etoposide eto  (Millipore)


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    Structured Review

    Millipore etoposide eto
    Etoposide Eto, supplied by Millipore, used in various techniques. Bioz Stars score: 98/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/etoposide eto/product/Millipore
    Average 98 stars, based on 7 article reviews
    Price from $9.99 to $1999.99
    etoposide eto - by Bioz Stars, 2019-12
    98/100 stars

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    Related Articles

    Cycling Probe Technology:

    Article Title: Ataxin-3 promotes genome integrity by stabilizing Chk1
    Article Snippet: Antibodies used in this study include: Chk1 (1:500, Santa Cruz Biotechnology, sc-8408 clone G-4), p-Chk1 (S345, 1:1000, Cell Signaling Technology, 2348 clone 133D3), Chk2 (1:10 000, Epitomics, 3428-1), p-Chk2 (T68, 1:1000, Cell Signaling Technology, 2661), ATX3 (1:10 000, Millipore, 5360), USP1 (1:1000, Cell Signaling Technology, D37B4), p-ATR (S428, 1:1000, Cell Signaling Technology, 2853), Myc (1:500, Santa Cruz Biotechnology, 9E10), Myc (1:1000, Covance, PRB-150P), M2 (1:2000, Sigma-Aldrich, F1804), polyclonal Flag (1:10 000, Sigma-Aldrich, 7425), HA (1:5000, Covance, PRB-101P), PCNA (1:500, Santa Cruz Biotechnology, PC10), H3 (1:10 000, Abmart, P30266), pH3 (S10, 1:1000, Cell Signaling Technology, D2C8), actin (1:10 000, proteintech, 00001-1), rabbit polyclonal ATX3 antibody was generated at Abmart. .. Camptothecin (CPT), hydroxyurea (HU), cycloheximide (CHX), nocodazole, reduced glutathione (rGTH), etoposide (ETO) and puromycin were from SIGMA, hydrogen peroxide (H2 O2 ) was from ACROS ORGANICS, proteasome inhibitor MG132 was from Selleckchem, ATM and ATR inhibitor CGK733 was from Merck. .. Polyethylenimine, linear (23 966) was from Polyscienses.

    Article Title: Natural Killer Cell-Mediated Shedding of ULBP2
    Article Snippet: Recombinant human IL-2 was from the National Cancer Institute-Frederick Cancer Research and Development Center (Frederick, MD, USA). .. Actinomycin D (ActD), Camptothecin (CPT) and Etoposide (ETO) were from Sigma-Aldrich. .. Z-VAD-FMK and Z-FA-FMK were from BD Biosciences.

    Article Title:
    Article Snippet: Bovine aprotinin, aphidicolin (APC), camptothecin (CPT), cisplatin (CDDP), methyl methanesulfonate (MMS), mitomycin C (MMC), and anti-Flag antibody (Ab) were from Sigma-Aldrich. .. Bleomycin (BLEO) (Invitrogen), etoposide (ETO) (Calbiochem), PMSF (Wako), monoclonal anti-GCN5 Ab (Chemicon), KOD FX DNA polymerase (Toyobo), all anti-acetylated histone antisera, and protein G-agarose/salmon sperm DNA (Millipore) were obtained.

    Article Title: Characterization of DDRI-18 (3,3?-(1H,3?H-5,5?-bibenzo[d]imidazole-2,2?-diyl)dianiline), a novel small molecule inhibitor modulating the DNA damage response
    Article Snippet: Doxorubicin (DOX), cisplatin, 5-fluorouracil (5-FU), KU55933 (C21 H17 NO3 S2 ) and NU7026 (C17 H15 NO3 ) were purchased from Calbiochem (San Diego, CA). .. Etoposide (ETO), camptothecin (CPT) and bleomycin were purchased from Sigma (St. Louis, MO, USA). .. Z-VAD-fmk was purchased from Tocris (Bristol, UK).

    Synthesized:

    Article Title: Impact of phase I metabolism on uptake, oxidative stress and genotoxicity of the emerging mycotoxin alternariol and its monomethyl ether in esophageal cells
    Article Snippet: 4-OH-AOH and 4-OH-AME were chemically synthesized as shown in Fig. . .. Etoposide (ETO) was bought from Sigma-Aldrich (Taufkirchen, Germany).

    Cytometry:

    Article Title: The role of antigen specificity in the binding of murine monoclonal anti-DNA antibodies to microparticles from apoptotic cells
    Article Snippet: Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes. .. Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes.

    Nuclear Magnetic Resonance:

    Article Title: Impact of phase I metabolism on uptake, oxidative stress and genotoxicity of the emerging mycotoxin alternariol and its monomethyl ether in esophageal cells
    Article Snippet: The products were purified by semi-preparative HPLC (Knauer, Germany), characterized by NMR spectroscopy (Bruker Avance UltraShield 400, Ettlingen, Germany) and HRMS analysis (Thermo Scientific LTQ Orbitrap XL Hybrid FTMS, Waltham, MA, USA), and their purity ( > 95 %) was analyzed by HPLC-DAD (Agilent 1200, Waldbronn, Germany) at a wavelength of 340 nm. .. Etoposide (ETO) was bought from Sigma-Aldrich (Taufkirchen, Germany).

    Modification:

    Article Title: miR-191 promotes tumorigenesis of human colorectal cancer through targeting C/EBPβ
    Article Snippet: The HEK293T cell line was maintained in Dulbecco's modified Eagle's medium (DMEM, HyClone, Logan, Utah, USA) supplemented with 10% fetal bovine serum and 1% penicillin–streptomycin at 37°C in 5% CO2 . .. 5-Fluorouracil (5-Fu), indomethacin (Indo), cisplatin (Cis) and etoposide (Eto) were obtained from Sigma Chemical Co. (St. Louis, MO, USA)

    Article Title: Direct and protective effects of single or combined addition of vincristine and ε-viniferin on human HepG2 cellular oxidative stress markers in vitro
    Article Snippet: In the second phase, oxidative stress formed in HepG2 cells via exogenically administered H2 O2 was studied. .. Dulbecco’s modified eagle medium (DMEM), phosphate buffered saline (PBS), Tris HCl, vincristine sulphate, and etoposide (ETO) were purchased from Sigma Aldrich Co (St. Louis, MO, USA). ε-Viniferin was provided by Actichem SA (Montauban, France). .. H2 O2 , chloroform and methanol were purchased from Merck company (Darmstadt, Germany).

    Article Title: Polycomb protein EZH2 regulates cancer cell fate decision in response to DNA damage
    Article Snippet: Other cell lines used in this study, including human osteosarcoma U2OS and Saos-2, lung caner H1299, breast cancer MCF7 and HEK epithelial 293 cells, were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA) and maintained in Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum and antibiotics in a 37°C humidified incubator containing 5% CO2 . .. Human breast epithelial MCF10A cells were obtained from ATCC and maintained, as recommended. adramycin (ADR), etoposide (ETO), nocodazole (Noco), proteasomal inhibitors MG132 and MG115 were purchased from Sigma-Aldrich (St. Louis, MO, USA).

    Derivative Assay:

    Article Title: Polycomb protein EZH2 regulates cancer cell fate decision in response to DNA damage
    Article Snippet: The human colorectal cancer HCT116 cells and its derived isogenic p53−/− HCT116 cells were kindly provided by Dr. Bert Vogelstein (John Hopkins University, MD, USA). .. Human breast epithelial MCF10A cells were obtained from ATCC and maintained, as recommended. adramycin (ADR), etoposide (ETO), nocodazole (Noco), proteasomal inhibitors MG132 and MG115 were purchased from Sigma-Aldrich (St. Louis, MO, USA).

    High Performance Liquid Chromatography:

    Article Title: Impact of phase I metabolism on uptake, oxidative stress and genotoxicity of the emerging mycotoxin alternariol and its monomethyl ether in esophageal cells
    Article Snippet: The products were purified by semi-preparative HPLC (Knauer, Germany), characterized by NMR spectroscopy (Bruker Avance UltraShield 400, Ettlingen, Germany) and HRMS analysis (Thermo Scientific LTQ Orbitrap XL Hybrid FTMS, Waltham, MA, USA), and their purity ( > 95 %) was analyzed by HPLC-DAD (Agilent 1200, Waldbronn, Germany) at a wavelength of 340 nm. .. Etoposide (ETO) was bought from Sigma-Aldrich (Taufkirchen, Germany).

    Flow Cytometry:

    Article Title: The role of antigen specificity in the binding of murine monoclonal anti-DNA antibodies to microparticles from apoptotic cells
    Article Snippet: Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes. .. Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes.

    Chromatography:

    Article Title: Sialic acid-binding lectin (leczyme) induces caspase-dependent apoptosis-mediated mitochondrial perturbation in Jurkat cells
    Article Snippet: SBL was isolated in sequential chromatography on Sephadex G-75, DEAE-cellulose, hydroxyapatite and SP-Sepharose as described previously ( ). .. Etoposide (ETO), doxorubicin (DOX) and anti-β-actin antibody were purchased from Sigma-Aldrich (Tokyo, Japan).

    Cell Culture:

    Article Title: miR-191 promotes tumorigenesis of human colorectal cancer through targeting C/EBPβ
    Article Snippet: Paragraph title: Cell culture and reagents ... 5-Fluorouracil (5-Fu), indomethacin (Indo), cisplatin (Cis) and etoposide (Eto) were obtained from Sigma Chemical Co. (St. Louis, MO, USA)

    Article Title: Nucleocytoplasmic Translocation of UBXN2A Is Required for Apoptosis during DNA Damage Stresses in Colon Cancer Cells
    Article Snippet: Etoposide (ETO) was purchased from Sigma-Aldrich. .. Etoposide (ETO) was purchased from Sigma-Aldrich.

    Article Title: The role of antigen specificity in the binding of murine monoclonal anti-DNA antibodies to microparticles from apoptotic cells
    Article Snippet: Jurkat and THP-1 human cell lines obtained from the Duke University Comprehensive Cancer Center Cell Culture Facility were cultured in RPMI 1640 medium (Invitrogen, Carlsbad, CA) supplemented with 10% fetal bovine serum (HyClone, Logan, UT) and 20 μg/ml gentamicin (Invitrogen). .. Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes. .. The cell free supernatant was centrifuged further at 16,000 × g for 45 minutes to pellet MPs.

    Article Title: Natural Killer Cell-Mediated Shedding of ULBP2
    Article Snippet: Purified NK cells were co-cultured with an equal number of Mitomycin C (Roche Diagnostics)-treated autologous PBL feeder cells in IMDM (Life Technologies) supplemented with 10% human AB serum (Sigma-Aldrich), 10% purified IL-2 (Hemagen Diagnostics), 200 U/ml recombinant human IL-2 for one week, and then expanded NK cells were cultured with IMDM supplemented with 10% human AB serum and rIL-2 (200 U/ml). .. Actinomycin D (ActD), Camptothecin (CPT) and Etoposide (ETO) were from Sigma-Aldrich.

    Article Title: Polycomb protein EZH2 regulates cancer cell fate decision in response to DNA damage
    Article Snippet: Paragraph title: Cell culture and drugs ... Human breast epithelial MCF10A cells were obtained from ATCC and maintained, as recommended. adramycin (ADR), etoposide (ETO), nocodazole (Noco), proteasomal inhibitors MG132 and MG115 were purchased from Sigma-Aldrich (St. Louis, MO, USA).

    Hemagglutination Assay:

    Article Title: Ataxin-3 promotes genome integrity by stabilizing Chk1
    Article Snippet: Antibodies used in this study include: Chk1 (1:500, Santa Cruz Biotechnology, sc-8408 clone G-4), p-Chk1 (S345, 1:1000, Cell Signaling Technology, 2348 clone 133D3), Chk2 (1:10 000, Epitomics, 3428-1), p-Chk2 (T68, 1:1000, Cell Signaling Technology, 2661), ATX3 (1:10 000, Millipore, 5360), USP1 (1:1000, Cell Signaling Technology, D37B4), p-ATR (S428, 1:1000, Cell Signaling Technology, 2853), Myc (1:500, Santa Cruz Biotechnology, 9E10), Myc (1:1000, Covance, PRB-150P), M2 (1:2000, Sigma-Aldrich, F1804), polyclonal Flag (1:10 000, Sigma-Aldrich, 7425), HA (1:5000, Covance, PRB-101P), PCNA (1:500, Santa Cruz Biotechnology, PC10), H3 (1:10 000, Abmart, P30266), pH3 (S10, 1:1000, Cell Signaling Technology, D2C8), actin (1:10 000, proteintech, 00001-1), rabbit polyclonal ATX3 antibody was generated at Abmart. .. Camptothecin (CPT), hydroxyurea (HU), cycloheximide (CHX), nocodazole, reduced glutathione (rGTH), etoposide (ETO) and puromycin were from SIGMA, hydrogen peroxide (H2 O2 ) was from ACROS ORGANICS, proteasome inhibitor MG132 was from Selleckchem, ATM and ATR inhibitor CGK733 was from Merck.

    Generated:

    Article Title: Ataxin-3 promotes genome integrity by stabilizing Chk1
    Article Snippet: Antibodies used in this study include: Chk1 (1:500, Santa Cruz Biotechnology, sc-8408 clone G-4), p-Chk1 (S345, 1:1000, Cell Signaling Technology, 2348 clone 133D3), Chk2 (1:10 000, Epitomics, 3428-1), p-Chk2 (T68, 1:1000, Cell Signaling Technology, 2661), ATX3 (1:10 000, Millipore, 5360), USP1 (1:1000, Cell Signaling Technology, D37B4), p-ATR (S428, 1:1000, Cell Signaling Technology, 2853), Myc (1:500, Santa Cruz Biotechnology, 9E10), Myc (1:1000, Covance, PRB-150P), M2 (1:2000, Sigma-Aldrich, F1804), polyclonal Flag (1:10 000, Sigma-Aldrich, 7425), HA (1:5000, Covance, PRB-101P), PCNA (1:500, Santa Cruz Biotechnology, PC10), H3 (1:10 000, Abmart, P30266), pH3 (S10, 1:1000, Cell Signaling Technology, D2C8), actin (1:10 000, proteintech, 00001-1), rabbit polyclonal ATX3 antibody was generated at Abmart. .. Camptothecin (CPT), hydroxyurea (HU), cycloheximide (CHX), nocodazole, reduced glutathione (rGTH), etoposide (ETO) and puromycin were from SIGMA, hydrogen peroxide (H2 O2 ) was from ACROS ORGANICS, proteasome inhibitor MG132 was from Selleckchem, ATM and ATR inhibitor CGK733 was from Merck.

    other:

    Article Title: The Endoplasmic Reticulum-Resident Chaperone Heat Shock Protein 47 Protects the Golgi Apparatus from the Effects of O-Glycosylation Inhibition
    Article Snippet: The chemical reagents used in this study included benzyl 2-acetamido-2-deoxy-α-d-galactopyranoside (GalNAc-bn), thapsigargin (Tg), tunicamycin(TM), staurosporine (STS), etoposide (Eto), and monensin (Sigma Chemical Co., St. Louis, MO, USA).

    Article Title: A Novel Receptor Tyrosine Kinase Switch Promotes Gastrointestinal Stromal Tumor Drug Resistance
    Article Snippet: Etoposide (Eto) was obtained from Calbiochem (La Jolla, CA, USA).

    Article Title: BRG1 Promotes chromatin remodeling around DNA damage sites
    Article Snippet: Puromycin, etoposide (ETO), and 4-hydroxy tamoxifen (4OHT) were obtained from Sigma Aldrich (St Louis, MO).

    Recombinant:

    Article Title: Natural Killer Cell-Mediated Shedding of ULBP2
    Article Snippet: Recombinant human IL-2 was from the National Cancer Institute-Frederick Cancer Research and Development Center (Frederick, MD, USA). .. Actinomycin D (ActD), Camptothecin (CPT) and Etoposide (ETO) were from Sigma-Aldrich.

    Isolation:

    Article Title: The role of antigen specificity in the binding of murine monoclonal anti-DNA antibodies to microparticles from apoptotic cells
    Article Snippet: Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes. .. The resulting MP pellet was resuspended in Ca++ and Mg++ free PBS (Invitrogen) and adjusted to a concentration of 5000 MPs/μl after counting by flow cytometry (described below, section 2.3).

    Article Title: Natural Killer Cell-Mediated Shedding of ULBP2
    Article Snippet: Human NK cells were isolated from peripheral blood lymphocytes of unidentified donors (NIH blood bank) by negative selection using the EasySep™ human NK cell enrichment kit (STEMCELL Technologies). .. Actinomycin D (ActD), Camptothecin (CPT) and Etoposide (ETO) were from Sigma-Aldrich.

    Article Title: Sialic acid-binding lectin (leczyme) induces caspase-dependent apoptosis-mediated mitochondrial perturbation in Jurkat cells
    Article Snippet: SBL was isolated in sequential chromatography on Sephadex G-75, DEAE-cellulose, hydroxyapatite and SP-Sepharose as described previously ( ). .. Etoposide (ETO), doxorubicin (DOX) and anti-β-actin antibody were purchased from Sigma-Aldrich (Tokyo, Japan).

    Purification:

    Article Title: Impact of phase I metabolism on uptake, oxidative stress and genotoxicity of the emerging mycotoxin alternariol and its monomethyl ether in esophageal cells
    Article Snippet: The products were purified by semi-preparative HPLC (Knauer, Germany), characterized by NMR spectroscopy (Bruker Avance UltraShield 400, Ettlingen, Germany) and HRMS analysis (Thermo Scientific LTQ Orbitrap XL Hybrid FTMS, Waltham, MA, USA), and their purity ( > 95 %) was analyzed by HPLC-DAD (Agilent 1200, Waldbronn, Germany) at a wavelength of 340 nm. .. Etoposide (ETO) was bought from Sigma-Aldrich (Taufkirchen, Germany).

    Article Title: Natural Killer Cell-Mediated Shedding of ULBP2
    Article Snippet: Purified NK cells were co-cultured with an equal number of Mitomycin C (Roche Diagnostics)-treated autologous PBL feeder cells in IMDM (Life Technologies) supplemented with 10% human AB serum (Sigma-Aldrich), 10% purified IL-2 (Hemagen Diagnostics), 200 U/ml recombinant human IL-2 for one week, and then expanded NK cells were cultured with IMDM supplemented with 10% human AB serum and rIL-2 (200 U/ml). .. Actinomycin D (ActD), Camptothecin (CPT) and Etoposide (ETO) were from Sigma-Aldrich.

    Spectroscopy:

    Article Title: Impact of phase I metabolism on uptake, oxidative stress and genotoxicity of the emerging mycotoxin alternariol and its monomethyl ether in esophageal cells
    Article Snippet: The products were purified by semi-preparative HPLC (Knauer, Germany), characterized by NMR spectroscopy (Bruker Avance UltraShield 400, Ettlingen, Germany) and HRMS analysis (Thermo Scientific LTQ Orbitrap XL Hybrid FTMS, Waltham, MA, USA), and their purity ( > 95 %) was analyzed by HPLC-DAD (Agilent 1200, Waldbronn, Germany) at a wavelength of 340 nm. .. Etoposide (ETO) was bought from Sigma-Aldrich (Taufkirchen, Germany).

    SDS Page:

    Article Title: Ataxin-3 promotes genome integrity by stabilizing Chk1
    Article Snippet: Proteins were separated by SDS-PAGE and blotted onto a polyvinylidene fluoride membrane (Millipore). .. Camptothecin (CPT), hydroxyurea (HU), cycloheximide (CHX), nocodazole, reduced glutathione (rGTH), etoposide (ETO) and puromycin were from SIGMA, hydrogen peroxide (H2 O2 ) was from ACROS ORGANICS, proteasome inhibitor MG132 was from Selleckchem, ATM and ATR inhibitor CGK733 was from Merck.

    Plasmid Preparation:

    Article Title: Ataxin-3 promotes genome integrity by stabilizing Chk1
    Article Snippet: Camptothecin (CPT), hydroxyurea (HU), cycloheximide (CHX), nocodazole, reduced glutathione (rGTH), etoposide (ETO) and puromycin were from SIGMA, hydrogen peroxide (H2 O2 ) was from ACROS ORGANICS, proteasome inhibitor MG132 was from Selleckchem, ATM and ATR inhibitor CGK733 was from Merck. .. Camptothecin (CPT), hydroxyurea (HU), cycloheximide (CHX), nocodazole, reduced glutathione (rGTH), etoposide (ETO) and puromycin were from SIGMA, hydrogen peroxide (H2 O2 ) was from ACROS ORGANICS, proteasome inhibitor MG132 was from Selleckchem, ATM and ATR inhibitor CGK733 was from Merck.

    Irradiation:

    Article Title: DNA damage response curtails detrimental replication stress and chromosomal instability induced by the dietary carcinogen PhIP
    Article Snippet: Etoposide (ETO) was obtained from Sigma-Aldrich and dissolved in DMSO to give a stock solution of 10 mM. .. All inhibitors (ATRi, ATMi, DNA-PKcs i, CHK1i) were dissolved in DMSO at a stock concentration of 10 mM and added to the cells 2 h before and 24 h after treatment with N-OH-PhIP as indicated.

    Selection:

    Article Title: Natural Killer Cell-Mediated Shedding of ULBP2
    Article Snippet: Human NK cells were isolated from peripheral blood lymphocytes of unidentified donors (NIH blood bank) by negative selection using the EasySep™ human NK cell enrichment kit (STEMCELL Technologies). .. Actinomycin D (ActD), Camptothecin (CPT) and Etoposide (ETO) were from Sigma-Aldrich.

    In Vitro:

    Article Title: The role of antigen specificity in the binding of murine monoclonal anti-DNA antibodies to microparticles from apoptotic cells
    Article Snippet: Paragraph title: 2.2 Preparation of microparticles from in vitro cell cultures ... Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes.

    Concentration Assay:

    Article Title: DNA damage response curtails detrimental replication stress and chromosomal instability induced by the dietary carcinogen PhIP
    Article Snippet: Both compounds were dissolved in DMSO at a final stock concentration of 30 and 10 mM, respectively. .. Etoposide (ETO) was obtained from Sigma-Aldrich and dissolved in DMSO to give a stock solution of 10 mM.

    Article Title: The role of antigen specificity in the binding of murine monoclonal anti-DNA antibodies to microparticles from apoptotic cells
    Article Snippet: Jurkat and THP-1 human cell lines obtained from the Duke University Comprehensive Cancer Center Cell Culture Facility were cultured in RPMI 1640 medium (Invitrogen, Carlsbad, CA) supplemented with 10% fetal bovine serum (HyClone, Logan, UT) and 20 μg/ml gentamicin (Invitrogen). .. Cells were cultured at 37°C and 5% CO2 , plated at a concentration of 2.5 × 106 cells/ml and induced to undergo apoptosis by treatment with 1 μM staurosporine (STS) or 10 μM etoposide (ETO) (Sigma) for 18 h. The cells in the treated culture medium were collected and washed by centrifuging at 500 × g for 5 minutes. .. The cell free supernatant was centrifuged further at 16,000 × g for 45 minutes to pellet MPs.

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  • 99
    Millipore etoposide
    Effects of either PI3K or MEK inhibition as well as the antioxidant Trolox on the up-regulation of CAV1 induced by Methotrexate and <t>Etoposide</t> Colon cancer (A) HT29(US) and (B) DLD-1 cells were treated with either the PI3K inhibitor LY294002 (LY, 10 μM), the MEK inhibitor PD98059 (PD, 50 μM) or the vitamin E analog, Trolox (TR, 2 mM) for 30 min before treatment with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Cells were harvested and total protein extracts were separated by SDS-PAGE (50 μg total protein per lane) and analyzed by Western blotting with antibodies against CAV1 and β-actin. The graphs show the expression of CAV1 normalized to β-actin (mean ± SEM) averaged from 3 independent experiments. Significant differences in comparison with the untreated condition (Basal) are indicated *** p ≤ 0.001, ** p ≤ 0.05, * p ≤ 0.01.
    Etoposide, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 147 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/etoposide/product/Millipore
    Average 99 stars, based on 147 article reviews
    Price from $9.99 to $1999.99
    etoposide - by Bioz Stars, 2019-12
    99/100 stars
      Buy from Supplier

    Image Search Results


    Effects of either PI3K or MEK inhibition as well as the antioxidant Trolox on the up-regulation of CAV1 induced by Methotrexate and Etoposide Colon cancer (A) HT29(US) and (B) DLD-1 cells were treated with either the PI3K inhibitor LY294002 (LY, 10 μM), the MEK inhibitor PD98059 (PD, 50 μM) or the vitamin E analog, Trolox (TR, 2 mM) for 30 min before treatment with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Cells were harvested and total protein extracts were separated by SDS-PAGE (50 μg total protein per lane) and analyzed by Western blotting with antibodies against CAV1 and β-actin. The graphs show the expression of CAV1 normalized to β-actin (mean ± SEM) averaged from 3 independent experiments. Significant differences in comparison with the untreated condition (Basal) are indicated *** p ≤ 0.001, ** p ≤ 0.05, * p ≤ 0.01.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Effects of either PI3K or MEK inhibition as well as the antioxidant Trolox on the up-regulation of CAV1 induced by Methotrexate and Etoposide Colon cancer (A) HT29(US) and (B) DLD-1 cells were treated with either the PI3K inhibitor LY294002 (LY, 10 μM), the MEK inhibitor PD98059 (PD, 50 μM) or the vitamin E analog, Trolox (TR, 2 mM) for 30 min before treatment with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Cells were harvested and total protein extracts were separated by SDS-PAGE (50 μg total protein per lane) and analyzed by Western blotting with antibodies against CAV1 and β-actin. The graphs show the expression of CAV1 normalized to β-actin (mean ± SEM) averaged from 3 independent experiments. Significant differences in comparison with the untreated condition (Basal) are indicated *** p ≤ 0.001, ** p ≤ 0.05, * p ≤ 0.01.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Inhibition, SDS Page, Western Blot, Expressing

    Effect of MEK and Src family kinase inhibition, as well as the anti-oxidants Trolox and Tiron on cell migration induced by Methotrexate and Etoposide (A) HT29(US) or (B) DLD-1 cells (6 x 10 5 ) were seeded in 6 cm plates 24 h before treatment with the MEK inhibitor PD98059 (PD, 50 μM), the Src family kinase inhibitor PP2 (1 mM), the vitamin E analog Trolox (TR, 2 mM) or the superoxide scavenger Tiron (Ti, 4 mM) for 30 min (PD, TR and Ti) or 1 h (PP2) before treatment with 100 nM Methotrexate or 10 μM Etoposide for 48 h. Cells (2 x 10 5 ) were then seeded in Boyden chambers coated with fibronectin (2 μg/ml) and allowed to migrate for 7 h (HT29(US) cells) or 5 h (DLD1 cells). The cells that migrated through the pores were stained and counted. Values were normalized to those obtained for cells without treatment (Basal). The graphs show the averages of results from 3 independent experiments (mean ± SEM). Significant differences are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Effect of MEK and Src family kinase inhibition, as well as the anti-oxidants Trolox and Tiron on cell migration induced by Methotrexate and Etoposide (A) HT29(US) or (B) DLD-1 cells (6 x 10 5 ) were seeded in 6 cm plates 24 h before treatment with the MEK inhibitor PD98059 (PD, 50 μM), the Src family kinase inhibitor PP2 (1 mM), the vitamin E analog Trolox (TR, 2 mM) or the superoxide scavenger Tiron (Ti, 4 mM) for 30 min (PD, TR and Ti) or 1 h (PP2) before treatment with 100 nM Methotrexate or 10 μM Etoposide for 48 h. Cells (2 x 10 5 ) were then seeded in Boyden chambers coated with fibronectin (2 μg/ml) and allowed to migrate for 7 h (HT29(US) cells) or 5 h (DLD1 cells). The cells that migrated through the pores were stained and counted. Values were normalized to those obtained for cells without treatment (Basal). The graphs show the averages of results from 3 independent experiments (mean ± SEM). Significant differences are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Inhibition, Migration, Staining

    Ex vivo treatment of colon carcinoma cells with Methotrexate and Etoposide increases the number of CAV1-expressing cells in ascites fluid and metastasis in vivo (A) Schematic summarizing events in the intraperitoneal carcinomatosis assay: Seven week-old BalbC/NoD/SciD mice (5 mice per group) were injected intraperitoneally with 1 x 10 6 sh-Scramble or sh-CAV1 (#5) HT29(US) cells treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h prior to injection. After 21 days, the animals were euthanized and paracentesis was analyzed. (B) The graph shows the number of viable cells in the ascitic fluid in each condition. (C) Representative images at 21 days post-cell injection showing the intestine, pancreas, spleen and stomach of mice injected with HT29(US) sh-Scramble (upper panel) or HT29(US) sh-CAV1 (#5) (lower panel) that prior to injection were either not treated (Basal) or treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. (D) The graph shows the total mass of the solid tumors and the infiltrated organs (intestine, pancreas, spleen and stomach) of mice injected with HT29(US) sh-Scramble or HT29(US) sh-CAV1 (#5) that were previously either not treated (Basal) or treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h, as well as the total weight of the organs in non-treated mice (NT). Significant differences are indicated, *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Ex vivo treatment of colon carcinoma cells with Methotrexate and Etoposide increases the number of CAV1-expressing cells in ascites fluid and metastasis in vivo (A) Schematic summarizing events in the intraperitoneal carcinomatosis assay: Seven week-old BalbC/NoD/SciD mice (5 mice per group) were injected intraperitoneally with 1 x 10 6 sh-Scramble or sh-CAV1 (#5) HT29(US) cells treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h prior to injection. After 21 days, the animals were euthanized and paracentesis was analyzed. (B) The graph shows the number of viable cells in the ascitic fluid in each condition. (C) Representative images at 21 days post-cell injection showing the intestine, pancreas, spleen and stomach of mice injected with HT29(US) sh-Scramble (upper panel) or HT29(US) sh-CAV1 (#5) (lower panel) that prior to injection were either not treated (Basal) or treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. (D) The graph shows the total mass of the solid tumors and the infiltrated organs (intestine, pancreas, spleen and stomach) of mice injected with HT29(US) sh-Scramble or HT29(US) sh-CAV1 (#5) that were previously either not treated (Basal) or treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h, as well as the total weight of the organs in non-treated mice (NT). Significant differences are indicated, *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Ex Vivo, Expressing, In Vivo, Mouse Assay, Injection

    CAV1 silencing precludes the increase in cell migration induced by Methotrexate and Etoposide in colon cancer cell lines Parental, sh-Scramble and sh-CAV1 (#5) (A) HT29(US) or (B) DLD-1 cells (6 x 10 5 ) were seeded in 6 cm plates 24 h before treatment with 100 nM Methotrexate or 10 μM Etoposide for 48 h. Cells (2 x 10 5 ) were then seeded in Boyden chambers coated with fibronectin (2 μg/ml) on the lower side and allowed to migrate for 7 h (HT29(US) cells) or 5 h (DLD1 cells). The cells that migrated through the pores were stained and counted. Values obtained were normalized to those obtained for parental cells without treatment. The graphs show the averages of values from 3 independent experiments (mean ± SEM). Significant differences are indicated ** p ≤ 0.01, * p ≤ 0.05.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: CAV1 silencing precludes the increase in cell migration induced by Methotrexate and Etoposide in colon cancer cell lines Parental, sh-Scramble and sh-CAV1 (#5) (A) HT29(US) or (B) DLD-1 cells (6 x 10 5 ) were seeded in 6 cm plates 24 h before treatment with 100 nM Methotrexate or 10 μM Etoposide for 48 h. Cells (2 x 10 5 ) were then seeded in Boyden chambers coated with fibronectin (2 μg/ml) on the lower side and allowed to migrate for 7 h (HT29(US) cells) or 5 h (DLD1 cells). The cells that migrated through the pores were stained and counted. Values obtained were normalized to those obtained for parental cells without treatment. The graphs show the averages of values from 3 independent experiments (mean ± SEM). Significant differences are indicated ** p ≤ 0.01, * p ≤ 0.05.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Migration, Staining

    CAV1 silencing decreases metalloproteinase activity induced by Methotrexate and Etoposide in colon cancer cell lines sh-Scramble and sh-CAV1 (#5) (A) HT29(US) or (B) DLD1 cells (6 X 10 5 ) were seeded in 6 cm plates 24 h before treatment with 100 nM Methotrexate or 10 μM Etoposide for 48 h. Cells were harvested and total protein extracts were subjected to gelatin zymography. The graphs show the densitometric analysis of gelatinolytic activity detected at 92 kDa (pro-MMP9) and 72 kDa (pro-MMP2) averaged from 3 independent experiments (mean ± SEM). Significant differences are indicated, *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: CAV1 silencing decreases metalloproteinase activity induced by Methotrexate and Etoposide in colon cancer cell lines sh-Scramble and sh-CAV1 (#5) (A) HT29(US) or (B) DLD1 cells (6 X 10 5 ) were seeded in 6 cm plates 24 h before treatment with 100 nM Methotrexate or 10 μM Etoposide for 48 h. Cells were harvested and total protein extracts were subjected to gelatin zymography. The graphs show the densitometric analysis of gelatinolytic activity detected at 92 kDa (pro-MMP9) and 72 kDa (pro-MMP2) averaged from 3 independent experiments (mean ± SEM). Significant differences are indicated, *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Activity Assay, Zymography

    Working model summarizing the main findings described in this study identifying the mechanisms by which cytotoxic drugs induce CAV1 expression Initially, CAV1 expression is repressed by methylation in the gene promoter region in tumor cells. Exposure to the anti-neoplastic drugs Methotrexate or Etoposide induces promoter demethylation that increases transcription and expression of CAV1 that is is mediated by ERK activation and ROS production. Likewise, ROS are shown to promote Src family kinase-dependent CAV1 phosphorylation that may lead to Rac1 and metalloproteinase activation, as well as increased migration, invasion and metastasis.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Working model summarizing the main findings described in this study identifying the mechanisms by which cytotoxic drugs induce CAV1 expression Initially, CAV1 expression is repressed by methylation in the gene promoter region in tumor cells. Exposure to the anti-neoplastic drugs Methotrexate or Etoposide induces promoter demethylation that increases transcription and expression of CAV1 that is is mediated by ERK activation and ROS production. Likewise, ROS are shown to promote Src family kinase-dependent CAV1 phosphorylation that may lead to Rac1 and metalloproteinase activation, as well as increased migration, invasion and metastasis.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Expressing, Methylation, Activation Assay, Migration

    Anti-neoplastic drugs increase CAV1 expression in colon and breast cancer cell lines Colon cancer cells (A) HT29(US), (B) DLD-1, (C) HT29(ATCC) and breast cancer cells (D) MCF-7 were treated with 100 nM Methotrexate (MT), 10 μM Etoposide (ET), 1 μM Doxorubicin (DX), 5 nM Staurosporine (ST), 5 nM Taxol (TX) or 100 nM Cisplatin (CP) for 24 h. Cells were harvested and total protein extracts were separated by SDS-PAGE (50 μg total protein per lane) and analyzed by Western blotting with antibodies against caveolin-1 (CAV1) and β-actin. The graphs show the expression of CAV1 normalized to β-actin (mean ± SEM) of 3 independent experiments. Significant differences in comparison with the untreated condition (Basal) are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05 .

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Anti-neoplastic drugs increase CAV1 expression in colon and breast cancer cell lines Colon cancer cells (A) HT29(US), (B) DLD-1, (C) HT29(ATCC) and breast cancer cells (D) MCF-7 were treated with 100 nM Methotrexate (MT), 10 μM Etoposide (ET), 1 μM Doxorubicin (DX), 5 nM Staurosporine (ST), 5 nM Taxol (TX) or 100 nM Cisplatin (CP) for 24 h. Cells were harvested and total protein extracts were separated by SDS-PAGE (50 μg total protein per lane) and analyzed by Western blotting with antibodies against caveolin-1 (CAV1) and β-actin. The graphs show the expression of CAV1 normalized to β-actin (mean ± SEM) of 3 independent experiments. Significant differences in comparison with the untreated condition (Basal) are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05 .

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Expressing, SDS Page, Western Blot

    Methotrexate and Etoposide induce an increase in CAV1 mRNA levels in colon cancer cell lines Colon cancer cells (A) HT29(US) and (B) DLD-1 were treated with 100 nM Methotrexate or 10 μM Etoposide for 12 and 24 h. CAV1 mRNA levels were evaluated by quantitative RT-PCR analysis, using β-actin as an internal control. Values obtained by analysis of three independent experiments are shown for CAV1 mRNA following standardization to β-actin (mean ± SEM) and after normalizing to the values obtained for untreated (0 h) samples. Statistically significant differences compared with the controls (time 0) are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Methotrexate and Etoposide induce an increase in CAV1 mRNA levels in colon cancer cell lines Colon cancer cells (A) HT29(US) and (B) DLD-1 were treated with 100 nM Methotrexate or 10 μM Etoposide for 12 and 24 h. CAV1 mRNA levels were evaluated by quantitative RT-PCR analysis, using β-actin as an internal control. Values obtained by analysis of three independent experiments are shown for CAV1 mRNA following standardization to β-actin (mean ± SEM) and after normalizing to the values obtained for untreated (0 h) samples. Statistically significant differences compared with the controls (time 0) are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Quantitative RT-PCR

    Methotrexate and Etoposide induce CAV1 promoter demethylation in colon cancer cells (A) HT29(US) and (B) DLD-1 colon cancer cells were treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Genomic DNA (gDNA) was denatured for 10 min at 95°C and then immunoprecipitated using the anti-5-methylcytidine antibody (5mC). Affinity purified DNA was then evaluated using qPCR analysis, defining the enrichment levels as a percentage of the input material. Specific primers were used to analyze the CAV1 proximal promoter (CAV1 promoter) or negative (CSa region) and positive control regions (IAP region). Statistically significant differences compared with the control (Basal, black bars) are indicated *** p ≤ 0.001, ** p ≤ 0.01.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Methotrexate and Etoposide induce CAV1 promoter demethylation in colon cancer cells (A) HT29(US) and (B) DLD-1 colon cancer cells were treated with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Genomic DNA (gDNA) was denatured for 10 min at 95°C and then immunoprecipitated using the anti-5-methylcytidine antibody (5mC). Affinity purified DNA was then evaluated using qPCR analysis, defining the enrichment levels as a percentage of the input material. Specific primers were used to analyze the CAV1 proximal promoter (CAV1 promoter) or negative (CSa region) and positive control regions (IAP region). Statistically significant differences compared with the control (Basal, black bars) are indicated *** p ≤ 0.001, ** p ≤ 0.01.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Immunoprecipitation, Affinity Purification, Real-time Polymerase Chain Reaction, Positive Control

    MEK inhibition reduces ROS production induced by Methotrexate and Etoposide (A) HT29(US) or (B) DLD-1 colon cancer cells (3 x 10 6 ) were seeded in 24-well plates and, after 24 h, were treated with the MEK inhibitor PD98059 (PD, 50 μM) for 30 min, added prior to treatment with 100 nM Methotrexate for 20 h or 10 μM Etoposide for the indicated time periods. Cells were washed 3 times with PBS and subsequently incubated with trypsin for 5 min. Once in suspension, cells were loaded with the probe DHR123 (1.4 μg/ml) in RPMI media without serum for 30 min and then the reaction was stopped on ice. The extent of DHR123 oxidation was determined by flow cytometry. The graphs show DHR123 fluorescence normalized to the untreated condition (Basal) (mean ± SEM) averaged from 3 independent experiments. Significant differences are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: MEK inhibition reduces ROS production induced by Methotrexate and Etoposide (A) HT29(US) or (B) DLD-1 colon cancer cells (3 x 10 6 ) were seeded in 24-well plates and, after 24 h, were treated with the MEK inhibitor PD98059 (PD, 50 μM) for 30 min, added prior to treatment with 100 nM Methotrexate for 20 h or 10 μM Etoposide for the indicated time periods. Cells were washed 3 times with PBS and subsequently incubated with trypsin for 5 min. Once in suspension, cells were loaded with the probe DHR123 (1.4 μg/ml) in RPMI media without serum for 30 min and then the reaction was stopped on ice. The extent of DHR123 oxidation was determined by flow cytometry. The graphs show DHR123 fluorescence normalized to the untreated condition (Basal) (mean ± SEM) averaged from 3 independent experiments. Significant differences are indicated *** p ≤ 0.001, ** p ≤ 0.01, * p ≤ 0.05.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Inhibition, Incubation, Flow Cytometry, Cytometry, Fluorescence

    Methotrexate and Etoposide increase cancer cell invasion in a Src family kinase and Rac1 dependent manner (A) HT29(US) or (B) DLD1 (6 X 10 5 ) cells were seeded in 6 cm plates 24 h before pre-treatment with either the Src family kinase inhibitor, PP2 (1 mM) or with the Tiam1 inhibitor, NS (NSC 23766, 100 mM) for 60 min followed by the treatment with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. (C) HT29(US) or (D) DLD1 (6 X 10 5 ) cells were transfected with GFP (white bars) or with the Rac1 dominant-negative, RacS17N (black bars), 24 h before the treatment with 100 mM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Then, cells (2 x 10 5 ) were seeded in Matrigel-coated chambers and allowed to invade the matrix for 24 h. The cells that accumulated on the lower surface of the membrane were then stained and counted. Values obtained were normalized to those obtained for cells without treatment (Basal). The graphs show the averages of results from 3 independent experiments (mean ± SEM). Significant differences are indicated, ** p ≤ 0.01, * p ≤ 0.05.

    Journal: Oncotarget

    Article Title: Anti-neoplastic drugs increase caveolin-1-dependent migration, invasion and metastasis of cancer cells

    doi: 10.18632/oncotarget.22955

    Figure Lengend Snippet: Methotrexate and Etoposide increase cancer cell invasion in a Src family kinase and Rac1 dependent manner (A) HT29(US) or (B) DLD1 (6 X 10 5 ) cells were seeded in 6 cm plates 24 h before pre-treatment with either the Src family kinase inhibitor, PP2 (1 mM) or with the Tiam1 inhibitor, NS (NSC 23766, 100 mM) for 60 min followed by the treatment with 100 nM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. (C) HT29(US) or (D) DLD1 (6 X 10 5 ) cells were transfected with GFP (white bars) or with the Rac1 dominant-negative, RacS17N (black bars), 24 h before the treatment with 100 mM Methotrexate (MT) or 10 μM Etoposide (ET) for 48 h. Then, cells (2 x 10 5 ) were seeded in Matrigel-coated chambers and allowed to invade the matrix for 24 h. The cells that accumulated on the lower surface of the membrane were then stained and counted. Values obtained were normalized to those obtained for cells without treatment (Basal). The graphs show the averages of results from 3 independent experiments (mean ± SEM). Significant differences are indicated, ** p ≤ 0.01, * p ≤ 0.05.

    Article Snippet: The anti-neoplastic drugs Methotrexate, Etoposide, Doxorubicin, Staurosporine and Cisplatin were from Calbiochem (La Jolla, CA, USA), while Taxol was from Molecular Probes (Eugene, OR, USA).

    Techniques: Transfection, Dominant Negative Mutation, Staining

    1p36 allelic loss is not observed in parental and HTLA-ER cells and miRNA-34a levels are reduced in both cell populations following etoposide treatment. ( A ) FISH analysis of HTLA-230 cells and HTLA-ER cells. Upper and lower left panels: metaphase of both HTLA-230 and HTLA-ER cells displays one normal chromosome 1 (close arrow) one 1 p arm derivative (close arrowhead) and 1 q arm derivative (open arrow); lower right panel: metaphase of HTLA-ER cells displaying one additional 1 p arm derivative (open arrowhead). ( B ) Expression levels of miRNA-34a in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Histograms summarize quantitative data of means ± S.E.M of three independent experiments. ** p

    Journal: Scientific Reports

    Article Title: Etoposide-resistance in a neuroblastoma model cell line is associated with 13q14.3 mono-allelic deletion and miRNA-15a/16-1 down-regulation

    doi: 10.1038/s41598-018-32195-7

    Figure Lengend Snippet: 1p36 allelic loss is not observed in parental and HTLA-ER cells and miRNA-34a levels are reduced in both cell populations following etoposide treatment. ( A ) FISH analysis of HTLA-230 cells and HTLA-ER cells. Upper and lower left panels: metaphase of both HTLA-230 and HTLA-ER cells displays one normal chromosome 1 (close arrow) one 1 p arm derivative (close arrowhead) and 1 q arm derivative (open arrow); lower right panel: metaphase of HTLA-ER cells displaying one additional 1 p arm derivative (open arrowhead). ( B ) Expression levels of miRNA-34a in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Histograms summarize quantitative data of means ± S.E.M of three independent experiments. ** p

    Article Snippet: Parental HTLA-230 and HTLA-ER cells were treated for 24 hrs with 1.25 µM etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany).

    Techniques: Fluorescence In Situ Hybridization, Expressing

    Parental and HTLA-ER cells express a non-inducible P53 protein carrying the homozygous TP53 missense mutation A161T. ( A ) Protein levels of P53 in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Immunoblots are representative of three independent experiments with essentially similar results. β-Actin is the internal loading control. ( B ) Transactivation ability of wild-type and mutant (A161T) P53 proteins in yLFM-P21-5′, yLFM-BAX A + B and yLFM-MDM2P2C yeast strains. The transactivation ability was determined at two different temperatures (30 °C and 36 °C) using a constitutive expression of P53 proteins (ADH1 promoter). Presented data are the fold of induction over empty vector (pRS315) and standard deviation of four biological replicates. # p

    Journal: Scientific Reports

    Article Title: Etoposide-resistance in a neuroblastoma model cell line is associated with 13q14.3 mono-allelic deletion and miRNA-15a/16-1 down-regulation

    doi: 10.1038/s41598-018-32195-7

    Figure Lengend Snippet: Parental and HTLA-ER cells express a non-inducible P53 protein carrying the homozygous TP53 missense mutation A161T. ( A ) Protein levels of P53 in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Immunoblots are representative of three independent experiments with essentially similar results. β-Actin is the internal loading control. ( B ) Transactivation ability of wild-type and mutant (A161T) P53 proteins in yLFM-P21-5′, yLFM-BAX A + B and yLFM-MDM2P2C yeast strains. The transactivation ability was determined at two different temperatures (30 °C and 36 °C) using a constitutive expression of P53 proteins (ADH1 promoter). Presented data are the fold of induction over empty vector (pRS315) and standard deviation of four biological replicates. # p

    Article Snippet: Parental HTLA-230 and HTLA-ER cells were treated for 24 hrs with 1.25 µM etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany).

    Techniques: Mutagenesis, Western Blot, Expressing, Plasmid Preparation, Standard Deviation

    Molecular mechanisms underlying the chemoresistance of HTLA-ER cells. This figure illustrates the observed molecular mechanisms underlying chemoresistance of HTLA-ER cells and the events leading to apoptosis in etoposide sensitive HTLA parental cells. Left panel: Short-term treatment with etoposide of HTLA-230 cells reduces oxidative phosphorylation and decreases glutathione (GSH) levels inducing reactive oxygen species (ROS) overproduction, thus leading to DNA damage (H2AX). Consequently, etoposide-induced genotoxic stress increases pro-apoptotic Bax, reduces anti-apoptotic Bcl2 and stimulates P53-Ser15 phosphorylation, two events leading to apoptosis and chemosensitivity. Right panel: HTLA-ER cells are able to efficiently counteract etoposide-induced ROS production by maintaining an efficient aerobic metabolism and increasing GSH levels. Long-term treatment with etoposide causes a deletion of the 13q14.3 locus and the consequent downregulation of miRNAs 15a/16-1, stimulating several pro-survival signals which contribute to inducing chemoresistance.

    Journal: Scientific Reports

    Article Title: Etoposide-resistance in a neuroblastoma model cell line is associated with 13q14.3 mono-allelic deletion and miRNA-15a/16-1 down-regulation

    doi: 10.1038/s41598-018-32195-7

    Figure Lengend Snippet: Molecular mechanisms underlying the chemoresistance of HTLA-ER cells. This figure illustrates the observed molecular mechanisms underlying chemoresistance of HTLA-ER cells and the events leading to apoptosis in etoposide sensitive HTLA parental cells. Left panel: Short-term treatment with etoposide of HTLA-230 cells reduces oxidative phosphorylation and decreases glutathione (GSH) levels inducing reactive oxygen species (ROS) overproduction, thus leading to DNA damage (H2AX). Consequently, etoposide-induced genotoxic stress increases pro-apoptotic Bax, reduces anti-apoptotic Bcl2 and stimulates P53-Ser15 phosphorylation, two events leading to apoptosis and chemosensitivity. Right panel: HTLA-ER cells are able to efficiently counteract etoposide-induced ROS production by maintaining an efficient aerobic metabolism and increasing GSH levels. Long-term treatment with etoposide causes a deletion of the 13q14.3 locus and the consequent downregulation of miRNAs 15a/16-1, stimulating several pro-survival signals which contribute to inducing chemoresistance.

    Article Snippet: Parental HTLA-230 and HTLA-ER cells were treated for 24 hrs with 1.25 µM etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany).

    Techniques:

    P53 Ser15 phosphorylation is detected only in etoposide-treated HTLA parental cells and is associated with PPM1D up-regulation. ( A ) Protein levels of phospho-(Ser15)-P53 in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Immunoblots are representative of three independent experiments with essentially similar results. β-Actin is the internal loading control. Histograms summarize quantitative data of phospho-P53/P53 ratio means ± S.E.M of three independent experiments. ** p

    Journal: Scientific Reports

    Article Title: Etoposide-resistance in a neuroblastoma model cell line is associated with 13q14.3 mono-allelic deletion and miRNA-15a/16-1 down-regulation

    doi: 10.1038/s41598-018-32195-7

    Figure Lengend Snippet: P53 Ser15 phosphorylation is detected only in etoposide-treated HTLA parental cells and is associated with PPM1D up-regulation. ( A ) Protein levels of phospho-(Ser15)-P53 in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Immunoblots are representative of three independent experiments with essentially similar results. β-Actin is the internal loading control. Histograms summarize quantitative data of phospho-P53/P53 ratio means ± S.E.M of three independent experiments. ** p

    Article Snippet: Parental HTLA-230 and HTLA-ER cells were treated for 24 hrs with 1.25 µM etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany).

    Techniques: Western Blot

    HTLA-ER cells have a deletion at the 13q14.3 locus which is associated with decreased levels of miRNAs 15a/16-1 in respect to parental cells. ( A ) FISH analysis of HTLA-230 and HTLA-ER cells: Upper panels: nuclei of HTLA-230 cells with two CEP12, two 13q34 and two D13S319 signals; nuclei of HTLA-ER cells with two CEP12, three 13q34 and one D13S319 signals. Lower panels: metaphase of HTLA-230 cells with two chromosomes 13 displaying 13q34 and D13S319 signals; metaphase of HTLA-ER cells with one chromosome 13 displaying 13q34 and D13S319 signals, one rearranged chromosome 13 displaying two 13q34, and one chromosome 12 displaying cep 12signal. ( B ) Expression levels of miRNA-15a (left panel) and miRNA-16 (right panel) in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Histograms summarize quantitative data of means ± S.E.M of three independent experiments. ** p

    Journal: Scientific Reports

    Article Title: Etoposide-resistance in a neuroblastoma model cell line is associated with 13q14.3 mono-allelic deletion and miRNA-15a/16-1 down-regulation

    doi: 10.1038/s41598-018-32195-7

    Figure Lengend Snippet: HTLA-ER cells have a deletion at the 13q14.3 locus which is associated with decreased levels of miRNAs 15a/16-1 in respect to parental cells. ( A ) FISH analysis of HTLA-230 and HTLA-ER cells: Upper panels: nuclei of HTLA-230 cells with two CEP12, two 13q34 and two D13S319 signals; nuclei of HTLA-ER cells with two CEP12, three 13q34 and one D13S319 signals. Lower panels: metaphase of HTLA-230 cells with two chromosomes 13 displaying 13q34 and D13S319 signals; metaphase of HTLA-ER cells with one chromosome 13 displaying 13q34 and D13S319 signals, one rearranged chromosome 13 displaying two 13q34, and one chromosome 12 displaying cep 12signal. ( B ) Expression levels of miRNA-15a (left panel) and miRNA-16 (right panel) in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Histograms summarize quantitative data of means ± S.E.M of three independent experiments. ** p

    Article Snippet: Parental HTLA-230 and HTLA-ER cells were treated for 24 hrs with 1.25 µM etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany).

    Techniques: Fluorescence In Situ Hybridization, Expressing

    The mitotic index of HTLA-ER cells and their Bax/Bcl2 ratio were not modified by acute etoposide exposure. ( A ) Mitotic index of HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Histograms summarize quantitative data of means ± S.D. of four independent experiments per experimental condition (at least 4 × 10 3 cells per experimental condition were counted) ** p

    Journal: Scientific Reports

    Article Title: Etoposide-resistance in a neuroblastoma model cell line is associated with 13q14.3 mono-allelic deletion and miRNA-15a/16-1 down-regulation

    doi: 10.1038/s41598-018-32195-7

    Figure Lengend Snippet: The mitotic index of HTLA-ER cells and their Bax/Bcl2 ratio were not modified by acute etoposide exposure. ( A ) Mitotic index of HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Histograms summarize quantitative data of means ± S.D. of four independent experiments per experimental condition (at least 4 × 10 3 cells per experimental condition were counted) ** p

    Article Snippet: Parental HTLA-230 and HTLA-ER cells were treated for 24 hrs with 1.25 µM etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany).

    Techniques: Modification

    BMI-1 overexpression with consequent p16 down-regulation is found in HTLA-ER cells in respect to parental cells. ( A ) Protein levels of BMI-1 in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Immunoblots are representative of three independent experiments with essentially similar results. β-Actin is the internal loading control. Histograms summarize quantitative data of protein expression levels means, normalized to β-actin expression ± S.E.M of three independent experiments. * p

    Journal: Scientific Reports

    Article Title: Etoposide-resistance in a neuroblastoma model cell line is associated with 13q14.3 mono-allelic deletion and miRNA-15a/16-1 down-regulation

    doi: 10.1038/s41598-018-32195-7

    Figure Lengend Snippet: BMI-1 overexpression with consequent p16 down-regulation is found in HTLA-ER cells in respect to parental cells. ( A ) Protein levels of BMI-1 in HTLA-230 and HTLA-ER cells untreated or treated for 24 hrs with 1.25 μM etoposide. Immunoblots are representative of three independent experiments with essentially similar results. β-Actin is the internal loading control. Histograms summarize quantitative data of protein expression levels means, normalized to β-actin expression ± S.E.M of three independent experiments. * p

    Article Snippet: Parental HTLA-230 and HTLA-ER cells were treated for 24 hrs with 1.25 µM etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany).

    Techniques: Over Expression, Western Blot, Expressing

    HTLA-Chr cells are characterized by higher GSH levels, a lower amount of a lipid peroxidation marker and up-regulation of GST activity A. Reduced and oxidized glutathione (GSH and GSSG) levels were analyzed in untreated and in 1.25 μM etoposide-treated (24 hrs) HTLA and HTLA-Chr cells. Results were reported as μM/μg protein. Histogram summarizes quantitative data of means ± S.E.M. of six independent experiments. ** p

    Journal: Oncotarget

    Article Title: Glutathione-mediated antioxidant response and aerobic metabolism: two crucial factors involved in determining the multi-drug resistance of high-risk neuroblastoma

    doi: 10.18632/oncotarget.12209

    Figure Lengend Snippet: HTLA-Chr cells are characterized by higher GSH levels, a lower amount of a lipid peroxidation marker and up-regulation of GST activity A. Reduced and oxidized glutathione (GSH and GSSG) levels were analyzed in untreated and in 1.25 μM etoposide-treated (24 hrs) HTLA and HTLA-Chr cells. Results were reported as μM/μg protein. Histogram summarizes quantitative data of means ± S.E.M. of six independent experiments. ** p

    Article Snippet: The Etoposide-Resistant cell line (HTLA-Chr) was selected by treating HTLA-230 cells for 6 months with increasing concentrations of etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany; up to 1.25 μM), and then maintaining them in a medium supplemented with 1.25 μM etoposide, the dose comparable to that clinically used [ ].

    Techniques: Marker, Activity Assay

    NAC treatment enhances GSH levels, decreases H 2 O 2 production and markedly promotes the tumorigenic potential of neuroblastoma cells A. GSH levels were analyzed in HTLA and HTLA-Chr cells treated with 2 mM NAC or pre-treated (1 hr) with 2 mM NAC and then exposed (24 hrs) to 1.25 μM etoposide. Histogram summarizes quantitative data of the means ± S.E.M. of three independent experiments. ** p

    Journal: Oncotarget

    Article Title: Glutathione-mediated antioxidant response and aerobic metabolism: two crucial factors involved in determining the multi-drug resistance of high-risk neuroblastoma

    doi: 10.18632/oncotarget.12209

    Figure Lengend Snippet: NAC treatment enhances GSH levels, decreases H 2 O 2 production and markedly promotes the tumorigenic potential of neuroblastoma cells A. GSH levels were analyzed in HTLA and HTLA-Chr cells treated with 2 mM NAC or pre-treated (1 hr) with 2 mM NAC and then exposed (24 hrs) to 1.25 μM etoposide. Histogram summarizes quantitative data of the means ± S.E.M. of three independent experiments. ** p

    Article Snippet: The Etoposide-Resistant cell line (HTLA-Chr) was selected by treating HTLA-230 cells for 6 months with increasing concentrations of etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany; up to 1.25 μM), and then maintaining them in a medium supplemented with 1.25 μM etoposide, the dose comparable to that clinically used [ ].

    Techniques:

    HTLA-Chr cells develop a multi-drug resistant phenotype Cell viability was determined by MTT assays in cells exposed to increasing concentrations of etoposide (1.25–100 μM) for 24 hrs A. of doxorubicin (0.046-14.72 μM) for 24, 48 and 72 hrs B. and of H 2 O 2 (250-1000 μM) for 3 hrs C. Histograms summarize quantitative data of the means ± S.E.M. of four independent experiments. * p

    Journal: Oncotarget

    Article Title: Glutathione-mediated antioxidant response and aerobic metabolism: two crucial factors involved in determining the multi-drug resistance of high-risk neuroblastoma

    doi: 10.18632/oncotarget.12209

    Figure Lengend Snippet: HTLA-Chr cells develop a multi-drug resistant phenotype Cell viability was determined by MTT assays in cells exposed to increasing concentrations of etoposide (1.25–100 μM) for 24 hrs A. of doxorubicin (0.046-14.72 μM) for 24, 48 and 72 hrs B. and of H 2 O 2 (250-1000 μM) for 3 hrs C. Histograms summarize quantitative data of the means ± S.E.M. of four independent experiments. * p

    Article Snippet: The Etoposide-Resistant cell line (HTLA-Chr) was selected by treating HTLA-230 cells for 6 months with increasing concentrations of etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany; up to 1.25 μM), and then maintaining them in a medium supplemented with 1.25 μM etoposide, the dose comparable to that clinically used [ ].

    Techniques: MTT Assay

    HTLA-Chr cells do not change H 2 O 2 production after treatment with etoposide or doxorubicin H 2 O 2 production was analyzed in HTLA and in HTLA-Chr cells incubated for 24 hrs with increasing concentrations of etoposide (1.25–100 μM) A. or doxorubicin (0.046-14.72 μM) B. Histograms summarize quantitative data of the means ± S.E.M. of three independent experiments. * p

    Journal: Oncotarget

    Article Title: Glutathione-mediated antioxidant response and aerobic metabolism: two crucial factors involved in determining the multi-drug resistance of high-risk neuroblastoma

    doi: 10.18632/oncotarget.12209

    Figure Lengend Snippet: HTLA-Chr cells do not change H 2 O 2 production after treatment with etoposide or doxorubicin H 2 O 2 production was analyzed in HTLA and in HTLA-Chr cells incubated for 24 hrs with increasing concentrations of etoposide (1.25–100 μM) A. or doxorubicin (0.046-14.72 μM) B. Histograms summarize quantitative data of the means ± S.E.M. of three independent experiments. * p

    Article Snippet: The Etoposide-Resistant cell line (HTLA-Chr) was selected by treating HTLA-230 cells for 6 months with increasing concentrations of etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany; up to 1.25 μM), and then maintaining them in a medium supplemented with 1.25 μM etoposide, the dose comparable to that clinically used [ ].

    Techniques: Incubation

    HTLA-Chr cells have a major oxygen consumption, an increased oxidative phosphorylation and an up-regulation of catalase activity A. The oxygen consumption rate (OCR) was evaluated in untreated and in 1.25 μM etoposide-treated (24 hrs) HTLA and HTLA-Chr cells. Results were reported as nmol O 2 /min/10 6 cells. Histogram summarizes quantitative data of means ± S.E.M. of three independent experiments. ** p

    Journal: Oncotarget

    Article Title: Glutathione-mediated antioxidant response and aerobic metabolism: two crucial factors involved in determining the multi-drug resistance of high-risk neuroblastoma

    doi: 10.18632/oncotarget.12209

    Figure Lengend Snippet: HTLA-Chr cells have a major oxygen consumption, an increased oxidative phosphorylation and an up-regulation of catalase activity A. The oxygen consumption rate (OCR) was evaluated in untreated and in 1.25 μM etoposide-treated (24 hrs) HTLA and HTLA-Chr cells. Results were reported as nmol O 2 /min/10 6 cells. Histogram summarizes quantitative data of means ± S.E.M. of three independent experiments. ** p

    Article Snippet: The Etoposide-Resistant cell line (HTLA-Chr) was selected by treating HTLA-230 cells for 6 months with increasing concentrations of etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany; up to 1.25 μM), and then maintaining them in a medium supplemented with 1.25 μM etoposide, the dose comparable to that clinically used [ ].

    Techniques: Activity Assay

    BSO treatment induces GSH depletion, increases H 2 O 2 production and markedly reduces the tumorigenic potential of etoposide-resistant cells A. GSH levels were analyzed in HTLA and HTLA-Chr cell treated with 1 mM BSO or pre-treated (1 hr) with 1 mM BSO and then exposed (24 hrs) to 1.25 μM etoposide. Results were reported as μM/μg protein. Histogram summarizes quantitative data of the means ± S.E.M. of three independent experiments. ** p

    Journal: Oncotarget

    Article Title: Glutathione-mediated antioxidant response and aerobic metabolism: two crucial factors involved in determining the multi-drug resistance of high-risk neuroblastoma

    doi: 10.18632/oncotarget.12209

    Figure Lengend Snippet: BSO treatment induces GSH depletion, increases H 2 O 2 production and markedly reduces the tumorigenic potential of etoposide-resistant cells A. GSH levels were analyzed in HTLA and HTLA-Chr cell treated with 1 mM BSO or pre-treated (1 hr) with 1 mM BSO and then exposed (24 hrs) to 1.25 μM etoposide. Results were reported as μM/μg protein. Histogram summarizes quantitative data of the means ± S.E.M. of three independent experiments. ** p

    Article Snippet: The Etoposide-Resistant cell line (HTLA-Chr) was selected by treating HTLA-230 cells for 6 months with increasing concentrations of etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany; up to 1.25 μM), and then maintaining them in a medium supplemented with 1.25 μM etoposide, the dose comparable to that clinically used [ ].

    Techniques:

    Chronically-etoposide-treated HTLA cells (HTLA-Chr) are less proliferating and tumorigenic than untreated HTLA parental cells and they evade apoptotic death induced by etoposide exposure A. Proliferation assay. HTLA parental cells and HTLA-Chr cells were incubated with CFDA-SE and the intensity of cellular CFDA-SE fluorescence was evaluated at 24 hrs and 48 hrs after 1.25 μM etoposide treatment. Results were expressed as proliferation index and are the means ±S.E.M. of three independent experiments. ** p

    Journal: Oncotarget

    Article Title: Glutathione-mediated antioxidant response and aerobic metabolism: two crucial factors involved in determining the multi-drug resistance of high-risk neuroblastoma

    doi: 10.18632/oncotarget.12209

    Figure Lengend Snippet: Chronically-etoposide-treated HTLA cells (HTLA-Chr) are less proliferating and tumorigenic than untreated HTLA parental cells and they evade apoptotic death induced by etoposide exposure A. Proliferation assay. HTLA parental cells and HTLA-Chr cells were incubated with CFDA-SE and the intensity of cellular CFDA-SE fluorescence was evaluated at 24 hrs and 48 hrs after 1.25 μM etoposide treatment. Results were expressed as proliferation index and are the means ±S.E.M. of three independent experiments. ** p

    Article Snippet: The Etoposide-Resistant cell line (HTLA-Chr) was selected by treating HTLA-230 cells for 6 months with increasing concentrations of etoposide (Calbiochem, Merck KGaA, Darmstadt, Germany; up to 1.25 μM), and then maintaining them in a medium supplemented with 1.25 μM etoposide, the dose comparable to that clinically used [ ].

    Techniques: Proliferation Assay, Incubation, Fluorescence

    Effect of PES on cell cycle events. a Representative flow cytometry profiles of MEFs, with control (no treatment), treatment with 1.5 µM etoposide for 18 h, pre-treatment with 30 μM of PFT-α followed by 18 h treatment with 1.5 µM etoposide or pre-treatment with 10 μM of PES followed by 18 h treatment with 1.5 µM etoposide. Percentage of cells having sub-G1 DNA content is indicated. Data shown are representative of six experiments with similar results. b Columns represent distribution of cells in G1, S and G2/M phases of cell cycle, as analyzed by flow cytometry. Data are mean + SD of three independent experiments with three replicates each. c MEFs were treated for various times with 15 µM of etoposide either alone or pre-treated with 10 μM of PES. Total RNA was isolated and reverse transcribed into cDNA. The expression of p21 cip1/waf1 was determined using specific primers in RT-PCR. β-actin was used as a loading control. Representative of three independent experiments. d MEFs were untreated (Con) or treated with 1.5 µM of etoposide for 3 or 6 h alone or pre-treated with 10 μM PES. Total cell proteins were probed with anti-Ser345 CHK1, anti-CHK1, anti-phospho-Tyr 15 CDK1, anti-CDK1, Cyclin B1 antibodies, and anti-vinculin as the loading control.

    Journal: Cancer Cell International

    Article Title: Etoposide induces cell death via mitochondrial-dependent actions of p53

    doi: 10.1186/s12935-015-0231-z

    Figure Lengend Snippet: Effect of PES on cell cycle events. a Representative flow cytometry profiles of MEFs, with control (no treatment), treatment with 1.5 µM etoposide for 18 h, pre-treatment with 30 μM of PFT-α followed by 18 h treatment with 1.5 µM etoposide or pre-treatment with 10 μM of PES followed by 18 h treatment with 1.5 µM etoposide. Percentage of cells having sub-G1 DNA content is indicated. Data shown are representative of six experiments with similar results. b Columns represent distribution of cells in G1, S and G2/M phases of cell cycle, as analyzed by flow cytometry. Data are mean + SD of three independent experiments with three replicates each. c MEFs were treated for various times with 15 µM of etoposide either alone or pre-treated with 10 μM of PES. Total RNA was isolated and reverse transcribed into cDNA. The expression of p21 cip1/waf1 was determined using specific primers in RT-PCR. β-actin was used as a loading control. Representative of three independent experiments. d MEFs were untreated (Con) or treated with 1.5 µM of etoposide for 3 or 6 h alone or pre-treated with 10 μM PES. Total cell proteins were probed with anti-Ser345 CHK1, anti-CHK1, anti-phospho-Tyr 15 CDK1, anti-CDK1, Cyclin B1 antibodies, and anti-vinculin as the loading control.

    Article Snippet: Etoposide, PFT-α and PES were purchased from Calbiochem (La Jolla, CA, USA).

    Techniques: Flow Cytometry, Cytometry, Isolation, Expressing, Reverse Transcription Polymerase Chain Reaction

    Effect of drugs on p53 acetylation and complex formation. a MEFs were either left untreated (Con) or treated with etoposide (Etop) alone or in combination with either PFT-α or PES for 3 h. Total cell extracts were prepared. The blot was probed with anti-Lys373/382 p53 antibody or anti-p53 as a loading control. b MEFs were either untreated (Con) or treated with etoposide (Etop) for 3 h, or pre-treated with either 30 µM of PFT-α or 10 μM of PES followed by 1.5 μM of etoposide for 3 h. Total cell extracts were immunoprecipitated with anti-p53 antibody and probed for HDAC1 and Mdm2. The first lane (IP Con IgG) represents PES and etoposide-treated extracts immunoprecipitated with rabbit IgG alone to indicate position of the IgG heavy chain (IgGH). Input Mdm2 represents the starting material unbound to the Agarose G beads, probed for the presence of Mdm2.

    Journal: Cancer Cell International

    Article Title: Etoposide induces cell death via mitochondrial-dependent actions of p53

    doi: 10.1186/s12935-015-0231-z

    Figure Lengend Snippet: Effect of drugs on p53 acetylation and complex formation. a MEFs were either left untreated (Con) or treated with etoposide (Etop) alone or in combination with either PFT-α or PES for 3 h. Total cell extracts were prepared. The blot was probed with anti-Lys373/382 p53 antibody or anti-p53 as a loading control. b MEFs were either untreated (Con) or treated with etoposide (Etop) for 3 h, or pre-treated with either 30 µM of PFT-α or 10 μM of PES followed by 1.5 μM of etoposide for 3 h. Total cell extracts were immunoprecipitated with anti-p53 antibody and probed for HDAC1 and Mdm2. The first lane (IP Con IgG) represents PES and etoposide-treated extracts immunoprecipitated with rabbit IgG alone to indicate position of the IgG heavy chain (IgGH). Input Mdm2 represents the starting material unbound to the Agarose G beads, probed for the presence of Mdm2.

    Article Snippet: Etoposide, PFT-α and PES were purchased from Calbiochem (La Jolla, CA, USA).

    Techniques: Immunoprecipitation

    Concentration-dependent etoposide-induced apoptosis. a Flow cytometry analysis of MEFs was performed to determine the percentage of cells with DNA content below the threshold for cells in G1, following 18 h treatment with 1.5, 15 or 150 µM etoposide. Data shown are mean + SD from seven independent experiments with three replicates each. b MEFs were treated with 1.5, 15 or 150 µM etoposide for 3, 6 or 18 h. Con indicates extract from untreated cells. Total cell proteins were probed with anti-Caspase-3 antibody. Vinculin was used as the loading control. Figures are representative of three independent experiments showing similar results.

    Journal: Cancer Cell International

    Article Title: Etoposide induces cell death via mitochondrial-dependent actions of p53

    doi: 10.1186/s12935-015-0231-z

    Figure Lengend Snippet: Concentration-dependent etoposide-induced apoptosis. a Flow cytometry analysis of MEFs was performed to determine the percentage of cells with DNA content below the threshold for cells in G1, following 18 h treatment with 1.5, 15 or 150 µM etoposide. Data shown are mean + SD from seven independent experiments with three replicates each. b MEFs were treated with 1.5, 15 or 150 µM etoposide for 3, 6 or 18 h. Con indicates extract from untreated cells. Total cell proteins were probed with anti-Caspase-3 antibody. Vinculin was used as the loading control. Figures are representative of three independent experiments showing similar results.

    Article Snippet: Etoposide, PFT-α and PES were purchased from Calbiochem (La Jolla, CA, USA).

    Techniques: Concentration Assay, Flow Cytometry, Cytometry

    Etoposide effect on p53-mediated transcriptional events. a MEFs were treated with 15 µM of etoposide ( upper panel ) or 1.5 µM of etoposide ( lower panel ) for 1, 3, 6 or 18 h. Total cell proteins were probed with anti-PUMA antibody. Vinculin or actin was used as the loading control. b MEFS were treated with 15 µM of etoposide ( upper panel ) or 1.5 µM of etoposide ( lower panel ) for 30 min, 1, 3, 6 or 18 h. Total RNA was isolated and reverse transcribed into cDNA. The expression of p21 CIP1/WAF1 was determined by RT-PCR using specific primers. β-actin was used a loading control. Figures are representative of three independent experiments.

    Journal: Cancer Cell International

    Article Title: Etoposide induces cell death via mitochondrial-dependent actions of p53

    doi: 10.1186/s12935-015-0231-z

    Figure Lengend Snippet: Etoposide effect on p53-mediated transcriptional events. a MEFs were treated with 15 µM of etoposide ( upper panel ) or 1.5 µM of etoposide ( lower panel ) for 1, 3, 6 or 18 h. Total cell proteins were probed with anti-PUMA antibody. Vinculin or actin was used as the loading control. b MEFS were treated with 15 µM of etoposide ( upper panel ) or 1.5 µM of etoposide ( lower panel ) for 30 min, 1, 3, 6 or 18 h. Total RNA was isolated and reverse transcribed into cDNA. The expression of p21 CIP1/WAF1 was determined by RT-PCR using specific primers. β-actin was used a loading control. Figures are representative of three independent experiments.

    Article Snippet: Etoposide, PFT-α and PES were purchased from Calbiochem (La Jolla, CA, USA).

    Techniques: Isolation, Expressing, Reverse Transcription Polymerase Chain Reaction

    Effect of PES on etoposide-induced cell death. a MEFs were either untreated (Con) or treated with etoposide (Etop) for 6 h, or pre-treated with 10 μM of PES followed by 1.5 μM of etoposide for 6 h. Mitochondrial extracts were immunoprecipitated with anti-p53 antibody and probed for Bcl-xL. The first lane (IP Con IgG) represents mitochondrial extracts from untreated (Con) cells immunoprecipitated with rabbit IgG alone Input p53 represents the starting material unbound to the Agarose G beads, probed for the presence of p53. b MEFs were treated with either 1.5 µM etoposide alone or pre-treated with 10 μM of PES and followed by 1.5 µM etoposide for various times. Columns represent percentage of cells having sub-G1 DNA content. A one-way ANOVA was carried out to compare the treatment groups. Post hoc comparison using the Tukey HSD test indicated significant inhibitory effect of PES treatment on the percentage of cells having sub-G1 DNA content. Data shown are mean + SD from 3 independent experiments with three replicates each. c Analysis of MEFs was done as in B; cells were pre-treated with 10 μM PES followed by 18 h treatment with 1.5, 15 or 150 µM etoposide. Cells were treated in parallel with UVC and followed for 18 h. Statistical analysis was as in b . Data are mean + SD of three repeat analyses.

    Journal: Cancer Cell International

    Article Title: Etoposide induces cell death via mitochondrial-dependent actions of p53

    doi: 10.1186/s12935-015-0231-z

    Figure Lengend Snippet: Effect of PES on etoposide-induced cell death. a MEFs were either untreated (Con) or treated with etoposide (Etop) for 6 h, or pre-treated with 10 μM of PES followed by 1.5 μM of etoposide for 6 h. Mitochondrial extracts were immunoprecipitated with anti-p53 antibody and probed for Bcl-xL. The first lane (IP Con IgG) represents mitochondrial extracts from untreated (Con) cells immunoprecipitated with rabbit IgG alone Input p53 represents the starting material unbound to the Agarose G beads, probed for the presence of p53. b MEFs were treated with either 1.5 µM etoposide alone or pre-treated with 10 μM of PES and followed by 1.5 µM etoposide for various times. Columns represent percentage of cells having sub-G1 DNA content. A one-way ANOVA was carried out to compare the treatment groups. Post hoc comparison using the Tukey HSD test indicated significant inhibitory effect of PES treatment on the percentage of cells having sub-G1 DNA content. Data shown are mean + SD from 3 independent experiments with three replicates each. c Analysis of MEFs was done as in B; cells were pre-treated with 10 μM PES followed by 18 h treatment with 1.5, 15 or 150 µM etoposide. Cells were treated in parallel with UVC and followed for 18 h. Statistical analysis was as in b . Data are mean + SD of three repeat analyses.

    Article Snippet: Etoposide, PFT-α and PES were purchased from Calbiochem (La Jolla, CA, USA).

    Techniques: Immunoprecipitation

    Effect of PFT-α on etoposide-induced apoptosis. a MEFs, pre-treated with 30 μM PFT-α followed by 18 h treatment with 1.5, 15 or 150 µM etoposide, were analyzed by flow cytometry to determine percentage of cells having sub-G1 DNA content. Cells were treated in parallel with UVC and analyzed after 18 h. Control indicates normally proliferating cells. Columns represent percentage of cells having sub-G1 DNA content, as analyzed by flow cytometry. Data are mean + SD of three independent experiments with three replicates each. b mRNA was extracted from untreated (0 h), 6 and 18 h treatment with 15 µM of etoposide ( black bars ) or pre-treatment with PFT-α followed by etoposide ( open bars ). The relative expression of p21 cip1/waf1 was determined by qRT-PCR (mean ± SD from three independent experiments). The values of p21 cip1/waf1 were normalized to β-actin. c , d Parallel studies used 1.5 or 15 µM etoposide. Sub-G1 population of MEFs was measured as in A for cells that were untreated (Control), pre-treated with 30 μM PFT-α followed by etoposide for 3 h (PFT-α-Etop 3 h), etoposide alone for 19 h (Etop 19 h), pre-treated with PFT-α followed by etoposide for 19 h (PFT-α-Etop 19 h), washed after 3 h of co-treatment, followed by further incubation for 18 h (PFT-α-Etop washed) and washed after 3 h, and PFT-α was added back for 18 h (PFT-α-Etop + PFT-α). Columns represent percentage of cells having sub-G1 DNA content; representative of three experiments with similar results.

    Journal: Cancer Cell International

    Article Title: Etoposide induces cell death via mitochondrial-dependent actions of p53

    doi: 10.1186/s12935-015-0231-z

    Figure Lengend Snippet: Effect of PFT-α on etoposide-induced apoptosis. a MEFs, pre-treated with 30 μM PFT-α followed by 18 h treatment with 1.5, 15 or 150 µM etoposide, were analyzed by flow cytometry to determine percentage of cells having sub-G1 DNA content. Cells were treated in parallel with UVC and analyzed after 18 h. Control indicates normally proliferating cells. Columns represent percentage of cells having sub-G1 DNA content, as analyzed by flow cytometry. Data are mean + SD of three independent experiments with three replicates each. b mRNA was extracted from untreated (0 h), 6 and 18 h treatment with 15 µM of etoposide ( black bars ) or pre-treatment with PFT-α followed by etoposide ( open bars ). The relative expression of p21 cip1/waf1 was determined by qRT-PCR (mean ± SD from three independent experiments). The values of p21 cip1/waf1 were normalized to β-actin. c , d Parallel studies used 1.5 or 15 µM etoposide. Sub-G1 population of MEFs was measured as in A for cells that were untreated (Control), pre-treated with 30 μM PFT-α followed by etoposide for 3 h (PFT-α-Etop 3 h), etoposide alone for 19 h (Etop 19 h), pre-treated with PFT-α followed by etoposide for 19 h (PFT-α-Etop 19 h), washed after 3 h of co-treatment, followed by further incubation for 18 h (PFT-α-Etop washed) and washed after 3 h, and PFT-α was added back for 18 h (PFT-α-Etop + PFT-α). Columns represent percentage of cells having sub-G1 DNA content; representative of three experiments with similar results.

    Article Snippet: Etoposide, PFT-α and PES were purchased from Calbiochem (La Jolla, CA, USA).

    Techniques: Flow Cytometry, Cytometry, Expressing, Quantitative RT-PCR, Incubation