microscope. The values in c represent the mean±S.D. from three independent samples ( n =3)

    Techniques Used: Fluorescence, Flow Cytometry, Cytometry, Microscopy, Western Blot, Fractionation, Translocation Assay, Marker

    β -Lapachone induces RIP1-dependent necroptosis. ( a ) SK-Hep1 cells stimulated with 4 μM  β -lapachone in the presence or absence of 60  μ M necrostatin-1 for 18 h. The cell morphologies were determined by interference light microscopy. Images were magnified × 200. ( b ,  c ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone in the presence or absence of 60  μ M necrostatin-1 for 18 h. Cell death was analyzed with the LDH release ( b ). Cells stained with propidium iodide, and PI uptake was determined by flow cytometry ( c ). ( d ) SK-Hep1 cells stimulated with 4  μ M  β -lapachone in the presence or absence of 60  μ M necrostatin-1 for 18 h. The levels of HMGB1 in whole cell and culture media were determined by immunoblotting with an anti-HMGB1 antibody. ( e ,  f ) SK-Hep1 cells were transfected with RIP1 siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of  β -lapachone for 18 h. Cell viability was analyzed using XTT assay (left panel). The RIP1 protein levels were determined by Western blotting. The level of actin was used as a loading control (right panel) ( e ). Cells stained with propidium iodide, and PI uptake was determined by flow cytometry ( f ). ( g ,  h ) MEF and RIP1−/− MEF cells were treated with the indicated concentrations of  β -lapachone for 18 h. Cell viability was analyzed using XTT assay (left panel). The RIP1 protein levels were determined by western blotting. The level of actin was used as a loading control (right panel) ( g ). Cells stained with propidium iodide, and PI uptake was determined by flow cytometry ( h ). The values in  b ,  c ,  e ,  f ,  g ,  h  represent the mean±S.D. from three independent samples ( n  =3). * P
    Figure Legend Snippet: β -Lapachone induces RIP1-dependent necroptosis. ( a ) SK-Hep1 cells stimulated with 4 μM β -lapachone in the presence or absence of 60  μ M necrostatin-1 for 18 h. The cell morphologies were determined by interference light microscopy. Images were magnified × 200. ( b , c ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone in the presence or absence of 60  μ M necrostatin-1 for 18 h. Cell death was analyzed with the LDH release ( b ). Cells stained with propidium iodide, and PI uptake was determined by flow cytometry ( c ). ( d ) SK-Hep1 cells stimulated with 4  μ M β -lapachone in the presence or absence of 60  μ M necrostatin-1 for 18 h. The levels of HMGB1 in whole cell and culture media were determined by immunoblotting with an anti-HMGB1 antibody. ( e , f ) SK-Hep1 cells were transfected with RIP1 siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of β -lapachone for 18 h. Cell viability was analyzed using XTT assay (left panel). The RIP1 protein levels were determined by Western blotting. The level of actin was used as a loading control (right panel) ( e ). Cells stained with propidium iodide, and PI uptake was determined by flow cytometry ( f ). ( g , h ) MEF and RIP1−/− MEF cells were treated with the indicated concentrations of β -lapachone for 18 h. Cell viability was analyzed using XTT assay (left panel). The RIP1 protein levels were determined by western blotting. The level of actin was used as a loading control (right panel) ( g ). Cells stained with propidium iodide, and PI uptake was determined by flow cytometry ( h ). The values in b , c , e , f , g , h represent the mean±S.D. from three independent samples ( n =3). * P

    Techniques Used: Light Microscopy, Staining, Flow Cytometry, Cytometry, Transfection, XTT Assay, Western Blot

    β -Lapachone-induced AIF translocation is associated with necroptosis. ( a ) SK-Hep1 cells were stained for mitochondria with MitoTracker Red, for AIF with an anti-AIF antibody followed by an FITC-conjugated secondary antibody and for nuclei with Hoechst staining. ( b ,  c ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone. Cytoplasmic and nucleus fractions were analyzed for cytosol and nucleus translocation of AIF. The protein levels of AIF and cytochrome C were determined by western blotting. The level of ERK was used as a loading control. Ref-1 was used as a nucleus loading control and MnSOD was used as a mitochondria loading control. ( d ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone and incubated with antibody against AIF followed by labeling with the FITC-conjugated secondary antibody. Nuclei were stained with propidium iodide (PI). Yellow, nuclear translocation of AIF is shown by overlap of AIF (green fluorescence) and nuclear PI staining (red fluorescence). Graph represents relative intensity of nuclear AIF (right panel). ( e ,  f ) SK-Hep1 cells were transfected with AIF siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of  β -lapachone for 18 h. Cell viability was analyzed using XTT assay ( e ). Cell death was analyzed using LDH assay ( f ). The AIF protein levels were determined by western blotting. The level of actin was used as a loading control ( e ,  f ). ( g ) SK-Hep1 cells were transfected with AIF siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of  β -lapachone for 5 days. After treatment, cells were stained with Coomassie Brilliant Blue dye. The AIF protein levels were determined by western blotting. The level of actin was used as a loading control. The values in  d ,  e ,  f  represent the mean±S.D. from three independent samples ( n  =3). * P
    Figure Legend Snippet: β -Lapachone-induced AIF translocation is associated with necroptosis. ( a ) SK-Hep1 cells were stained for mitochondria with MitoTracker Red, for AIF with an anti-AIF antibody followed by an FITC-conjugated secondary antibody and for nuclei with Hoechst staining. ( b , c ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone. Cytoplasmic and nucleus fractions were analyzed for cytosol and nucleus translocation of AIF. The protein levels of AIF and cytochrome C were determined by western blotting. The level of ERK was used as a loading control. Ref-1 was used as a nucleus loading control and MnSOD was used as a mitochondria loading control. ( d ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone and incubated with antibody against AIF followed by labeling with the FITC-conjugated secondary antibody. Nuclei were stained with propidium iodide (PI). Yellow, nuclear translocation of AIF is shown by overlap of AIF (green fluorescence) and nuclear PI staining (red fluorescence). Graph represents relative intensity of nuclear AIF (right panel). ( e , f ) SK-Hep1 cells were transfected with AIF siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of β -lapachone for 18 h. Cell viability was analyzed using XTT assay ( e ). Cell death was analyzed using LDH assay ( f ). The AIF protein levels were determined by western blotting. The level of actin was used as a loading control ( e , f ). ( g ) SK-Hep1 cells were transfected with AIF siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of β -lapachone for 5 days. After treatment, cells were stained with Coomassie Brilliant Blue dye. The AIF protein levels were determined by western blotting. The level of actin was used as a loading control. The values in d , e , f represent the mean±S.D. from three independent samples ( n =3). * P

    Techniques Used: Translocation Assay, Staining, Western Blot, Incubation, Labeling, Fluorescence, Transfection, XTT Assay, Lactate Dehydrogenase Assay

    β -Lapachone induces necrotic cell death in human hepatocellular carcinoma SK-Hep1 cells. ( a – c ) SK-Hep1 cells were treated with the indicated concentrations of  β -lapachone for 18 h. The cell morphologies were determined by interference light microscopy. Images were magnified × 200 ( a ). Cell viability was analyzed using XTT assay ( b ). Cells were stained with propidium iodide (PI), and PI uptake was determined by flow cytometry ( c ). ( d – f ) SK-Hep1 cells were stimulated with 4  μ M  β -lapachone or 5  μ M MG132 for 18 h in the presence or absence of zVAD-fmk (50  μ M) for 18 h. The cell morphologies were determined by interference light microscopy. Images were magnified × 200 ( d ). Cell viability was analyzed using XTT assay ( e ). Cells were stained with propidium iodide, and PI uptake was determined by flow cytometry ( f ). ( g ) SK-Hep1 cells were treated with the indicated concentrations of  β -lapachone or 5  μ M MG132 for 18 h. Caspase activities were determined with colorimetric assays using caspase-3 DEVDase assay kits. ( h ) Transmission electron microscopic observation was conducted on SK-Hep1 cells treated with 4  μ M  β -lapachone for 18 h. ( i ) SK-Hep1 cells were treated with 4  μ M  β -lapachone or 5  μ M MG132 for 18 h, collected and stained with 7-AAD and Annexin V. Cell death was determined by flow cytometry. Values correspond to the percentage of cells in those quadrants. ( j ) SK-Hep1 cells were treated with 4  μ M  β -lapachone for 18 h, collected and stained with propidium iodide. PI uptake was determined by flow cytometry. ( k ) SK-Hep1 cells were treated with 4  μ M  β -lapachone for 18 h, stained with PI and Hoechst33342, and analyzed with a fluorescence microscope. ( l ) SK-Hep1 cells were treated with the indicated concentrations of  β -lapachone for 18 h (left panel) or 4  μ M  β -lapachone for indicated time periods (right panel). The levels of HMGB1 in whole cell and culture media were determined by immunoblotting with an anti-HMGB1 antibody. The values in  b ,  c ,  e ,  f ,  g , and  j  represent the mean±S.D. from three independent samples ( n  =3). * P
    Figure Legend Snippet: β -Lapachone induces necrotic cell death in human hepatocellular carcinoma SK-Hep1 cells. ( a – c ) SK-Hep1 cells were treated with the indicated concentrations of β -lapachone for 18 h. The cell morphologies were determined by interference light microscopy. Images were magnified × 200 ( a ). Cell viability was analyzed using XTT assay ( b ). Cells were stained with propidium iodide (PI), and PI uptake was determined by flow cytometry ( c ). ( d – f ) SK-Hep1 cells were stimulated with 4  μ M β -lapachone or 5  μ M MG132 for 18 h in the presence or absence of zVAD-fmk (50  μ M) for 18 h. The cell morphologies were determined by interference light microscopy. Images were magnified × 200 ( d ). Cell viability was analyzed using XTT assay ( e ). Cells were stained with propidium iodide, and PI uptake was determined by flow cytometry ( f ). ( g ) SK-Hep1 cells were treated with the indicated concentrations of β -lapachone or 5  μ M MG132 for 18 h. Caspase activities were determined with colorimetric assays using caspase-3 DEVDase assay kits. ( h ) Transmission electron microscopic observation was conducted on SK-Hep1 cells treated with 4  μ M β -lapachone for 18 h. ( i ) SK-Hep1 cells were treated with 4  μ M β -lapachone or 5  μ M MG132 for 18 h, collected and stained with 7-AAD and Annexin V. Cell death was determined by flow cytometry. Values correspond to the percentage of cells in those quadrants. ( j ) SK-Hep1 cells were treated with 4  μ M β -lapachone for 18 h, collected and stained with propidium iodide. PI uptake was determined by flow cytometry. ( k ) SK-Hep1 cells were treated with 4  μ M β -lapachone for 18 h, stained with PI and Hoechst33342, and analyzed with a fluorescence microscope. ( l ) SK-Hep1 cells were treated with the indicated concentrations of β -lapachone for 18 h (left panel) or 4  μ M β -lapachone for indicated time periods (right panel). The levels of HMGB1 in whole cell and culture media were determined by immunoblotting with an anti-HMGB1 antibody. The values in b , c , e , f , g , and j represent the mean±S.D. from three independent samples ( n =3). * P

    Techniques Used: Light Microscopy, XTT Assay, Staining, Flow Cytometry, Cytometry, Transmission Assay, Fluorescence, Microscopy

    β -Lapachone induces programmed necrosis through NQO1 enzyme activity in SK-Hep1 cells. ( a ) SK-Hep1 cells stimulated with 4  μ M  β -lapachone in the presence or absence of dicoumarol (50  μ M) for 18 h. Cell death was analyzed with LDH release. ( b ) SK-Hep1 cells were transfected with NQO1 siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of  β -lapachone for 18 h. Western blotting analysis and LDH assay were performed. ( c ) Vector cells (SK-Hep1/Vector) and NQO1 overexpressed cells (SK-Hep1/NQO1) were treated with 1  μ M  β -lapachone for 18 h. Cell death was analyzed using LDH assay (left panel). Western blotting analysis was performed for confirmed overexpression of NQO1 (right panel). ( d ) SK-Hep1 cells were transfected with vector, NQO1 and NQO1 shRNA. Twenty-four hours after transfection, cell were treated with 4  μ M  β -lapachone for 1 h, and loaded with fluorescent dye, H 2 DCF-DA. H 2 DCF-DA fluorescence intensity was analyzed with a fluorescent microscope. The NQO1 protein levels were determined by western blotting. The level of actin was used as a loading control. ( e ) The NQO1 protein levels were determined by western blotting. The level of actin was used as a loading control (upper panel). NQO1 activity was determined in cell extracts prepared from U343, T98G, Caki, ACHN, MDA-MB-361, MDA-MB-231 and SK-Hep1 cells (lower panel). ( f ,  g ) U343, T98G, Caki, ACHN, MDA-MB-361, MDA-MB-231 and SK-Hep1 cells were treated with the indicated concentrations of  β -lapachone for 14 h. Cell viability was analyzed using XTT assay ( f ). Cell death was analyzed with LDH assay ( g ). The values in  a ,  b ,  c ,  e ,  f ,  g  represent the mean±S.D. from three independent samples ( n  =3). * P
    Figure Legend Snippet: β -Lapachone induces programmed necrosis through NQO1 enzyme activity in SK-Hep1 cells. ( a ) SK-Hep1 cells stimulated with 4  μ M β -lapachone in the presence or absence of dicoumarol (50  μ M) for 18 h. Cell death was analyzed with LDH release. ( b ) SK-Hep1 cells were transfected with NQO1 siRNA or control siRNA. Twenty-four hours after transfection, cells were treated with the indicated concentrations of β -lapachone for 18 h. Western blotting analysis and LDH assay were performed. ( c ) Vector cells (SK-Hep1/Vector) and NQO1 overexpressed cells (SK-Hep1/NQO1) were treated with 1  μ M β -lapachone for 18 h. Cell death was analyzed using LDH assay (left panel). Western blotting analysis was performed for confirmed overexpression of NQO1 (right panel). ( d ) SK-Hep1 cells were transfected with vector, NQO1 and NQO1 shRNA. Twenty-four hours after transfection, cell were treated with 4  μ M β -lapachone for 1 h, and loaded with fluorescent dye, H 2 DCF-DA. H 2 DCF-DA fluorescence intensity was analyzed with a fluorescent microscope. The NQO1 protein levels were determined by western blotting. The level of actin was used as a loading control. ( e ) The NQO1 protein levels were determined by western blotting. The level of actin was used as a loading control (upper panel). NQO1 activity was determined in cell extracts prepared from U343, T98G, Caki, ACHN, MDA-MB-361, MDA-MB-231 and SK-Hep1 cells (lower panel). ( f , g ) U343, T98G, Caki, ACHN, MDA-MB-361, MDA-MB-231 and SK-Hep1 cells were treated with the indicated concentrations of β -lapachone for 14 h. Cell viability was analyzed using XTT assay ( f ). Cell death was analyzed with LDH assay ( g ). The values in a , b , c , e , f , g represent the mean±S.D. from three independent samples ( n =3). * P

    Techniques Used: Activity Assay, Transfection, Western Blot, Lactate Dehydrogenase Assay, Plasmid Preparation, Over Expression, shRNA, Fluorescence, Microscopy, Multiple Displacement Amplification, XTT Assay

    3) Product Images from "Terpinen-4-ol, tyrosol, and β-lapachone as potential antifungals against dimorphic fungi"

    Article Title: Terpinen-4-ol, tyrosol, and β-lapachone as potential antifungals against dimorphic fungi

    Journal: Brazilian Journal of Microbiology

    doi: 10.1016/j.bjm.2016.07.015

    Effect of terpinen-4-ol (TER), tyrosol (TIR), β-lapachone (B-LAP), amphotericin B (AMB, control drug), and isoniazid (INH, positive control drug) at MIC and MIC/2 (μg/mL) on the permeability of the cell membrane of the ten strains of C. posadasii strains (1a–b), eight strains of filamentous (F) H. capsulatum strains (1c–d) and eight yeast strains (Y) of H. capsulatum (1e–f) resulting in the leakage of nucleic acids (260 nm) (1a, 1c, 1e) and proteins (280 nm) (1b, 1d, 1f). These values represent the mean ± standard error of the absorbance values obtained for all tested strains. Control: fungal growth without drug.
    Figure Legend Snippet: Effect of terpinen-4-ol (TER), tyrosol (TIR), β-lapachone (B-LAP), amphotericin B (AMB, control drug), and isoniazid (INH, positive control drug) at MIC and MIC/2 (μg/mL) on the permeability of the cell membrane of the ten strains of C. posadasii strains (1a–b), eight strains of filamentous (F) H. capsulatum strains (1c–d) and eight yeast strains (Y) of H. capsulatum (1e–f) resulting in the leakage of nucleic acids (260 nm) (1a, 1c, 1e) and proteins (280 nm) (1b, 1d, 1f). These values represent the mean ± standard error of the absorbance values obtained for all tested strains. Control: fungal growth without drug.

    Techniques Used: Positive Control, Permeability

    Quantification of ergosterol from the ten strains of Coccidioides posadasii (2a), eight filamentous (F) strains of Histoplasma capsulatum (2b), and eight yeast (Y) strain of H. capsulatum (2c) after exposure to sub-inhibitory concentrations of terpinen-4-ol (TER), tyrosol (TIR), β-lapachone (B-LAP), itraconazole (ITC, control drug), and amphotericin B (AMB, control drug). These values represent the average results for all strains tested for each species. Control: fungal growth control without drug.
    Figure Legend Snippet: Quantification of ergosterol from the ten strains of Coccidioides posadasii (2a), eight filamentous (F) strains of Histoplasma capsulatum (2b), and eight yeast (Y) strain of H. capsulatum (2c) after exposure to sub-inhibitory concentrations of terpinen-4-ol (TER), tyrosol (TIR), β-lapachone (B-LAP), itraconazole (ITC, control drug), and amphotericin B (AMB, control drug). These values represent the average results for all strains tested for each species. Control: fungal growth control without drug.

    Techniques Used:

    4) Product Images from "?-LAPACHONE AMELIORIZATION OF EXPERIMENTAL AUTOIMMUNE ENCEPHALOMYELITIS"

    Article Title: ?-LAPACHONE AMELIORIZATION OF EXPERIMENTAL AUTOIMMUNE ENCEPHALOMYELITIS

    Journal: Journal of neuroimmunology

    doi: 10.1016/j.jneuroim.2012.09.004

    Administration of β-lapachone inhibits IL-12 family subunit mRNA expression in EAE mice
    Figure Legend Snippet: Administration of β-lapachone inhibits IL-12 family subunit mRNA expression in EAE mice

    Techniques Used: Expressing, Mouse Assay

    Administration of β-lapachone inhibits mRNA expression of IL-17RA, IL-23R and IFN- γ in EAE mice
    Figure Legend Snippet: Administration of β-lapachone inhibits mRNA expression of IL-17RA, IL-23R and IFN- γ in EAE mice

    Techniques Used: Expressing, Mouse Assay

    Effects of β-lapachone on the production of IL-12 family cytokines by bone marrow derived dendritic cells (BM-DCs)
    Figure Legend Snippet: Effects of β-lapachone on the production of IL-12 family cytokines by bone marrow derived dendritic cells (BM-DCs)

    Techniques Used: Derivative Assay

    Effects of β-lapachone on IL-17A production in CD4 + T cells treated with microglial conditioned medium
    Figure Legend Snippet: Effects of β-lapachone on IL-17A production in CD4 + T cells treated with microglial conditioned medium

    Techniques Used:

    Treatment with β-lapachone suppresses the clinical severity of EAE
    Figure Legend Snippet: Treatment with β-lapachone suppresses the clinical severity of EAE

    Techniques Used:

    Administration of β-lapachone inhibits mRNA expression of TLR signaling molecules in EAE mice
    Figure Legend Snippet: Administration of β-lapachone inhibits mRNA expression of TLR signaling molecules in EAE mice

    Techniques Used: Expressing, Mouse Assay

    Effects of β-lapachone on the production of IL-12 family cytokines by mouse primary microglia
    Figure Legend Snippet: Effects of β-lapachone on the production of IL-12 family cytokines by mouse primary microglia

    Techniques Used:

    5) Product Images from "Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells"

    Article Title: Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0088122

    Dicoumarol, an NQO1 inhibitor, blocks the apoptotic effects of β-lapachone. (A) CL1-1 cells (top) or CL1-5 cells (bottom) were left untreated or were incubated for 6 h with 5 µM β-lapachone and/or 10 µM dicoumarol, then stained with Annexin V-FITC and the Annexin V fluorescence measured by flow cytometry. (B) Morphological changes after drug treatment. CL1-1 or CL1-5 cells were left untreated (CTL) or were incubated for 24 h with 5 µM β-lapachone with or without 10 µM dicoumarol, then stained with acridine orange to observe the morphology of the cell nucleus. The scale bar represents 50 µm.
    Figure Legend Snippet: Dicoumarol, an NQO1 inhibitor, blocks the apoptotic effects of β-lapachone. (A) CL1-1 cells (top) or CL1-5 cells (bottom) were left untreated or were incubated for 6 h with 5 µM β-lapachone and/or 10 µM dicoumarol, then stained with Annexin V-FITC and the Annexin V fluorescence measured by flow cytometry. (B) Morphological changes after drug treatment. CL1-1 or CL1-5 cells were left untreated (CTL) or were incubated for 24 h with 5 µM β-lapachone with or without 10 µM dicoumarol, then stained with acridine orange to observe the morphology of the cell nucleus. The scale bar represents 50 µm.

    Techniques Used: Incubation, Staining, Fluorescence, Flow Cytometry, Cytometry, CTL Assay

    β-lapachone-induced cell death is associated with NQO1 expression levels. (A) Percentage survival of the lung cancer cell lines CL1-1, CL1-5, and A549. Cells were treated with 0–10 µM β-lapachone for 12 h, then cell viability was determined by crystal violet staining assay and expressed as a percentage of the value for cultures with no β-lapachone. (B–D) NQO1 activity levels (B), NQO1 RNA expression levels (C), and NQO1 protein expression levels (D) in the three lung cancer cell lines grown under normal culture conditions. (E) Percentage survival of A549 cells (left panel), CL1-1 cells (center panel), and CL1-5 cells (right panel) incubated with the indicated concentration of β-lapachone for 3, 6, 12, or 24 h examined by crystal violet staining and expressed as percentage survival compared to the untreated cells. The results are the mean ± SD for 3 independent experiments, each in triplicate.
    Figure Legend Snippet: β-lapachone-induced cell death is associated with NQO1 expression levels. (A) Percentage survival of the lung cancer cell lines CL1-1, CL1-5, and A549. Cells were treated with 0–10 µM β-lapachone for 12 h, then cell viability was determined by crystal violet staining assay and expressed as a percentage of the value for cultures with no β-lapachone. (B–D) NQO1 activity levels (B), NQO1 RNA expression levels (C), and NQO1 protein expression levels (D) in the three lung cancer cell lines grown under normal culture conditions. (E) Percentage survival of A549 cells (left panel), CL1-1 cells (center panel), and CL1-5 cells (right panel) incubated with the indicated concentration of β-lapachone for 3, 6, 12, or 24 h examined by crystal violet staining and expressed as percentage survival compared to the untreated cells. The results are the mean ± SD for 3 independent experiments, each in triplicate.

    Techniques Used: Expressing, Staining, Activity Assay, RNA Expression, Incubation, Concentration Assay

    Signaling pathway components involved in β-lapachone-induced apoptosis. (A) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for the indicated time, then levels of p-ERK, ERK, p-JNK, and JNK were measured by Western blotting. (B) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then levels of p-PI3K and p-AKT were examined by Western blotting. (C) CL1-1 cells (left) or CL1-5 cells (right) were pretreated with the indicated concentrations of the JNK inhibitor sp600125 for 6 h, and then treated with or without 5 µM β-lapachone for 24 h.Cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells.* p
    Figure Legend Snippet: Signaling pathway components involved in β-lapachone-induced apoptosis. (A) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for the indicated time, then levels of p-ERK, ERK, p-JNK, and JNK were measured by Western blotting. (B) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then levels of p-PI3K and p-AKT were examined by Western blotting. (C) CL1-1 cells (left) or CL1-5 cells (right) were pretreated with the indicated concentrations of the JNK inhibitor sp600125 for 6 h, and then treated with or without 5 µM β-lapachone for 24 h.Cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells.* p

    Techniques Used: Incubation, Western Blot, MTT Assay

    NQO1 siRNA transfection significantly inhibits the effect of sulindac and its metabolites on β-lapachone-induced cell death. CL1-1 cells (left) or CL1-5 cells (right) were transfected with control siRNA (−) or NQO1 siRNA (+) for 24 h, then were left untreated or were incubated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C), then 2 µM β-lapachone or medium was added and the cells incubated for 12 h, when cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: NQO1 siRNA transfection significantly inhibits the effect of sulindac and its metabolites on β-lapachone-induced cell death. CL1-1 cells (left) or CL1-5 cells (right) were transfected with control siRNA (−) or NQO1 siRNA (+) for 24 h, then were left untreated or were incubated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C), then 2 µM β-lapachone or medium was added and the cells incubated for 12 h, when cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Transfection, Incubation, Staining

    The cytotoxicity of β-lapachone for CL1-1 and CL1-5 cells is enhanced by sulindac and its metabolites. (A) CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with the indicated concentration of sulindac, sulindac sulfone, and sulindac sulfide, then 2 µM β-lapachone was added for 12 h, then cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: The cytotoxicity of β-lapachone for CL1-1 and CL1-5 cells is enhanced by sulindac and its metabolites. (A) CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with the indicated concentration of sulindac, sulindac sulfone, and sulindac sulfide, then 2 µM β-lapachone was added for 12 h, then cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Concentration Assay, Staining

    The increase in β-lapachone-induced cell death caused by sulindac and its metabolites is blocked by the NQO1 inhibitor, dicoumarol. CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C) with or without 10 µM dicoumarol, then were incubated for a further 12 h with or without addition of 2 µM β-lapachone, then cell survival was measured by crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: The increase in β-lapachone-induced cell death caused by sulindac and its metabolites is blocked by the NQO1 inhibitor, dicoumarol. CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C) with or without 10 µM dicoumarol, then were incubated for a further 12 h with or without addition of 2 µM β-lapachone, then cell survival was measured by crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Incubation, Staining

    The β-lapachone-induced apoptosis of CL1-1 and CL1-5 cells is partly due to an intracellular calcium increase. (A) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then were examined for apoptosis using Annexin V. (B) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for the indicated time, then intracellular calcium levels were measured using Fluo-4 staining and flow cytometry. The intensity of Fluo-4 staining was increased by β-lapachone treatment, especially at 1 h (arrows). (C) CL1-1 cells (left panel) or CL1-5 cells (right panel) were left untreated or were incubated for 24 h with the indicated concentration of BAPTA-AM, an intracellular calcium chelator, and/or 5 µM β-lapachone, then cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells. * p
    Figure Legend Snippet: The β-lapachone-induced apoptosis of CL1-1 and CL1-5 cells is partly due to an intracellular calcium increase. (A) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then were examined for apoptosis using Annexin V. (B) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for the indicated time, then intracellular calcium levels were measured using Fluo-4 staining and flow cytometry. The intensity of Fluo-4 staining was increased by β-lapachone treatment, especially at 1 h (arrows). (C) CL1-1 cells (left panel) or CL1-5 cells (right panel) were left untreated or were incubated for 24 h with the indicated concentration of BAPTA-AM, an intracellular calcium chelator, and/or 5 µM β-lapachone, then cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells. * p

    Techniques Used: Incubation, Staining, Flow Cytometry, Cytometry, Concentration Assay, MTT Assay

    6) Product Images from "Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells"

    Article Title: Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0088122

    Dicoumarol, an NQO1 inhibitor, blocks the apoptotic effects of β-lapachone. (A) CL1-1 cells (top) or CL1-5 cells (bottom) were left untreated or were incubated for 6 h with 5 µM β-lapachone and/or 10 µM dicoumarol, then stained with Annexin V-FITC and the Annexin V fluorescence measured by flow cytometry. (B) Morphological changes after drug treatment. CL1-1 or CL1-5 cells were left untreated (CTL) or were incubated for 24 h with 5 µM β-lapachone with or without 10 µM dicoumarol, then stained with acridine orange to observe the morphology of the cell nucleus. The scale bar represents 50 µm.
    Figure Legend Snippet: Dicoumarol, an NQO1 inhibitor, blocks the apoptotic effects of β-lapachone. (A) CL1-1 cells (top) or CL1-5 cells (bottom) were left untreated or were incubated for 6 h with 5 µM β-lapachone and/or 10 µM dicoumarol, then stained with Annexin V-FITC and the Annexin V fluorescence measured by flow cytometry. (B) Morphological changes after drug treatment. CL1-1 or CL1-5 cells were left untreated (CTL) or were incubated for 24 h with 5 µM β-lapachone with or without 10 µM dicoumarol, then stained with acridine orange to observe the morphology of the cell nucleus. The scale bar represents 50 µm.

    Techniques Used: Incubation, Staining, Fluorescence, Flow Cytometry, Cytometry, CTL Assay

    β-lapachone-induced cell death is associated with NQO1 expression levels. (A) Percentage survival of the lung cancer cell lines CL1-1, CL1-5, and A549. Cells were treated with 0–10 µM β-lapachone for 12 h, then cell viability was determined by crystal violet staining assay and expressed as a percentage of the value for cultures with no β-lapachone. (B–D) NQO1 activity levels (B), NQO1 RNA expression levels (C), and NQO1 protein expression levels (D) in the three lung cancer cell lines grown under normal culture conditions. (E) Percentage survival of A549 cells (left panel), CL1-1 cells (center panel), and CL1-5 cells (right panel) incubated with the indicated concentration of β-lapachone for 3, 6, 12, or 24 h examined by crystal violet staining and expressed as percentage survival compared to the untreated cells. The results are the mean ± SD for 3 independent experiments, each in triplicate.
    Figure Legend Snippet: β-lapachone-induced cell death is associated with NQO1 expression levels. (A) Percentage survival of the lung cancer cell lines CL1-1, CL1-5, and A549. Cells were treated with 0–10 µM β-lapachone for 12 h, then cell viability was determined by crystal violet staining assay and expressed as a percentage of the value for cultures with no β-lapachone. (B–D) NQO1 activity levels (B), NQO1 RNA expression levels (C), and NQO1 protein expression levels (D) in the three lung cancer cell lines grown under normal culture conditions. (E) Percentage survival of A549 cells (left panel), CL1-1 cells (center panel), and CL1-5 cells (right panel) incubated with the indicated concentration of β-lapachone for 3, 6, 12, or 24 h examined by crystal violet staining and expressed as percentage survival compared to the untreated cells. The results are the mean ± SD for 3 independent experiments, each in triplicate.

    Techniques Used: Expressing, Staining, Activity Assay, RNA Expression, Incubation, Concentration Assay

    Signaling pathway components involved in β-lapachone-induced apoptosis. (A) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for the indicated time, then levels of p-ERK, ERK, p-JNK, and JNK were measured by Western blotting. (B) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then levels of p-PI3K and p-AKT were examined by Western blotting. (C) CL1-1 cells (left) or CL1-5 cells (right) were pretreated with the indicated concentrations of the JNK inhibitor sp600125 for 6 h, and then treated with or without 5 µM β-lapachone for 24 h.Cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells.* p
    Figure Legend Snippet: Signaling pathway components involved in β-lapachone-induced apoptosis. (A) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for the indicated time, then levels of p-ERK, ERK, p-JNK, and JNK were measured by Western blotting. (B) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then levels of p-PI3K and p-AKT were examined by Western blotting. (C) CL1-1 cells (left) or CL1-5 cells (right) were pretreated with the indicated concentrations of the JNK inhibitor sp600125 for 6 h, and then treated with or without 5 µM β-lapachone for 24 h.Cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells.* p

    Techniques Used: Incubation, Western Blot, MTT Assay

    NQO1 siRNA transfection significantly inhibits the effect of sulindac and its metabolites on β-lapachone-induced cell death. CL1-1 cells (left) or CL1-5 cells (right) were transfected with control siRNA (−) or NQO1 siRNA (+) for 24 h, then were left untreated or were incubated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C), then 2 µM β-lapachone or medium was added and the cells incubated for 12 h, when cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: NQO1 siRNA transfection significantly inhibits the effect of sulindac and its metabolites on β-lapachone-induced cell death. CL1-1 cells (left) or CL1-5 cells (right) were transfected with control siRNA (−) or NQO1 siRNA (+) for 24 h, then were left untreated or were incubated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C), then 2 µM β-lapachone or medium was added and the cells incubated for 12 h, when cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Transfection, Incubation, Staining

    The cytotoxicity of β-lapachone for CL1-1 and CL1-5 cells is enhanced by sulindac and its metabolites. (A) CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with the indicated concentration of sulindac, sulindac sulfone, and sulindac sulfide, then 2 µM β-lapachone was added for 12 h, then cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: The cytotoxicity of β-lapachone for CL1-1 and CL1-5 cells is enhanced by sulindac and its metabolites. (A) CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with the indicated concentration of sulindac, sulindac sulfone, and sulindac sulfide, then 2 µM β-lapachone was added for 12 h, then cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Concentration Assay, Staining

    The increase in β-lapachone-induced cell death caused by sulindac and its metabolites is blocked by the NQO1 inhibitor, dicoumarol. CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C) with or without 10 µM dicoumarol, then were incubated for a further 12 h with or without addition of 2 µM β-lapachone, then cell survival was measured by crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: The increase in β-lapachone-induced cell death caused by sulindac and its metabolites is blocked by the NQO1 inhibitor, dicoumarol. CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C) with or without 10 µM dicoumarol, then were incubated for a further 12 h with or without addition of 2 µM β-lapachone, then cell survival was measured by crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Incubation, Staining

    The β-lapachone-induced apoptosis of CL1-1 and CL1-5 cells is partly due to an intracellular calcium increase. (A) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then were examined for apoptosis using Annexin V. (B) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for the indicated time, then intracellular calcium levels were measured using Fluo-4 staining and flow cytometry. The intensity of Fluo-4 staining was increased by β-lapachone treatment, especially at 1 h (arrows). (C) CL1-1 cells (left panel) or CL1-5 cells (right panel) were left untreated or were incubated for 24 h with the indicated concentration of BAPTA-AM, an intracellular calcium chelator, and/or 5 µM β-lapachone, then cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells. * p
    Figure Legend Snippet: The β-lapachone-induced apoptosis of CL1-1 and CL1-5 cells is partly due to an intracellular calcium increase. (A) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then were examined for apoptosis using Annexin V. (B) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for the indicated time, then intracellular calcium levels were measured using Fluo-4 staining and flow cytometry. The intensity of Fluo-4 staining was increased by β-lapachone treatment, especially at 1 h (arrows). (C) CL1-1 cells (left panel) or CL1-5 cells (right panel) were left untreated or were incubated for 24 h with the indicated concentration of BAPTA-AM, an intracellular calcium chelator, and/or 5 µM β-lapachone, then cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells. * p

    Techniques Used: Incubation, Staining, Flow Cytometry, Cytometry, Concentration Assay, MTT Assay

    7) Product Images from "?-Lapachone induces heart morphogenetic and functional defects by promoting the death of erythrocytes and the endocardium in zebrafish embryos"

    Article Title: ?-Lapachone induces heart morphogenetic and functional defects by promoting the death of erythrocytes and the endocardium in zebrafish embryos

    Journal: Journal of Biomedical Science

    doi: 10.1186/1423-0127-18-70

    Defects in heart-looping, valve formation, and contractile performance were detected in β-lapachone-treated embryos . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 48 and 72 hpf for nppa and cmlc2 hybridization (a-h). A Q-RT-PCR analysis indicated nppa expression levels in β-lapachone-treated 48- and 72-hpf embryos (i). B: Images of respective hearts with ventricles at either end-diastolic volume of DMSO (a), β-lapachone-treated embryo containing few erythrocytes (b), and β-lapachone-treated embryo containing no erythrocytes (c), or at end-systolic volume of DMSO (a'), β-lapachone-treated embryo containing few erythrocytes (b'), and β-lapachone-treated embryo containing no erythrocytes (c') are shown. Fractional shortening (FS) of the atrial and ventricular chamber of DMSO or β-lapachone-treated 52-hpf embryos was measured and calculated according to the formula, FS = (ED - ES)/ED × 100%, where ED is the end-diastolic diameter and ES is the end-systolic diameter of either the atrial or ventricular chambers (d). In β-lapachone-treated embryos, embryos containing few or no erythrocytes were recorded. Error bars indicate the standard error. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p
    Figure Legend Snippet: Defects in heart-looping, valve formation, and contractile performance were detected in β-lapachone-treated embryos . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 48 and 72 hpf for nppa and cmlc2 hybridization (a-h). A Q-RT-PCR analysis indicated nppa expression levels in β-lapachone-treated 48- and 72-hpf embryos (i). B: Images of respective hearts with ventricles at either end-diastolic volume of DMSO (a), β-lapachone-treated embryo containing few erythrocytes (b), and β-lapachone-treated embryo containing no erythrocytes (c), or at end-systolic volume of DMSO (a'), β-lapachone-treated embryo containing few erythrocytes (b'), and β-lapachone-treated embryo containing no erythrocytes (c') are shown. Fractional shortening (FS) of the atrial and ventricular chamber of DMSO or β-lapachone-treated 52-hpf embryos was measured and calculated according to the formula, FS = (ED - ES)/ED × 100%, where ED is the end-diastolic diameter and ES is the end-systolic diameter of either the atrial or ventricular chambers (d). In β-lapachone-treated embryos, embryos containing few or no erythrocytes were recorded. Error bars indicate the standard error. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p

    Techniques Used: Hybridization, Reverse Transcription Polymerase Chain Reaction, Expressing

    Both dicoumarol and BAPTA-AM rescued the erythrocyte-deficiency in circulation and heart-looping defect phenotypes in β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with either 0.2% DMSO (A, B), 5 μM dicoumarol (C, D), 2 μM β-lapachone (E, F), or 2 μM β-lapachone and 5 μM dicoumarol (G, H) for 4 h and fixed for nppa hybridization and o -dianisidine (ODA) staining at 48 hpf. Higher-magnification ODA-stained images of the ventral tail region are shown (B, D, F, H). (I, J) Evaluation of the rescue effects by dicoumarol or BAPTA-AM. Error bars indicate the standard error. Scale bars represent 100 μm.
    Figure Legend Snippet: Both dicoumarol and BAPTA-AM rescued the erythrocyte-deficiency in circulation and heart-looping defect phenotypes in β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with either 0.2% DMSO (A, B), 5 μM dicoumarol (C, D), 2 μM β-lapachone (E, F), or 2 μM β-lapachone and 5 μM dicoumarol (G, H) for 4 h and fixed for nppa hybridization and o -dianisidine (ODA) staining at 48 hpf. Higher-magnification ODA-stained images of the ventral tail region are shown (B, D, F, H). (I, J) Evaluation of the rescue effects by dicoumarol or BAPTA-AM. Error bars indicate the standard error. Scale bars represent 100 μm.

    Techniques Used: Hybridization, Staining

    Phenotype and circulation defect of β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and examined at 30 (A, B), 52 (C-F), 72 (G-J), and 96 hpf (K, L). Dextran rhodamine dye-injected and DMSO-treated 48-hpf embryos 2 min after dye injection (M, N), and dextran rhodamine dye-injected β-lapachone-treated 48-hpf embryos 2 min (O) and 16 min after dye injection (P) are shown. Arrowheads indicate erythrocytes. The star indicates yolk edema. Black arrows point to the linear arrangement of the heart chambers and pericardial edema, while white arrows in panels N-P indicate dye injection sites. Scale bars represent 100 μm.
    Figure Legend Snippet: Phenotype and circulation defect of β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and examined at 30 (A, B), 52 (C-F), 72 (G-J), and 96 hpf (K, L). Dextran rhodamine dye-injected and DMSO-treated 48-hpf embryos 2 min after dye injection (M, N), and dextran rhodamine dye-injected β-lapachone-treated 48-hpf embryos 2 min (O) and 16 min after dye injection (P) are shown. Arrowheads indicate erythrocytes. The star indicates yolk edema. Black arrows point to the linear arrangement of the heart chambers and pericardial edema, while white arrows in panels N-P indicate dye injection sites. Scale bars represent 100 μm.

    Techniques Used: Injection

    Induction of ROS and DNA fragmentation in erythrocytes and the endocardium by β-lapachone treatment . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 30 and 48 hpf for either hbae1 hybridization or o -dianisidine (ODA) staining. B: DMSO- and β-lapachone-treated embryos were incubated with CM-H 2 DCFDA for 1 h at 29 hpf, and both bright-field (a, c) and fluorescent (b, d-f) images under a green fluorescent protein (GFP) filter were recorded. Atrial boundary is depicted by white dotted lines. Arrows indicate flowing erythrocytes with green fluorescence from the common cardinal vein to the atrium of the heart (d-f) of β-lapachone-treated embryos. C: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h, fixed at 52 hpf, and stained with ODA. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled erythrocytes were detected in β-lapachone-treated embryos (f, h). DIC images are shown (a, e), and the inset figure in panel a shows the position of sectioning. Tail border is illustrated by white dotted lines. Arrows in panels a and e indicate ODA-stained erythrocytes. D: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h and fixed at 52 hpf. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled cells located in the endocardium were detected in β-lapachone-treated embryos (f, h, j, l). In addition, fluorescein-dUTP-labeled erythrocytes were detected in the yolk near the heart (f, j). Red dotted lines indicate borders of head and yolk while white dotted lines illustrate ventricle boundaries. * indicates erythrocytes. V, ventricle. Scale bars represent 100 μm.
    Figure Legend Snippet: Induction of ROS and DNA fragmentation in erythrocytes and the endocardium by β-lapachone treatment . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 30 and 48 hpf for either hbae1 hybridization or o -dianisidine (ODA) staining. B: DMSO- and β-lapachone-treated embryos were incubated with CM-H 2 DCFDA for 1 h at 29 hpf, and both bright-field (a, c) and fluorescent (b, d-f) images under a green fluorescent protein (GFP) filter were recorded. Atrial boundary is depicted by white dotted lines. Arrows indicate flowing erythrocytes with green fluorescence from the common cardinal vein to the atrium of the heart (d-f) of β-lapachone-treated embryos. C: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h, fixed at 52 hpf, and stained with ODA. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled erythrocytes were detected in β-lapachone-treated embryos (f, h). DIC images are shown (a, e), and the inset figure in panel a shows the position of sectioning. Tail border is illustrated by white dotted lines. Arrows in panels a and e indicate ODA-stained erythrocytes. D: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h and fixed at 52 hpf. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled cells located in the endocardium were detected in β-lapachone-treated embryos (f, h, j, l). In addition, fluorescein-dUTP-labeled erythrocytes were detected in the yolk near the heart (f, j). Red dotted lines indicate borders of head and yolk while white dotted lines illustrate ventricle boundaries. * indicates erythrocytes. V, ventricle. Scale bars represent 100 μm.

    Techniques Used: Hybridization, Staining, Incubation, Fluorescence, TUNEL Assay, Labeling

    Decreased wall shear stress was detected in β-lapachone-treated embryos . Tg( gata1 : DsRed ) was treated with DMSO or β-lapachone for 4 h at 24 h post-fertilization (hpf), and blood cell circulation was recorded at 30 hpf. A: A DIC image of 30-hpf embryos is shown. The inset figure in panel A indicates the recorded region, and DsRed-labeled erythrocytes were recorded under the TRITC mode (a-f). Images corresponding to an individual DsRed-labeled erythrocyte (arrow) at time 0 and after traveling for some distance at time t are shown for respective DMSO-treated (a, b) and 2 representative β-lapachone-treated (c, d; e, f) embryos. B: The relative wall shear stress was calculated based on τ = 4 μQ/πR 3 where μ is viscosity, Q is the flow rate, and R is the blood vessel diameter. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p
    Figure Legend Snippet: Decreased wall shear stress was detected in β-lapachone-treated embryos . Tg( gata1 : DsRed ) was treated with DMSO or β-lapachone for 4 h at 24 h post-fertilization (hpf), and blood cell circulation was recorded at 30 hpf. A: A DIC image of 30-hpf embryos is shown. The inset figure in panel A indicates the recorded region, and DsRed-labeled erythrocytes were recorded under the TRITC mode (a-f). Images corresponding to an individual DsRed-labeled erythrocyte (arrow) at time 0 and after traveling for some distance at time t are shown for respective DMSO-treated (a, b) and 2 representative β-lapachone-treated (c, d; e, f) embryos. B: The relative wall shear stress was calculated based on τ = 4 μQ/πR 3 where μ is viscosity, Q is the flow rate, and R is the blood vessel diameter. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p

    Techniques Used: Labeling, Flow Cytometry

    8) Product Images from "Gemcitabine Functions Epigenetically by Inhibiting Repair Mediated DNA Demethylation"

    Article Title: Gemcitabine Functions Epigenetically by Inhibiting Repair Mediated DNA Demethylation

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0014060

    Gemcitabine inhibits Gadd45a mediated gene activation. ( A–B ) Luciferase reporter assays of HEK293T cells transiently transfected with HpaII in vitro methylated Gal-responsive reporter, together with either Gadd45a (A) or Gal-Elk1 (B, specificity control). Cells were treated with DMSO (control, Ctrl), gemcitabine (Gem), camptothecin (Cpt), etoposide (Eto), β-lapachone (βLap), merbarone (Mer) as indicated. Shown is the fold activation by Gadd45a (A) or Gal-Elk1 (B) over control transfected cells. Error bars represent standard deviation. Significance was assessed via unpaired Student's t-test using the control sample as reference: ** = p
    Figure Legend Snippet: Gemcitabine inhibits Gadd45a mediated gene activation. ( A–B ) Luciferase reporter assays of HEK293T cells transiently transfected with HpaII in vitro methylated Gal-responsive reporter, together with either Gadd45a (A) or Gal-Elk1 (B, specificity control). Cells were treated with DMSO (control, Ctrl), gemcitabine (Gem), camptothecin (Cpt), etoposide (Eto), β-lapachone (βLap), merbarone (Mer) as indicated. Shown is the fold activation by Gadd45a (A) or Gal-Elk1 (B) over control transfected cells. Error bars represent standard deviation. Significance was assessed via unpaired Student's t-test using the control sample as reference: ** = p

    Techniques Used: Activation Assay, Luciferase, Transfection, In Vitro, Methylation, Cycling Probe Technology, Standard Deviation

    9) Product Images from "DNA breaks and chromatin structural changes enhance the transcription of autoimmune regulator target genes"

    Article Title: DNA breaks and chromatin structural changes enhance the transcription of autoimmune regulator target genes

    Journal: The Journal of Biological Chemistry

    doi: 10.1074/jbc.M116.764704

    AIRE-dependent gene expression relies on the DNA cleavage activity of both TOP2 and TOP1. A , schematic of the mechanisms of TOP1/2 inhibition by Etop, camptothecin (CPT), merbarone (Mer), and β-lapachone (β-lap). The gray ellipses and the helix represent the topoisomerase homodimer and the DNA molecule, respectively. SSB , single-stranded break; DSB , double-stranded break. B , percentage of uninduced (Ctrl) and induced ( Dox ) AIRE-Tet cells with DNA breaks after mock treatment with DMSO, TOP2-specific (Etop and Mer), and TOP1-specific inhibitors (CPT and β-lap) using Br-dUTP-labeled nicked DNA measured by FACS. The data are derived from three independent experiments. Dashed lines connect experimental groups from the same experiment. C , expression of the AIRE target genes S100A8 and CEACAM5 and the AIRE-independent gene PSMD4 after DMSO, Etop, CPT, Mer, and β-lap treatment in Ctrl and Dox AIRE-Tet cells. Log 10 -transformed data points together with their mean ± S.D. are from five independent experiments. Statistical significance was assessed by two-sample t test comparing the inhibitor treatments to the mock treatment under Ctrl and Dox conditions separately (*, p
    Figure Legend Snippet: AIRE-dependent gene expression relies on the DNA cleavage activity of both TOP2 and TOP1. A , schematic of the mechanisms of TOP1/2 inhibition by Etop, camptothecin (CPT), merbarone (Mer), and β-lapachone (β-lap). The gray ellipses and the helix represent the topoisomerase homodimer and the DNA molecule, respectively. SSB , single-stranded break; DSB , double-stranded break. B , percentage of uninduced (Ctrl) and induced ( Dox ) AIRE-Tet cells with DNA breaks after mock treatment with DMSO, TOP2-specific (Etop and Mer), and TOP1-specific inhibitors (CPT and β-lap) using Br-dUTP-labeled nicked DNA measured by FACS. The data are derived from three independent experiments. Dashed lines connect experimental groups from the same experiment. C , expression of the AIRE target genes S100A8 and CEACAM5 and the AIRE-independent gene PSMD4 after DMSO, Etop, CPT, Mer, and β-lap treatment in Ctrl and Dox AIRE-Tet cells. Log 10 -transformed data points together with their mean ± S.D. are from five independent experiments. Statistical significance was assessed by two-sample t test comparing the inhibitor treatments to the mock treatment under Ctrl and Dox conditions separately (*, p

    Techniques Used: Expressing, Activity Assay, Inhibition, Cycling Probe Technology, Labeling, FACS, Derivative Assay, Transformation Assay

    10) Product Images from "An NQO1 Substrate with Potent Antitumor Activity That Selectively Kills by PARP1-Induced Programmed Necrosis"

    Article Title: An NQO1 Substrate with Potent Antitumor Activity That Selectively Kills by PARP1-Induced Programmed Necrosis

    Journal: Cancer research

    doi: 10.1158/0008-5472.CAN-11-3135

    Deoxynyboquinone shows equivalent efficacy to β-lapachone at a 6-fold lower dose. In A–C, mice bearing visible tumors were treated once every other day for 5 injections as in the text, and average wet weight tumor volumes assessed (A and B). Tumor nodules were confirmed visually (A) and histologically (not shown), and average wet weights calculated (B). DNQ-HPβCD (5 or 10 mg/kg) or β-Lap-HPβCD (30 mg/kg) caused tumor weight decreases that were equivalent ( P > 0.6). Normal tissues were assessed with no long-term pathologic injury noted; see H E-stained livers (C). D, Kaplan-Meier survival curves showed significant ( P ≤ 0.04) survival advantages of deoxynyboquinone (5 mg/kg) or β-lapachone-treated (30 mg/kg) groups. Groups were HPβCD alone (1,000 mg/kg), deoxynyboquinone (2.5, 5.0 mg/kg), or β-lapachone (30 mg/kg) in HPβCD. *, P ≤ 0.04; **, P ≤ 0.01. In E and F, PAR formation and ATP loss confirmed programmed necrotic mechanism of deoxynyboquinone- or β-lapachone–treated LLC tumors where animals were treated as in A–C with 3 doses and 24 hours later lungs were removed. Note the lack of response of associated normal lung tissue. **, P
    Figure Legend Snippet: Deoxynyboquinone shows equivalent efficacy to β-lapachone at a 6-fold lower dose. In A–C, mice bearing visible tumors were treated once every other day for 5 injections as in the text, and average wet weight tumor volumes assessed (A and B). Tumor nodules were confirmed visually (A) and histologically (not shown), and average wet weights calculated (B). DNQ-HPβCD (5 or 10 mg/kg) or β-Lap-HPβCD (30 mg/kg) caused tumor weight decreases that were equivalent ( P > 0.6). Normal tissues were assessed with no long-term pathologic injury noted; see H E-stained livers (C). D, Kaplan-Meier survival curves showed significant ( P ≤ 0.04) survival advantages of deoxynyboquinone (5 mg/kg) or β-lapachone-treated (30 mg/kg) groups. Groups were HPβCD alone (1,000 mg/kg), deoxynyboquinone (2.5, 5.0 mg/kg), or β-lapachone (30 mg/kg) in HPβCD. *, P ≤ 0.04; **, P ≤ 0.01. In E and F, PAR formation and ATP loss confirmed programmed necrotic mechanism of deoxynyboquinone- or β-lapachone–treated LLC tumors where animals were treated as in A–C with 3 doses and 24 hours later lungs were removed. Note the lack of response of associated normal lung tissue. **, P

    Techniques Used: Mouse Assay, Staining

    Elevated ROS in deoxynyboquinone-treated endogenous NQO1 + ) measured by staining intensity from ≥100 cells using NIH Image J. F, deoxynyboquinone (µmol/L, 2 hours) -exposed A549 cells were cotreated with ± catalase (CAT, 1,000 U) and clonogenic survival monitored. ***, P ≤ 0.001, comparing deoxynyboquinone treated A549 cells ± catalase cotreatment. Dic, dicoumarol; DNQ, deoxynyboquinone; β-lap, β-lapachone.
    Figure Legend Snippet: Elevated ROS in deoxynyboquinone-treated endogenous NQO1 + ) measured by staining intensity from ≥100 cells using NIH Image J. F, deoxynyboquinone (µmol/L, 2 hours) -exposed A549 cells were cotreated with ± catalase (CAT, 1,000 U) and clonogenic survival monitored. ***, P ≤ 0.001, comparing deoxynyboquinone treated A549 cells ± catalase cotreatment. Dic, dicoumarol; DNQ, deoxynyboquinone; β-lap, β-lapachone.

    Techniques Used: Staining

    Deoxynyboquinone induced lethality in endogenous NQO1 + cells. A, structures of deoxynyboquinone, β-lapachone, and menadione. B, endogenous, NQO1 overexpressing A549 or MCF-7 cells were treated with or without various deoxynyboquinone doses (µmol/L, 2 hours), ± dicoumarol (40 µmol/L, 2 hours). Lethality was monitored by relative survival. Data are means, ±SE for sextuplets carried out thrice. C, cells from B were treated with or without various β-lapachone doses (µmol/L, 2 hours), ± dicoumarol. D, cells were treated with or without menadione (µmol/L, 2 hours), ± dicoumarol as in B. Dicoumarol potentiated menadione toxicity in MCF-7, but not in A549, cells. Control cells in B–D were treated with identical DMSO concentrations (
    Figure Legend Snippet: Deoxynyboquinone induced lethality in endogenous NQO1 + cells. A, structures of deoxynyboquinone, β-lapachone, and menadione. B, endogenous, NQO1 overexpressing A549 or MCF-7 cells were treated with or without various deoxynyboquinone doses (µmol/L, 2 hours), ± dicoumarol (40 µmol/L, 2 hours). Lethality was monitored by relative survival. Data are means, ±SE for sextuplets carried out thrice. C, cells from B were treated with or without various β-lapachone doses (µmol/L, 2 hours), ± dicoumarol. D, cells were treated with or without menadione (µmol/L, 2 hours), ± dicoumarol as in B. Dicoumarol potentiated menadione toxicity in MCF-7, but not in A549, cells. Control cells in B–D were treated with identical DMSO concentrations (

    Techniques Used:

    11) Product Images from "?-Lapachone induces heart morphogenetic and functional defects by promoting the death of erythrocytes and the endocardium in zebrafish embryos"

    Article Title: ?-Lapachone induces heart morphogenetic and functional defects by promoting the death of erythrocytes and the endocardium in zebrafish embryos

    Journal: Journal of Biomedical Science

    doi: 10.1186/1423-0127-18-70

    Defects in heart-looping, valve formation, and contractile performance were detected in β-lapachone-treated embryos . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 48 and 72 hpf for nppa and cmlc2 hybridization (a-h). A Q-RT-PCR analysis indicated nppa expression levels in β-lapachone-treated 48- and 72-hpf embryos (i). B: Images of respective hearts with ventricles at either end-diastolic volume of DMSO (a), β-lapachone-treated embryo containing few erythrocytes (b), and β-lapachone-treated embryo containing no erythrocytes (c), or at end-systolic volume of DMSO (a'), β-lapachone-treated embryo containing few erythrocytes (b'), and β-lapachone-treated embryo containing no erythrocytes (c') are shown. Fractional shortening (FS) of the atrial and ventricular chamber of DMSO or β-lapachone-treated 52-hpf embryos was measured and calculated according to the formula, FS = (ED - ES)/ED × 100%, where ED is the end-diastolic diameter and ES is the end-systolic diameter of either the atrial or ventricular chambers (d). In β-lapachone-treated embryos, embryos containing few or no erythrocytes were recorded. Error bars indicate the standard error. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p
    Figure Legend Snippet: Defects in heart-looping, valve formation, and contractile performance were detected in β-lapachone-treated embryos . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 48 and 72 hpf for nppa and cmlc2 hybridization (a-h). A Q-RT-PCR analysis indicated nppa expression levels in β-lapachone-treated 48- and 72-hpf embryos (i). B: Images of respective hearts with ventricles at either end-diastolic volume of DMSO (a), β-lapachone-treated embryo containing few erythrocytes (b), and β-lapachone-treated embryo containing no erythrocytes (c), or at end-systolic volume of DMSO (a'), β-lapachone-treated embryo containing few erythrocytes (b'), and β-lapachone-treated embryo containing no erythrocytes (c') are shown. Fractional shortening (FS) of the atrial and ventricular chamber of DMSO or β-lapachone-treated 52-hpf embryos was measured and calculated according to the formula, FS = (ED - ES)/ED × 100%, where ED is the end-diastolic diameter and ES is the end-systolic diameter of either the atrial or ventricular chambers (d). In β-lapachone-treated embryos, embryos containing few or no erythrocytes were recorded. Error bars indicate the standard error. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p

    Techniques Used: Hybridization, Reverse Transcription Polymerase Chain Reaction, Expressing

    Both dicoumarol and BAPTA-AM rescued the erythrocyte-deficiency in circulation and heart-looping defect phenotypes in β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with either 0.2% DMSO (A, B), 5 μM dicoumarol (C, D), 2 μM β-lapachone (E, F), or 2 μM β-lapachone and 5 μM dicoumarol (G, H) for 4 h and fixed for nppa hybridization and o -dianisidine (ODA) staining at 48 hpf. Higher-magnification ODA-stained images of the ventral tail region are shown (B, D, F, H). (I, J) Evaluation of the rescue effects by dicoumarol or BAPTA-AM. Error bars indicate the standard error. Scale bars represent 100 μm.
    Figure Legend Snippet: Both dicoumarol and BAPTA-AM rescued the erythrocyte-deficiency in circulation and heart-looping defect phenotypes in β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with either 0.2% DMSO (A, B), 5 μM dicoumarol (C, D), 2 μM β-lapachone (E, F), or 2 μM β-lapachone and 5 μM dicoumarol (G, H) for 4 h and fixed for nppa hybridization and o -dianisidine (ODA) staining at 48 hpf. Higher-magnification ODA-stained images of the ventral tail region are shown (B, D, F, H). (I, J) Evaluation of the rescue effects by dicoumarol or BAPTA-AM. Error bars indicate the standard error. Scale bars represent 100 μm.

    Techniques Used: Hybridization, Staining

    Phenotype and circulation defect of β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and examined at 30 (A, B), 52 (C-F), 72 (G-J), and 96 hpf (K, L). Dextran rhodamine dye-injected and DMSO-treated 48-hpf embryos 2 min after dye injection (M, N), and dextran rhodamine dye-injected β-lapachone-treated 48-hpf embryos 2 min (O) and 16 min after dye injection (P) are shown. Arrowheads indicate erythrocytes. The star indicates yolk edema. Black arrows point to the linear arrangement of the heart chambers and pericardial edema, while white arrows in panels N-P indicate dye injection sites. Scale bars represent 100 μm.
    Figure Legend Snippet: Phenotype and circulation defect of β-lapachone-treated embryos . Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and examined at 30 (A, B), 52 (C-F), 72 (G-J), and 96 hpf (K, L). Dextran rhodamine dye-injected and DMSO-treated 48-hpf embryos 2 min after dye injection (M, N), and dextran rhodamine dye-injected β-lapachone-treated 48-hpf embryos 2 min (O) and 16 min after dye injection (P) are shown. Arrowheads indicate erythrocytes. The star indicates yolk edema. Black arrows point to the linear arrangement of the heart chambers and pericardial edema, while white arrows in panels N-P indicate dye injection sites. Scale bars represent 100 μm.

    Techniques Used: Injection

    Induction of ROS and DNA fragmentation in erythrocytes and the endocardium by β-lapachone treatment . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 30 and 48 hpf for either hbae1 hybridization or o -dianisidine (ODA) staining. B: DMSO- and β-lapachone-treated embryos were incubated with CM-H 2 DCFDA for 1 h at 29 hpf, and both bright-field (a, c) and fluorescent (b, d-f) images under a green fluorescent protein (GFP) filter were recorded. Atrial boundary is depicted by white dotted lines. Arrows indicate flowing erythrocytes with green fluorescence from the common cardinal vein to the atrium of the heart (d-f) of β-lapachone-treated embryos. C: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h, fixed at 52 hpf, and stained with ODA. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled erythrocytes were detected in β-lapachone-treated embryos (f, h). DIC images are shown (a, e), and the inset figure in panel a shows the position of sectioning. Tail border is illustrated by white dotted lines. Arrows in panels a and e indicate ODA-stained erythrocytes. D: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h and fixed at 52 hpf. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled cells located in the endocardium were detected in β-lapachone-treated embryos (f, h, j, l). In addition, fluorescein-dUTP-labeled erythrocytes were detected in the yolk near the heart (f, j). Red dotted lines indicate borders of head and yolk while white dotted lines illustrate ventricle boundaries. * indicates erythrocytes. V, ventricle. Scale bars represent 100 μm.
    Figure Legend Snippet: Induction of ROS and DNA fragmentation in erythrocytes and the endocardium by β-lapachone treatment . A: Embryos at 24 hours post-fertilization (hpf) were treated with DMSO or β-lapachone for 4 h and fixed at 30 and 48 hpf for either hbae1 hybridization or o -dianisidine (ODA) staining. B: DMSO- and β-lapachone-treated embryos were incubated with CM-H 2 DCFDA for 1 h at 29 hpf, and both bright-field (a, c) and fluorescent (b, d-f) images under a green fluorescent protein (GFP) filter were recorded. Atrial boundary is depicted by white dotted lines. Arrows indicate flowing erythrocytes with green fluorescence from the common cardinal vein to the atrium of the heart (d-f) of β-lapachone-treated embryos. C: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h, fixed at 52 hpf, and stained with ODA. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled erythrocytes were detected in β-lapachone-treated embryos (f, h). DIC images are shown (a, e), and the inset figure in panel a shows the position of sectioning. Tail border is illustrated by white dotted lines. Arrows in panels a and e indicate ODA-stained erythrocytes. D: Embryos were treated with DMSO or β-lapachone at 48 hpf for 4 h and fixed at 52 hpf. After paraffin sectioning, TUNEL reactions were conducted, and fluorescein-dUTP-labeled cells located in the endocardium were detected in β-lapachone-treated embryos (f, h, j, l). In addition, fluorescein-dUTP-labeled erythrocytes were detected in the yolk near the heart (f, j). Red dotted lines indicate borders of head and yolk while white dotted lines illustrate ventricle boundaries. * indicates erythrocytes. V, ventricle. Scale bars represent 100 μm.

    Techniques Used: Hybridization, Staining, Incubation, Fluorescence, TUNEL Assay, Labeling

    Decreased wall shear stress was detected in β-lapachone-treated embryos . Tg( gata1 : DsRed ) was treated with DMSO or β-lapachone for 4 h at 24 h post-fertilization (hpf), and blood cell circulation was recorded at 30 hpf. A: A DIC image of 30-hpf embryos is shown. The inset figure in panel A indicates the recorded region, and DsRed-labeled erythrocytes were recorded under the TRITC mode (a-f). Images corresponding to an individual DsRed-labeled erythrocyte (arrow) at time 0 and after traveling for some distance at time t are shown for respective DMSO-treated (a, b) and 2 representative β-lapachone-treated (c, d; e, f) embryos. B: The relative wall shear stress was calculated based on τ = 4 μQ/πR 3 where μ is viscosity, Q is the flow rate, and R is the blood vessel diameter. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p
    Figure Legend Snippet: Decreased wall shear stress was detected in β-lapachone-treated embryos . Tg( gata1 : DsRed ) was treated with DMSO or β-lapachone for 4 h at 24 h post-fertilization (hpf), and blood cell circulation was recorded at 30 hpf. A: A DIC image of 30-hpf embryos is shown. The inset figure in panel A indicates the recorded region, and DsRed-labeled erythrocytes were recorded under the TRITC mode (a-f). Images corresponding to an individual DsRed-labeled erythrocyte (arrow) at time 0 and after traveling for some distance at time t are shown for respective DMSO-treated (a, b) and 2 representative β-lapachone-treated (c, d; e, f) embryos. B: The relative wall shear stress was calculated based on τ = 4 μQ/πR 3 where μ is viscosity, Q is the flow rate, and R is the blood vessel diameter. Student's t -test was used to compare DMSO- and β-lapachone-treated embryos. * p

    Techniques Used: Labeling, Flow Cytometry

    12) Product Images from "The Tumor-Selective Cytotoxic Agent ?-Lapachone is a Potent Inhibitor of IDO1"

    Article Title: The Tumor-Selective Cytotoxic Agent ?-Lapachone is a Potent Inhibitor of IDO1

    Journal: International Journal of Tryptophan Research : IJTR

    doi: 10.4137/IJTR.S12094

    β-Lapachone inhibits IDO1 activity through uncompetitive inhibition. Enzyme assays with purified recombinant huIDO1 were performed in the presence of varying concentrations of L-Trp and β-lapachone and the concentration of the product kynurenine was measured at 15 minutes. ( A ) Michaelis-Menton nonlinear regression plot of substrate-velocity curves at the five different inhibitor concentrations. These data best fit an uncompetitive inhibition model with a global (shared) R 2 value of 0.96 and a K i of 450 nM. ( B ) Corresponding Lineweaver-Burk plot. These plots show decreasing K m and V max as the inhibitor concentration is increased consistent with uncompetitive inhibition.
    Figure Legend Snippet: β-Lapachone inhibits IDO1 activity through uncompetitive inhibition. Enzyme assays with purified recombinant huIDO1 were performed in the presence of varying concentrations of L-Trp and β-lapachone and the concentration of the product kynurenine was measured at 15 minutes. ( A ) Michaelis-Menton nonlinear regression plot of substrate-velocity curves at the five different inhibitor concentrations. These data best fit an uncompetitive inhibition model with a global (shared) R 2 value of 0.96 and a K i of 450 nM. ( B ) Corresponding Lineweaver-Burk plot. These plots show decreasing K m and V max as the inhibitor concentration is increased consistent with uncompetitive inhibition.

    Techniques Used: Activity Assay, Inhibition, Purification, Recombinant, Concentration Assay

    Inhibition of human IDO1 enzyme activity by β-lapachone. Dose-response assessment of increasing β-lapachone concentration on the activity of purified recombinant huIDO1. The assay was performed with 100 μM L-Trp substrate and the concentration of the product kynurenine was measured at 15 minutes while the enzyme was still active in the log phase. The data are plotted as percent inhibition of kynurenine production. The IC 50 of 440 nM was determined by nonlinear curve fitting.
    Figure Legend Snippet: Inhibition of human IDO1 enzyme activity by β-lapachone. Dose-response assessment of increasing β-lapachone concentration on the activity of purified recombinant huIDO1. The assay was performed with 100 μM L-Trp substrate and the concentration of the product kynurenine was measured at 15 minutes while the enzyme was still active in the log phase. The data are plotted as percent inhibition of kynurenine production. The IC 50 of 440 nM was determined by nonlinear curve fitting.

    Techniques Used: Inhibition, Activity Assay, Concentration Assay, Purification, Recombinant

    Inhibition of intracellular IDO1 activity by β-lapachone. ( A ) Dose-response assessment of increasing β-lapachone concentration on IDO1 activity in HeLa cells following 24 hrs of induction with IFNγ (100 ng/mL). Data are plotted as percent inhibition of kynurenine production with an IC 50 of 1.0 μM determined by nonlinear curve fitting. ( B ) SRB-based evaluation of viable cell numbers at the conclusion of the β-lapachone dose response assay shown in ( A ). (C) Western blot analysis of IDO1 protein in whole cell lysates from HeLa cells induced with IFNγ (100 ng/mL) and treated with β-lapachone (5 μM) as indicated. ( D ) β-lapachone dose-response assessment performed in parallel with ( A ) with the addition of the NQO1 inhibitor dicoumarol (50 μM). Data are plotted as percent inhibition of kynurenine production with an IC 50 of 4.2 μM determined by nonlinear curve fitting.
    Figure Legend Snippet: Inhibition of intracellular IDO1 activity by β-lapachone. ( A ) Dose-response assessment of increasing β-lapachone concentration on IDO1 activity in HeLa cells following 24 hrs of induction with IFNγ (100 ng/mL). Data are plotted as percent inhibition of kynurenine production with an IC 50 of 1.0 μM determined by nonlinear curve fitting. ( B ) SRB-based evaluation of viable cell numbers at the conclusion of the β-lapachone dose response assay shown in ( A ). (C) Western blot analysis of IDO1 protein in whole cell lysates from HeLa cells induced with IFNγ (100 ng/mL) and treated with β-lapachone (5 μM) as indicated. ( D ) β-lapachone dose-response assessment performed in parallel with ( A ) with the addition of the NQO1 inhibitor dicoumarol (50 μM). Data are plotted as percent inhibition of kynurenine production with an IC 50 of 4.2 μM determined by nonlinear curve fitting.

    Techniques Used: Inhibition, Activity Assay, Concentration Assay, Sulforhodamine B Assay, Western Blot

    β-Lapachone is predicted to bind in the IDO1 active site. Computational modeling at the IDO1 active site and IC 50 values for inhibition of purified human recombinant IDO1 for ( A ) dehydro-α-lapachone (2,2-dimethyl-2H-benzo[g]chromene-5,10-dione) and ( B ) β-lapachone (3,4-dihydro-2,2-dimethyl-2 H -naphthol[1,2- b ]pyran-5,6-dione).
    Figure Legend Snippet: β-Lapachone is predicted to bind in the IDO1 active site. Computational modeling at the IDO1 active site and IC 50 values for inhibition of purified human recombinant IDO1 for ( A ) dehydro-α-lapachone (2,2-dimethyl-2H-benzo[g]chromene-5,10-dione) and ( B ) β-lapachone (3,4-dihydro-2,2-dimethyl-2 H -naphthol[1,2- b ]pyran-5,6-dione).

    Techniques Used: Inhibition, Purification, Recombinant

    13) Product Images from "Inactivation of human DGAT2 by oxidative stress on cysteine residues"

    Article Title: Inactivation of human DGAT2 by oxidative stress on cysteine residues

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0181076

    Multimeric complex of human DGAT2 formed by ROS-induced intermolecular disulfide crosslinking in vitro . Membrane extracts from human DGAT2-overexpressing Sf9 insect cells were treated with H 2 O 2 (A) or β-lapachone (B) in the presence or absence of 20 mM DTT and subjected to Western blot analysis using anti-DGAT2 antibody. The amount of monomeric human DGAT2 proteins presented as redDGAT2 in (A) and (B) was quantified and the amount of relative redDGAT2 protein was calculated by setting the values from samples treated with PBS (C) or DMSO (D) to 100%. Asterisk indicates a non-specific band.
    Figure Legend Snippet: Multimeric complex of human DGAT2 formed by ROS-induced intermolecular disulfide crosslinking in vitro . Membrane extracts from human DGAT2-overexpressing Sf9 insect cells were treated with H 2 O 2 (A) or β-lapachone (B) in the presence or absence of 20 mM DTT and subjected to Western blot analysis using anti-DGAT2 antibody. The amount of monomeric human DGAT2 proteins presented as redDGAT2 in (A) and (B) was quantified and the amount of relative redDGAT2 protein was calculated by setting the values from samples treated with PBS (C) or DMSO (D) to 100%. Asterisk indicates a non-specific band.

    Techniques Used: In Vitro, Western Blot

    Inhibitory effect of ROS and ROS generator on human DGAT2 catalytic activity. Membrane extracts from human DGAT2-overexpressing Sf9 insect cells were treated with indicated concentrations of H 2 O 2 (A) or β-lapachone (B) in the presence or absence of 20 mM DTT. Human DGAT2 activity was measured by using the conventional extraction-based in vitro assays which are described in detail in the Materials and Methods section. The activities of membrane extracts treated with PBS (instead of H 2 O 2 ) or DMSO (instead of β-lapachone) in the absence of DTT were defined as 100%. The mean values and standard deviations were determined from four independent experiments.
    Figure Legend Snippet: Inhibitory effect of ROS and ROS generator on human DGAT2 catalytic activity. Membrane extracts from human DGAT2-overexpressing Sf9 insect cells were treated with indicated concentrations of H 2 O 2 (A) or β-lapachone (B) in the presence or absence of 20 mM DTT. Human DGAT2 activity was measured by using the conventional extraction-based in vitro assays which are described in detail in the Materials and Methods section. The activities of membrane extracts treated with PBS (instead of H 2 O 2 ) or DMSO (instead of β-lapachone) in the absence of DTT were defined as 100%. The mean values and standard deviations were determined from four independent experiments.

    Techniques Used: Activity Assay, In Vitro

    14) Product Images from "Potential mechanism of phytochemical-induced apoptosis in human prostate adenocarcinoma cells: Therapeutic synergy in genistein and ?-lapachone combination treatment"

    Article Title: Potential mechanism of phytochemical-induced apoptosis in human prostate adenocarcinoma cells: Therapeutic synergy in genistein and ?-lapachone combination treatment

    Journal: Cancer Cell International

    doi: 10.1186/1475-2867-4-5

    CPP32 is the major pathway in genistein-induced apoptosis in PC3 cells. PC3 cells (2.5 × 10 3 cells/well) were cultured in 48-well culture plates; treated with/without 100 μM caspase inhibitor (zVAD-fmk) for 2 hr; then with 1–8 μM β-Lapachone (bLap) for 4 hr as described in the methods. Cells were then analyzed for caspase (CPP32) activity and corresponding apoptosis. Data pointsare the means ± SEM of two independent experiments performed in triplicates
    Figure Legend Snippet: CPP32 is the major pathway in genistein-induced apoptosis in PC3 cells. PC3 cells (2.5 × 10 3 cells/well) were cultured in 48-well culture plates; treated with/without 100 μM caspase inhibitor (zVAD-fmk) for 2 hr; then with 1–8 μM β-Lapachone (bLap) for 4 hr as described in the methods. Cells were then analyzed for caspase (CPP32) activity and corresponding apoptosis. Data pointsare the means ± SEM of two independent experiments performed in triplicates

    Techniques Used: Cell Culture, Activity Assay

    Genistein (Gn)/β-Lapachone combination treatment of PC3. Cells were treated as described in the methods and subjected to post-treatment viability with MTS colorimetric assay. Data points represent the means ± SEM of three independent experiments performed in triplicates.
    Figure Legend Snippet: Genistein (Gn)/β-Lapachone combination treatment of PC3. Cells were treated as described in the methods and subjected to post-treatment viability with MTS colorimetric assay. Data points represent the means ± SEM of three independent experiments performed in triplicates.

    Techniques Used: Colorimetric Assay

    Single and combination of PC3 cells with genistein (Gn) and β-lapachone (bLap) βLap. Briefly, PC3 cells were seeded at 1 × 10 4 cells/well in 48-well MTP and co-cultured with Gn 0-70 with/without bLap (1.2 μM); followed by determination of treatment-induced cytotoxicity as described in the methods. Data points represent means ± SEM of three experiments performed in triplicates
    Figure Legend Snippet: Single and combination of PC3 cells with genistein (Gn) and β-lapachone (bLap) βLap. Briefly, PC3 cells were seeded at 1 × 10 4 cells/well in 48-well MTP and co-cultured with Gn 0-70 with/without bLap (1.2 μM); followed by determination of treatment-induced cytotoxicity as described in the methods. Data points represent means ± SEM of three experiments performed in triplicates

    Techniques Used: Cell Culture

    β-lapachone-induced growth inhibition in PC3. PC3 cells (1 × 10 4 cells/well) were cultured in 24-well plates for 48 hr to allow 85–90% confluence; treated with varyingconcentrations of bLap and assessed for post-treatment viabilitywith the MTS assay. Note the dose-dependent growth inhibitionin PC3. Data points represent means ± SEM of three independentexperiments performed in triplicates
    Figure Legend Snippet: β-lapachone-induced growth inhibition in PC3. PC3 cells (1 × 10 4 cells/well) were cultured in 24-well plates for 48 hr to allow 85–90% confluence; treated with varyingconcentrations of bLap and assessed for post-treatment viabilitywith the MTS assay. Note the dose-dependent growth inhibitionin PC3. Data points represent means ± SEM of three independentexperiments performed in triplicates

    Techniques Used: Inhibition, Cell Culture, MTS Assay

    15) Product Images from "Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells"

    Article Title: Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells

    Journal: PLoS ONE

    doi: 10.1371/journal.pone.0088122

    Dicoumarol, an NQO1 inhibitor, blocks the apoptotic effects of β-lapachone. (A) CL1-1 cells (top) or CL1-5 cells (bottom) were left untreated or were incubated for 6 h with 5 µM β-lapachone and/or 10 µM dicoumarol, then stained with Annexin V-FITC and the Annexin V fluorescence measured by flow cytometry. (B) Morphological changes after drug treatment. CL1-1 or CL1-5 cells were left untreated (CTL) or were incubated for 24 h with 5 µM β-lapachone with or without 10 µM dicoumarol, then stained with acridine orange to observe the morphology of the cell nucleus. The scale bar represents 50 µm.
    Figure Legend Snippet: Dicoumarol, an NQO1 inhibitor, blocks the apoptotic effects of β-lapachone. (A) CL1-1 cells (top) or CL1-5 cells (bottom) were left untreated or were incubated for 6 h with 5 µM β-lapachone and/or 10 µM dicoumarol, then stained with Annexin V-FITC and the Annexin V fluorescence measured by flow cytometry. (B) Morphological changes after drug treatment. CL1-1 or CL1-5 cells were left untreated (CTL) or were incubated for 24 h with 5 µM β-lapachone with or without 10 µM dicoumarol, then stained with acridine orange to observe the morphology of the cell nucleus. The scale bar represents 50 µm.

    Techniques Used: Incubation, Staining, Fluorescence, Flow Cytometry, Cytometry, CTL Assay

    β-lapachone-induced cell death is associated with NQO1 expression levels. (A) Percentage survival of the lung cancer cell lines CL1-1, CL1-5, and A549. Cells were treated with 0–10 µM β-lapachone for 12 h, then cell viability was determined by crystal violet staining assay and expressed as a percentage of the value for cultures with no β-lapachone. (B–D) NQO1 activity levels (B), NQO1 RNA expression levels (C), and NQO1 protein expression levels (D) in the three lung cancer cell lines grown under normal culture conditions. (E) Percentage survival of A549 cells (left panel), CL1-1 cells (center panel), and CL1-5 cells (right panel) incubated with the indicated concentration of β-lapachone for 3, 6, 12, or 24 h examined by crystal violet staining and expressed as percentage survival compared to the untreated cells. The results are the mean ± SD for 3 independent experiments, each in triplicate.
    Figure Legend Snippet: β-lapachone-induced cell death is associated with NQO1 expression levels. (A) Percentage survival of the lung cancer cell lines CL1-1, CL1-5, and A549. Cells were treated with 0–10 µM β-lapachone for 12 h, then cell viability was determined by crystal violet staining assay and expressed as a percentage of the value for cultures with no β-lapachone. (B–D) NQO1 activity levels (B), NQO1 RNA expression levels (C), and NQO1 protein expression levels (D) in the three lung cancer cell lines grown under normal culture conditions. (E) Percentage survival of A549 cells (left panel), CL1-1 cells (center panel), and CL1-5 cells (right panel) incubated with the indicated concentration of β-lapachone for 3, 6, 12, or 24 h examined by crystal violet staining and expressed as percentage survival compared to the untreated cells. The results are the mean ± SD for 3 independent experiments, each in triplicate.

    Techniques Used: Expressing, Staining, Activity Assay, RNA Expression, Incubation, Concentration Assay

    Signaling pathway components involved in β-lapachone-induced apoptosis. (A) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for the indicated time, then levels of p-ERK, ERK, p-JNK, and JNK were measured by Western blotting. (B) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then levels of p-PI3K and p-AKT were examined by Western blotting. (C) CL1-1 cells (left) or CL1-5 cells (right) were pretreated with the indicated concentrations of the JNK inhibitor sp600125 for 6 h, and then treated with or without 5 µM β-lapachone for 24 h.Cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells.* p
    Figure Legend Snippet: Signaling pathway components involved in β-lapachone-induced apoptosis. (A) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for the indicated time, then levels of p-ERK, ERK, p-JNK, and JNK were measured by Western blotting. (B) CL1-1 cells (left) or CL1-5 cells (right) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then levels of p-PI3K and p-AKT were examined by Western blotting. (C) CL1-1 cells (left) or CL1-5 cells (right) were pretreated with the indicated concentrations of the JNK inhibitor sp600125 for 6 h, and then treated with or without 5 µM β-lapachone for 24 h.Cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells.* p

    Techniques Used: Incubation, Western Blot, MTT Assay

    NQO1 siRNA transfection significantly inhibits the effect of sulindac and its metabolites on β-lapachone-induced cell death. CL1-1 cells (left) or CL1-5 cells (right) were transfected with control siRNA (−) or NQO1 siRNA (+) for 24 h, then were left untreated or were incubated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C), then 2 µM β-lapachone or medium was added and the cells incubated for 12 h, when cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: NQO1 siRNA transfection significantly inhibits the effect of sulindac and its metabolites on β-lapachone-induced cell death. CL1-1 cells (left) or CL1-5 cells (right) were transfected with control siRNA (−) or NQO1 siRNA (+) for 24 h, then were left untreated or were incubated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C), then 2 µM β-lapachone or medium was added and the cells incubated for 12 h, when cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Transfection, Incubation, Staining

    The cytotoxicity of β-lapachone for CL1-1 and CL1-5 cells is enhanced by sulindac and its metabolites. (A) CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with the indicated concentration of sulindac, sulindac sulfone, and sulindac sulfide, then 2 µM β-lapachone was added for 12 h, then cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: The cytotoxicity of β-lapachone for CL1-1 and CL1-5 cells is enhanced by sulindac and its metabolites. (A) CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with the indicated concentration of sulindac, sulindac sulfone, and sulindac sulfide, then 2 µM β-lapachone was added for 12 h, then cell survival was measured using crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Concentration Assay, Staining

    The increase in β-lapachone-induced cell death caused by sulindac and its metabolites is blocked by the NQO1 inhibitor, dicoumarol. CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C) with or without 10 µM dicoumarol, then were incubated for a further 12 h with or without addition of 2 µM β-lapachone, then cell survival was measured by crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p
    Figure Legend Snippet: The increase in β-lapachone-induced cell death caused by sulindac and its metabolites is blocked by the NQO1 inhibitor, dicoumarol. CL1-1 cells (left) or CL1-5 cells (right) were left untreated or were pretreated for 6 h with 100 or 250 µM sulindac (A), sulindac sulfone (B), or sulindac sulfide (C) with or without 10 µM dicoumarol, then were incubated for a further 12 h with or without addition of 2 µM β-lapachone, then cell survival was measured by crystal violet staining and expressed as percentage survival compared to the untreated cells. * : p

    Techniques Used: Incubation, Staining

    The β-lapachone-induced apoptosis of CL1-1 and CL1-5 cells is partly due to an intracellular calcium increase. (A) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then were examined for apoptosis using Annexin V. (B) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for the indicated time, then intracellular calcium levels were measured using Fluo-4 staining and flow cytometry. The intensity of Fluo-4 staining was increased by β-lapachone treatment, especially at 1 h (arrows). (C) CL1-1 cells (left panel) or CL1-5 cells (right panel) were left untreated or were incubated for 24 h with the indicated concentration of BAPTA-AM, an intracellular calcium chelator, and/or 5 µM β-lapachone, then cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells. * p
    Figure Legend Snippet: The β-lapachone-induced apoptosis of CL1-1 and CL1-5 cells is partly due to an intracellular calcium increase. (A) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for 0, 3, 6, or 9 h, then were examined for apoptosis using Annexin V. (B) CL1-1 cells (left panel) or CL1-5 cells (right panel) were incubated with 5 µM β-lapachone for the indicated time, then intracellular calcium levels were measured using Fluo-4 staining and flow cytometry. The intensity of Fluo-4 staining was increased by β-lapachone treatment, especially at 1 h (arrows). (C) CL1-1 cells (left panel) or CL1-5 cells (right panel) were left untreated or were incubated for 24 h with the indicated concentration of BAPTA-AM, an intracellular calcium chelator, and/or 5 µM β-lapachone, then cell survival was measured by the MTT assay and expressed as percentage survival compared to the untreated cells. * p

    Techniques Used: Incubation, Staining, Flow Cytometry, Cytometry, Concentration Assay, MTT Assay

    Related Articles

    Cell Culture:

    Article Title: Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells
    Article Snippet: .. Acridine Orange (AO) Staining Cells (5x104 ) cultured on cover-slides in 24-well plates were incubated for 14 h in RPMI 1640 medium containing 2% fetal calf serum, preincubated with sulindac sulfide for 6h, and then treated with or without β-lapachone for 24 h, then were immediately fixed in 4% paraformaldehyde in phosphate-buffered saline (PBS) for 10 min at room temperature (RT), and stained for 10 min with 0.5 ml of AO (10 mg/ml in PBS) (Sigma). .. After several PBS washes, the cells were examined on an Olympus BH-2 inverted microscope equipped with a fluorescence attachment.

    Modification:

    Article Title: β-Lapachone induces programmed necrosis through the RIP1-PARP-AIF-dependent pathway in human hepatocellular carcinoma SK-Hep1 cells
    Article Snippet: .. The culture medium used throughout these experiments was Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS), 20 mM HEPES buffer and 100 μ g/ml gentamycin. β -Lapachone, MG132, N-acetyl-cysteine (NAC), glutathione (GSH), diphenyliodonium (DPI), DPQ and dicoumarol were purchased from Sigma Chemical Co. (St. Louis, MO, USA). .. Necrostatin-1 and zVAD-fmk were obtained from Merck millipore (Bedford, MA, USA).

    Incubation:

    Article Title: Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells
    Article Snippet: .. Acridine Orange (AO) Staining Cells (5x104 ) cultured on cover-slides in 24-well plates were incubated for 14 h in RPMI 1640 medium containing 2% fetal calf serum, preincubated with sulindac sulfide for 6h, and then treated with or without β-lapachone for 24 h, then were immediately fixed in 4% paraformaldehyde in phosphate-buffered saline (PBS) for 10 min at room temperature (RT), and stained for 10 min with 0.5 ml of AO (10 mg/ml in PBS) (Sigma). .. After several PBS washes, the cells were examined on an Olympus BH-2 inverted microscope equipped with a fluorescence attachment.

    Article Title: ?-Lapachone induces heart morphogenetic and functional defects by promoting the death of erythrocytes and the endocardium in zebrafish embryos
    Article Snippet: .. After several washes with PBST (1 × PBS and 0.1% Tween-20), β-lapachone- and DMSO-treated 48-hpf embryos were incubated in H2 O2 (20 μl/ml)-activated ODA staining buffer (0.6 mg/ml ODA (Sigma-Aldrich, St. Louis, MO, USA) in 10 mM sodium acetate (pH 5.2) and 4% ethanol) for 15 min in the dark at room temperature and then washed with PBST several times. .. ROS assay β-Lapachone- and DMSO-treated embryos were incubated in 5-(and 6-)-chloromethyl-2',7'-dichloro-dihydrofluorescein diacetate (CM-H2 DCFDA) (Invitrogen, Carlsbad, CA, USA) at a final concentration of 500 ng/ml for 1 h in the dark at room temperature.

    other:

    Article Title: Terpinen-4-ol, tyrosol, and β-lapachone as potential antifungals against dimorphic fungi
    Article Snippet: Antimicrobial agents For the assays, terpinen-4-ol, tyrosol, and β-lapachone (all from Sigma Chemical Corporation, USA) were used.

    Standard Deviation:

    Article Title: Gemcitabine Functions Epigenetically by Inhibiting Repair Mediated DNA Demethylation
    Article Snippet: .. Where indicated, cells were treated with 67 nM gemcitabine, 26 nM camptothecin (MP Biomedicals), 43 nM etoposide (Sigma-Aldrich), 30 nM β-lapachone (Calbiochem) or 20 nM merbarone (Calbiochem) for 18 h. Results are shown as the mean of triplicates and error bars indicate standard deviation. .. Quantitative RT-PCR (qPCR) RNA was isolated using the RNeasy Kit (Qiagen) and reverse transcribed with the SuperScript II reverse transcriptase (Invitrogen).

    Staining:

    Article Title: Sulindac Compounds Facilitate the Cytotoxicity of ?-Lapachone by Up-Regulation of NAD(P)H Quinone Oxidoreductase in Human Lung Cancer Cells
    Article Snippet: .. Acridine Orange (AO) Staining Cells (5x104 ) cultured on cover-slides in 24-well plates were incubated for 14 h in RPMI 1640 medium containing 2% fetal calf serum, preincubated with sulindac sulfide for 6h, and then treated with or without β-lapachone for 24 h, then were immediately fixed in 4% paraformaldehyde in phosphate-buffered saline (PBS) for 10 min at room temperature (RT), and stained for 10 min with 0.5 ml of AO (10 mg/ml in PBS) (Sigma). .. After several PBS washes, the cells were examined on an Olympus BH-2 inverted microscope equipped with a fluorescence attachment.

    Article Title: ?-Lapachone induces heart morphogenetic and functional defects by promoting the death of erythrocytes and the endocardium in zebrafish embryos
    Article Snippet: .. After several washes with PBST (1 × PBS and 0.1% Tween-20), β-lapachone- and DMSO-treated 48-hpf embryos were incubated in H2 O2 (20 μl/ml)-activated ODA staining buffer (0.6 mg/ml ODA (Sigma-Aldrich, St. Louis, MO, USA) in 10 mM sodium acetate (pH 5.2) and 4% ethanol) for 15 min in the dark at room temperature and then washed with PBST several times. .. ROS assay β-Lapachone- and DMSO-treated embryos were incubated in 5-(and 6-)-chloromethyl-2',7'-dichloro-dihydrofluorescein diacetate (CM-H2 DCFDA) (Invitrogen, Carlsbad, CA, USA) at a final concentration of 500 ng/ml for 1 h in the dark at room temperature.

    Similar Products

  • Logo
  • About
  • News
  • Press Release
  • Team
  • Advisors
  • Partners
  • Contact
  • Bioz Stars
  • Bioz vStars
  • 94
    Millipore β lapachone
    Gemcitabine inhibits Gadd45a mediated gene activation. ( A–B ) Luciferase reporter assays of HEK293T cells transiently transfected with HpaII in vitro methylated Gal-responsive reporter, together with either Gadd45a (A) or Gal-Elk1 (B, specificity control). Cells were treated with DMSO (control, Ctrl), gemcitabine (Gem), camptothecin (Cpt), etoposide (Eto), <t>β-lapachone</t> (βLap), merbarone (Mer) as indicated. Shown is the fold activation by Gadd45a (A) or Gal-Elk1 (B) over control transfected cells. Error bars represent standard deviation. Significance was assessed via unpaired Student's t-test using the control sample as reference: ** = p
    β Lapachone, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β lapachone/product/Millipore
    Average 94 stars, based on 6 article reviews
    Price from $9.99 to $1999.99
    β lapachone - by Bioz Stars, 2020-07
    94/100 stars
      Buy from Supplier

    Image Search Results


    Gemcitabine inhibits Gadd45a mediated gene activation. ( A–B ) Luciferase reporter assays of HEK293T cells transiently transfected with HpaII in vitro methylated Gal-responsive reporter, together with either Gadd45a (A) or Gal-Elk1 (B, specificity control). Cells were treated with DMSO (control, Ctrl), gemcitabine (Gem), camptothecin (Cpt), etoposide (Eto), β-lapachone (βLap), merbarone (Mer) as indicated. Shown is the fold activation by Gadd45a (A) or Gal-Elk1 (B) over control transfected cells. Error bars represent standard deviation. Significance was assessed via unpaired Student's t-test using the control sample as reference: ** = p

    Journal: PLoS ONE

    Article Title: Gemcitabine Functions Epigenetically by Inhibiting Repair Mediated DNA Demethylation

    doi: 10.1371/journal.pone.0014060

    Figure Lengend Snippet: Gemcitabine inhibits Gadd45a mediated gene activation. ( A–B ) Luciferase reporter assays of HEK293T cells transiently transfected with HpaII in vitro methylated Gal-responsive reporter, together with either Gadd45a (A) or Gal-Elk1 (B, specificity control). Cells were treated with DMSO (control, Ctrl), gemcitabine (Gem), camptothecin (Cpt), etoposide (Eto), β-lapachone (βLap), merbarone (Mer) as indicated. Shown is the fold activation by Gadd45a (A) or Gal-Elk1 (B) over control transfected cells. Error bars represent standard deviation. Significance was assessed via unpaired Student's t-test using the control sample as reference: ** = p

    Article Snippet: Where indicated, cells were treated with 67 nM gemcitabine, 26 nM camptothecin (MP Biomedicals), 43 nM etoposide (Sigma-Aldrich), 30 nM β-lapachone (Calbiochem) or 20 nM merbarone (Calbiochem) for 18 h. Results are shown as the mean of triplicates and error bars indicate standard deviation.

    Techniques: Activation Assay, Luciferase, Transfection, In Vitro, Methylation, Cycling Probe Technology, Standard Deviation