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    Structured Review

    Millipore dimethyl sulfoxide dmso
    Both <t>resveratrol</t> and JNK inhibitor SP600125 inhibited TNFα-induced IL6 and IL8 mRNA expression in DPSCs. A and B: DPSCs were pretreated with <t>DMSO</t> (0 μM RSV), 50 μM RSV, 100 μM RSV for 1 h followed by TNFα treatment
    Dimethyl Sulfoxide Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 439 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Resveratrol represses tumor necrosis factor α/c-Jun N-terminal kinase signaling via autophagy in human dental pulp stem cells"

    Article Title: Resveratrol represses tumor necrosis factor α/c-Jun N-terminal kinase signaling via autophagy in human dental pulp stem cells

    Journal: Archives of oral biology

    doi: 10.1016/j.archoralbio.2018.10.020

    Both resveratrol and JNK inhibitor SP600125 inhibited TNFα-induced IL6 and IL8 mRNA expression in DPSCs. A and B: DPSCs were pretreated with DMSO (0 μM RSV), 50 μM RSV, 100 μM RSV for 1 h followed by TNFα treatment
    Figure Legend Snippet: Both resveratrol and JNK inhibitor SP600125 inhibited TNFα-induced IL6 and IL8 mRNA expression in DPSCs. A and B: DPSCs were pretreated with DMSO (0 μM RSV), 50 μM RSV, 100 μM RSV for 1 h followed by TNFα treatment

    Techniques Used: Expressing

    Resveratrol (RSV) inhibited TNFα-induced JNK activation but not NF-κB activation in DPSCs. A: DPSCs were pretreated with DMSO only (0 μM RSV), 50 μM RSV, 100 μM RSV for 1 h followed by TNFα treatment at
    Figure Legend Snippet: Resveratrol (RSV) inhibited TNFα-induced JNK activation but not NF-κB activation in DPSCs. A: DPSCs were pretreated with DMSO only (0 μM RSV), 50 μM RSV, 100 μM RSV for 1 h followed by TNFα treatment at

    Techniques Used: Activation Assay

    2) Product Images from "Docosahexaenoic acid-induced unfolded protein response, cell cycle arrest, and apoptosis in vascular smooth muscle cells are triggered by Ca2+-dependent induction of oxidative stress"

    Article Title: Docosahexaenoic acid-induced unfolded protein response, cell cycle arrest, and apoptosis in vascular smooth muscle cells are triggered by Ca2+-dependent induction of oxidative stress

    Journal: Free Radical Biology & Medicine

    doi: 10.1016/j.freeradbiomed.2012.02.036

    Free radical scavenger Tempol attenuates DHA-induced oxidative stress and UPR and improves cell proliferation. (A) Cells were loaded with 10 μM H 2 DCFDA for 30 min followed by exposure to veh or DHA (100 μM) for indicated time periods up to 5 h, followed by flow cytometry. For the 24-h time point cells were incubated with veh or DHA for 23.5 h and then loaded with H 2 DCFDA for 30 min in the presence of veh or DHA, followed by flow cytometry. (B) Cells were preincubated with Tempol (150 μM) or veh (DMSO) for 2 h (90 min before loading and during 30-min loading with H 2 DCFDA). Thereafter, the cells were exposed to DHA in the presence or absence of Tempol for 3 h, followed by ROS measurement. (C) [ 3 H]Thymidine incorporation in veh- or DHA-treated hPASMCs in the absence and presence of Tempol. (D) Representative Western blot of cyclin D1 protein expression in veh- or DHA-treated hPASMCs in the presence and absence of Tempol. (E) Representative Western blot of phosphorylated eIF2α in veh- or DHA-treated hPASMCs in the presence and absence of Tempol. Total eIF2α served as loading control. Data are given as means ± SEM of at least five independent experiments. * P
    Figure Legend Snippet: Free radical scavenger Tempol attenuates DHA-induced oxidative stress and UPR and improves cell proliferation. (A) Cells were loaded with 10 μM H 2 DCFDA for 30 min followed by exposure to veh or DHA (100 μM) for indicated time periods up to 5 h, followed by flow cytometry. For the 24-h time point cells were incubated with veh or DHA for 23.5 h and then loaded with H 2 DCFDA for 30 min in the presence of veh or DHA, followed by flow cytometry. (B) Cells were preincubated with Tempol (150 μM) or veh (DMSO) for 2 h (90 min before loading and during 30-min loading with H 2 DCFDA). Thereafter, the cells were exposed to DHA in the presence or absence of Tempol for 3 h, followed by ROS measurement. (C) [ 3 H]Thymidine incorporation in veh- or DHA-treated hPASMCs in the absence and presence of Tempol. (D) Representative Western blot of cyclin D1 protein expression in veh- or DHA-treated hPASMCs in the presence and absence of Tempol. (E) Representative Western blot of phosphorylated eIF2α in veh- or DHA-treated hPASMCs in the presence and absence of Tempol. Total eIF2α served as loading control. Data are given as means ± SEM of at least five independent experiments. * P

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

    3) Product Images from "H2S mediates carotid body response to hypoxia but not anoxia"

    Article Title: H2S mediates carotid body response to hypoxia but not anoxia

    Journal: Respiratory physiology & neurobiology

    doi: 10.1016/j.resp.2018.08.001

    Comparison of DL-PAG efficacy dissolved in saline versus DMSO. Average data (mean ± SEM; n=5 run in triplicate) of the effects of DL-PAG dissolved in saline ( A ) or DMSO ( B ) on CB H 2 S levels. C . Average data (mean ± SEM) of hypoxia (Hx; pO 2 = 37± 3mmHg)-evoked [Ca 2+ ] i responses of glomus cells treated with DL-PAG (50μM) dissolved either in saline (n=40 cells) or DMSO (n=37 cells), and presented as stimulus-evoked response minus baseline levels (Δ[Ca 2+ ] i , nM). *, P
    Figure Legend Snippet: Comparison of DL-PAG efficacy dissolved in saline versus DMSO. Average data (mean ± SEM; n=5 run in triplicate) of the effects of DL-PAG dissolved in saline ( A ) or DMSO ( B ) on CB H 2 S levels. C . Average data (mean ± SEM) of hypoxia (Hx; pO 2 = 37± 3mmHg)-evoked [Ca 2+ ] i responses of glomus cells treated with DL-PAG (50μM) dissolved either in saline (n=40 cells) or DMSO (n=37 cells), and presented as stimulus-evoked response minus baseline levels (Δ[Ca 2+ ] i , nM). *, P

    Techniques Used:

    Related Articles

    MTT Assay:

    Article Title: Shikonin Attenuates Acetaminophen-Induced Hepatotoxicity by Upregulation of Nrf2 through Akt/GSK3β Signaling
    Article Snippet: .. SHK, dimethyl sulfoxide (DMSO), LY294002, Lithium chloride (LiCl, a conventional GSK3β inhibitor), 2′,7′-Dichlorofluorescin diacetate (DCFH-DA) and 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide (MTT) were obtained from Sigma-Aldrich (St. Louis, MO, USA). ..

    Transfection:

    Article Title: Resveratrol represses tumor necrosis factor α/c-Jun N-terminal kinase signaling via autophagy in human dental pulp stem cells
    Article Snippet: Minimum Essential Medium (MEM)-α without ascorbic acid (A10490), Gibco™Fetal Bovine Serum (FBS, A3160401), OPTI-MEM I (31985–070), Lipofectamine RNAiMAX transfection reagent (A13778100), ECL Western Blotting substrate (#32106), Alexa Fluor® 488 goat anti-rabbit IgG (H + L) conjugate (A-11008), Alexa Fluor® 633 phalloidin (A22284), and ProLong™Gold Antifade Mountant with 4′,6-diamidino-2-phenylindole (DAPI) ( ) were obtained from Thermo Fisher Scientific (Waltham, MA, USA). .. Dimethyl sulfoxide (DMSO), resveratrol (R5010) and c-Jun N-terminal kinase (JNK) inhibitor SP600125 were obtained from Sigma-Aldrich (St. Louis, MO, USA).

    Modification:

    Article Title: In vitro assessment of bio-augmented minerals from peanut oil cakes fermented by Aspergillus oryzae through Caco-2 cells
    Article Snippet: .. Dulbeccos Modified Eagles Medium DMEM, l -glutamine, dimethyl sulfoxide (DMSO), Trypsin EDTA solution (0.25%), phenol red and streptomycin sulfate salt were procured from Sigma Aldrich, St. Louis, MO, USA. .. Human ferritin ELISA kit (SEA518Hu 96 Tests) from Cloud Clone-Corp. Houston, USA.

    Article Title: Circlular RNA BARD1 (Hsa_circ_0001098) overexpression in breast cancer cells with TCDD treatment could promote cell apoptosis via miR-3942/BARD1 axis
    Article Snippet: Mixture of DMEM (Dulbecco’s modified Eagle’s medium, Thermo Fisher Scientific, Waltham, USA) supplemented with 10% FBS, 100 units/mL penicillin and 100 ug/mL streptomycin (Sigma-Aldrich, St. Louis, MO, USA) were used to culture the cells and were stored in a humidified incubator with 5% CO2 at 37°C. .. At 70% confluency, cells were cultured with either dimethyl sulfoxide (DMSO) or 10 nM TCDD (Sigma-Aldrich, St. Louis, MO, USA) diluted in DMSO for 24 hours.

    Enzyme-linked Immunosorbent Assay:

    Article Title: In vitro assessment of bio-augmented minerals from peanut oil cakes fermented by Aspergillus oryzae through Caco-2 cells
    Article Snippet: Dulbeccos Modified Eagles Medium DMEM, l -glutamine, dimethyl sulfoxide (DMSO), Trypsin EDTA solution (0.25%), phenol red and streptomycin sulfate salt were procured from Sigma Aldrich, St. Louis, MO, USA. .. Human ferritin ELISA kit (SEA518Hu 96 Tests) from Cloud Clone-Corp. Houston, USA.

    Isolation:

    Article Title: The impact of skin banking and the use of its cadaveric skin allografts for severe burn victims in Singapore.
    Article Snippet: Random small skin samples are also isolated for microbiology testing at this step. .. Skin processing All the folded gauze is subsequently soaked in DMEM with 10% dimethyl sulfoxide (DMSO) [Sigma] to allow penetration of the cryoprotective agent to the tissues.

    Cell Culture:

    Article Title: In vitro assessment of bio-augmented minerals from peanut oil cakes fermented by Aspergillus oryzae through Caco-2 cells
    Article Snippet: Paragraph title: Cell culture ... Dulbeccos Modified Eagles Medium DMEM, l -glutamine, dimethyl sulfoxide (DMSO), Trypsin EDTA solution (0.25%), phenol red and streptomycin sulfate salt were procured from Sigma Aldrich, St. Louis, MO, USA.

    Article Title: Circlular RNA BARD1 (Hsa_circ_0001098) overexpression in breast cancer cells with TCDD treatment could promote cell apoptosis via miR-3942/BARD1 axis
    Article Snippet: .. At 70% confluency, cells were cultured with either dimethyl sulfoxide (DMSO) or 10 nM TCDD (Sigma-Aldrich, St. Louis, MO, USA) diluted in DMSO for 24 hours. .. The concentration of TCDD has been proved to reproducibly cause differential gene expression in variety of cells.

    SYBR Green Assay:

    Article Title: A systematic comparison of lipopolymers for siRNA delivery to multiple breast cancer cell lines: In vitro studies.
    Article Snippet: Small interfering RNA (siRNA) therapy is a promising approach for treatment of a wide range of cancers, including breast cancers that display variable phenotypic features. .. Small interfering RNA (siRNA) therapy is a promising approach for treatment of a wide range of cancers, including breast cancers that display variable phenotypic features.

    Concentration Assay:

    Article Title: Glycogen synthase kinase-3β inhibition in the medial prefrontal cortex mediates paradoxical amphetamine action in a mouse model of ADHD
    Article Snippet: Drugs and Doses d-Amphetamine hemisulfate (Amph), methylphenidate hydrochloride, lithium chloride (LiCl), TDZD-8, and dimethyl sulfoxide (DMSO) were from Sigma-Aldrich (USA). .. TDZD-8 stock solution was prepared in 100% DMSO and then dissolved in saline to achieve a final DMSO concentration of 0.5% v/v.

    Article Title: Circlular RNA BARD1 (Hsa_circ_0001098) overexpression in breast cancer cells with TCDD treatment could promote cell apoptosis via miR-3942/BARD1 axis
    Article Snippet: At 70% confluency, cells were cultured with either dimethyl sulfoxide (DMSO) or 10 nM TCDD (Sigma-Aldrich, St. Louis, MO, USA) diluted in DMSO for 24 hours. .. The concentration of TCDD has been proved to reproducibly cause differential gene expression in variety of cells.

    Cell Counting:

    Article Title: Continuous porcine cell lines developed from alveolar macrophages: partial characterization and virus susceptibility.
    Article Snippet: Viability after freezing Suspensions containing 10 7 cells/ml were frozen in freezing medium containing 10% dimethyl sulfoxide (DMSO) (Sigma) and 10% RPMI in FBS for 4 weeks in liquid nitrogen. .. The dead cell count at 18 h post seeding was deducted from the viable count immediately after thawing and used to estimate cell attachment/survival 18 h after seeding.

    other:

    Article Title: H2S mediates carotid body response to hypoxia but not anoxia
    Article Snippet: Aminooxyacetic acid (AOAA), Catalase, Sodium dithionite, DL-propargylglycine (DL-PAG), Dimethyl sulfoxide (DMSO), and Glucose Oxidase were obtained from commercial sources (Sigma Aldrich).

    Expressing:

    Article Title: Circlular RNA BARD1 (Hsa_circ_0001098) overexpression in breast cancer cells with TCDD treatment could promote cell apoptosis via miR-3942/BARD1 axis
    Article Snippet: At 70% confluency, cells were cultured with either dimethyl sulfoxide (DMSO) or 10 nM TCDD (Sigma-Aldrich, St. Louis, MO, USA) diluted in DMSO for 24 hours. .. The concentration of TCDD has been proved to reproducibly cause differential gene expression in variety of cells.

    Western Blot:

    Article Title: Resveratrol represses tumor necrosis factor α/c-Jun N-terminal kinase signaling via autophagy in human dental pulp stem cells
    Article Snippet: Minimum Essential Medium (MEM)-α without ascorbic acid (A10490), Gibco™Fetal Bovine Serum (FBS, A3160401), OPTI-MEM I (31985–070), Lipofectamine RNAiMAX transfection reagent (A13778100), ECL Western Blotting substrate (#32106), Alexa Fluor® 488 goat anti-rabbit IgG (H + L) conjugate (A-11008), Alexa Fluor® 633 phalloidin (A22284), and ProLong™Gold Antifade Mountant with 4′,6-diamidino-2-phenylindole (DAPI) ( ) were obtained from Thermo Fisher Scientific (Waltham, MA, USA). .. Dimethyl sulfoxide (DMSO), resveratrol (R5010) and c-Jun N-terminal kinase (JNK) inhibitor SP600125 were obtained from Sigma-Aldrich (St. Louis, MO, USA).

    Polymerase Chain Reaction:

    Article Title: Docosahexaenoic acid-induced unfolded protein response, cell cycle arrest, and apoptosis in vascular smooth muscle cells are triggered by Ca2+-dependent induction of oxidative stress
    Article Snippet: Taq polymerase was from Solis Biodyne (Tartu, Estonia), 10× PCR buffer and dNTP mix were from Promega (Madison, WI, USA). .. DHA, dimethyl sulfoxide (DMSO), ethanol, CaCl2 , KCl, MgCl2 , NaCl, Hepes, EGTA, glucose, 4-hydroxy-Tempol (Tempol), bovine serum albumin (BSA), Triton X-100, and propidium iodide were from Sigma (Steinheim, Germany).

    Lysis:

    Article Title: INHIBITORY ACTIVITY OF MANGIFERIN ON HELICOBACTER PYLORI-INDUCED INFLAMMATION IN HUMAN GASTRIC CARCINOMA AGS CELLS
    Article Snippet: .. Chemicals Mangiferin (98.0%), trypsin, ethylene-diamine-tetraacetic acid (EDTA), glutamine, cell lysis buffer, urease reagent, isopropanol, dimethyl sulfoxide (DMSO), and formalin were purchased from Sigma-Aldrich (MO, USA). .. F12 Hans medium, fetal bovine serum (FBS), penicillin-streptomycin, amoxicillin (AMX), phosphate buffered saline (PBS) were bought from Thermo Fisher Scientific Gibco (NY, USA).

    Recombinant:

    Article Title: Resveratrol represses tumor necrosis factor α/c-Jun N-terminal kinase signaling via autophagy in human dental pulp stem cells
    Article Snippet: Dimethyl sulfoxide (DMSO), resveratrol (R5010) and c-Jun N-terminal kinase (JNK) inhibitor SP600125 were obtained from Sigma-Aldrich (St. Louis, MO, USA). .. Recombinant human TNFα was purchased from R & D Systems (Minneapolis, MN, USA).

    Molecular Weight:

    Article Title: A systematic comparison of lipopolymers for siRNA delivery to multiple breast cancer cell lines: In vitro studies.
    Article Snippet: .. Small interfering RNA (siRNA) therapy is a promising approach for treatment of a wide range of cancers, including breast cancers that display variable phenotypic features. .. Small interfering RNA (siRNA) therapy is a promising approach for treatment of a wide range of cancers, including breast cancers that display variable phenotypic features.

    Staining:

    Article Title: STRIPAK directs PP2A activity toward MAP4K4 to promote oncogenic transformation of human cells
    Article Snippet: After 4 to 5 weeks, cells were stained with 0.005% crystal violet (Sigma) in PBS and colonies were counted. .. For MAP4K4 inhibitor experiments, dimethyl sulfoxide (DMSO) (Sigma) or inhibitor (compound 29) ( ) were used at the indicated concentrations in both the bottom and top soft agar layers and included in refeedings.

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    Millipore dmso
    Inhibition of the proteasome disrupts neuronal differentiation. ( A–D ) Hippocampal neurons were cultured in the presence of solvent <t>(DMSO),</t> 1.5 μM clasto-Lactacystin β-Lactone (Lactacystin), 40 μM <t>ALLN,</t> or 1.5 μM MG-132 (all dissolved in DMSO) for 48 h and analysed at 3 d.i.v. (stage 3) by staining with an anti-MAP2 antibody (A; red), the Tau-1 (blue), and an anti-Rap1 (red) antibody. Lactacystin induced the formation of multiple axons (asterisks) whose growth cones were all positive for Rap1B (A, arrows). Insets show higher magnifications of the marked growth cones. Axons identified by Tau-1 immunoreactivity are marked by asterisks. The scale bar is 12 μm. (B–D) The effect of Lactacystin (L), ALLN (A), or MG-132 (M) was analysed by determining the number of axons (B) or minor neurites (C) per cell and the length of axons (E) (means±s.e.m.; * P
    Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 24 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dmso/product/Millipore
    Average 99 stars, based on 24 article reviews
    Price from $9.99 to $1999.99
    dmso - by Bioz Stars, 2020-04
    99/100 stars
      Buy from Supplier

    92
    Millipore dmso stock
    Mef2d is necessary for the activating effect of <t>Fgf</t> on Myod expression at the neurula stage. (A) Embryos were injected unilaterally either with Fgf8b synthetic mRNA alone (5 pg), or with moMyod1 or with moMef2d1, fixed at the gastrula stage and submitted to in situ hybridization for Xbra , Myod or Mef2d . Co-injection of Mef2dF mRNA with moMef2d1 was able to rescue the phenotype of moMef2d1 embryos (+Mef2dF). (B) Unilateral injection of Fgf8b mRNA induced the lateral expansion of Mef2d expression domain at stage 16. (C) Embryos were injected unilaterally with 20 ng of XIMOF8 and fixed at stage 16. Co-injection with Mef2dF mRNA was able to rescue the phenotype of XIMOF8 embryos (+Mef2dF). (D) Embryos were injected at stage 11.5/12 with <t>DMSO</t> or 5 mM SU5402, an inhibitor of Fgf signaling, and fixed at stage 16. A first unilateral injection of Mef2dF mRNA at the two-cell stage was able to rescue the phenotype of treated embryos (+Mef2dF). Injected side (*) at the bottom except in (A), on the left. Bracket indicates the position of lateral myogenic cells. Probes are in a framed box and indicated for each panel. For complete statistical data, see supporting information, figure S2 .
    Dmso Stock, supplied by Millipore, used in various techniques. Bioz Stars score: 92/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/dmso stock/product/Millipore
    Average 92 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    dmso stock - by Bioz Stars, 2020-04
    92/100 stars
      Buy from Supplier

    Image Search Results


    Inhibition of the proteasome disrupts neuronal differentiation. ( A–D ) Hippocampal neurons were cultured in the presence of solvent (DMSO), 1.5 μM clasto-Lactacystin β-Lactone (Lactacystin), 40 μM ALLN, or 1.5 μM MG-132 (all dissolved in DMSO) for 48 h and analysed at 3 d.i.v. (stage 3) by staining with an anti-MAP2 antibody (A; red), the Tau-1 (blue), and an anti-Rap1 (red) antibody. Lactacystin induced the formation of multiple axons (asterisks) whose growth cones were all positive for Rap1B (A, arrows). Insets show higher magnifications of the marked growth cones. Axons identified by Tau-1 immunoreactivity are marked by asterisks. The scale bar is 12 μm. (B–D) The effect of Lactacystin (L), ALLN (A), or MG-132 (M) was analysed by determining the number of axons (B) or minor neurites (C) per cell and the length of axons (E) (means±s.e.m.; * P

    Journal: The EMBO Journal

    Article Title: Ubiquitination of the GTPase Rap1B by the ubiquitin ligase Smurf2 is required for the establishment of neuronal polarity

    doi: 10.1038/sj.emboj.7601580

    Figure Lengend Snippet: Inhibition of the proteasome disrupts neuronal differentiation. ( A–D ) Hippocampal neurons were cultured in the presence of solvent (DMSO), 1.5 μM clasto-Lactacystin β-Lactone (Lactacystin), 40 μM ALLN, or 1.5 μM MG-132 (all dissolved in DMSO) for 48 h and analysed at 3 d.i.v. (stage 3) by staining with an anti-MAP2 antibody (A; red), the Tau-1 (blue), and an anti-Rap1 (red) antibody. Lactacystin induced the formation of multiple axons (asterisks) whose growth cones were all positive for Rap1B (A, arrows). Insets show higher magnifications of the marked growth cones. Axons identified by Tau-1 immunoreactivity are marked by asterisks. The scale bar is 12 μm. (B–D) The effect of Lactacystin (L), ALLN (A), or MG-132 (M) was analysed by determining the number of axons (B) or minor neurites (C) per cell and the length of axons (E) (means±s.e.m.; * P

    Article Snippet: DMSO, ALLN, clasto-Lactacystin β-Lactone, and MG-132 (Calbiochem) were directly added to neuronal cultures 16 h after plating to a final concentration of 30, 40, 1.5, and 1.5 μM, respectively ( ).

    Techniques: Inhibition, Cell Culture, Staining

    TBZ inhibits angiogenesis in vivo in Xenopus embryos. Formation of Xenopus embryo veins is disrupted, marked by expression of vascular reporter genes (A, B) erg and (C, D) aplnr , contrasting treatment with 1% DMSO control only (A, C) with 1% DMSO, 250 µM TBZ, treated at stage 31 and imaged at stages 35–36 (B, D). PCV, posterior cardinal vein; ISV, intersomitic vein; VV, vitellin veins. Similarly, TBZ disrupts vasculature imaged within a living Xenopus embryo and visualized by vascular specific GFP in kdr:GFP frogs from [19] , contrasting the vasculature of stage 46 animals treated from stage 41 with the 1% DMSO control (E) or 1% DMSO, 250 µM TBZ (F). Scale bar, 200 µm.

    Journal: PLoS Biology

    Article Title: Evolutionarily Repurposed Networks Reveal the Well-Known Antifungal Drug Thiabendazole to Be a Novel Vascular Disrupting AgentEvolution-Based Drug Discovery: Antifungal Also Disrupts Blood Vessel Formation

    doi: 10.1371/journal.pbio.1001379

    Figure Lengend Snippet: TBZ inhibits angiogenesis in vivo in Xenopus embryos. Formation of Xenopus embryo veins is disrupted, marked by expression of vascular reporter genes (A, B) erg and (C, D) aplnr , contrasting treatment with 1% DMSO control only (A, C) with 1% DMSO, 250 µM TBZ, treated at stage 31 and imaged at stages 35–36 (B, D). PCV, posterior cardinal vein; ISV, intersomitic vein; VV, vitellin veins. Similarly, TBZ disrupts vasculature imaged within a living Xenopus embryo and visualized by vascular specific GFP in kdr:GFP frogs from [19] , contrasting the vasculature of stage 46 animals treated from stage 41 with the 1% DMSO control (E) or 1% DMSO, 250 µM TBZ (F). Scale bar, 200 µm.

    Article Snippet: Mass Spectrometry HUVECs were treated with 1% DMSO or 1% DMSO, 250 µM TBZ for 24 h, and lysed by Dounce homogenization in low salt buffer (10 mM Tris-HCl, pH 8.8, 10 mM KCl, 1.5 mM MgCl2 ) with 0.5 mM DTT and protease inhibitor mixture (Calbiochem).

    Techniques: In Vivo, Expressing

    TBZ significantly disrupts tube formation in cultured human umbilical vein endothelial cells (HUVECs), an in vitro capillary model. Here, we show effects of 1% DMSO-treated control (A) versus 1% DMSO, 100 µM TBZ (B) and 1% DMSO, 250 µM TBZ (C). Scale bar, 100 µm. (D) Tube disruption is dose-dependent and comparable to that from silencing known pro-angiogenic gene HOXA9 .

    Journal: PLoS Biology

    Article Title: Evolutionarily Repurposed Networks Reveal the Well-Known Antifungal Drug Thiabendazole to Be a Novel Vascular Disrupting AgentEvolution-Based Drug Discovery: Antifungal Also Disrupts Blood Vessel Formation

    doi: 10.1371/journal.pbio.1001379

    Figure Lengend Snippet: TBZ significantly disrupts tube formation in cultured human umbilical vein endothelial cells (HUVECs), an in vitro capillary model. Here, we show effects of 1% DMSO-treated control (A) versus 1% DMSO, 100 µM TBZ (B) and 1% DMSO, 250 µM TBZ (C). Scale bar, 100 µm. (D) Tube disruption is dose-dependent and comparable to that from silencing known pro-angiogenic gene HOXA9 .

    Article Snippet: Mass Spectrometry HUVECs were treated with 1% DMSO or 1% DMSO, 250 µM TBZ for 24 h, and lysed by Dounce homogenization in low salt buffer (10 mM Tris-HCl, pH 8.8, 10 mM KCl, 1.5 mM MgCl2 ) with 0.5 mM DTT and protease inhibitor mixture (Calbiochem).

    Techniques: Cell Culture, In Vitro

    TBZ impedes migration of HUVECs in a wound scratch assay, but treatment with the Rho Kinase inhibitor Y27632 reverses TBZ's effects. (A) The effects of 1% DMSO-treated control versus 1% DMSO, 250 µM TBZ, and 1% DMSO, 250 µM TBZ, 10 µM Y27632. Scale bar, 200 µm. (B) quantifies the dose-dependent suppression of TBZ inhibition by Y27632. Error bars represent the mean ± 1 s.d. across 3 wells (1 of 3 trials). TBZ results in disorganization of actin stress fibers, as shown in (C) for 1% DMSO-treated control versus 1% DMSO, 250 µM TBZ-treated cells. Scale bar, 20 µm.

    Journal: PLoS Biology

    Article Title: Evolutionarily Repurposed Networks Reveal the Well-Known Antifungal Drug Thiabendazole to Be a Novel Vascular Disrupting AgentEvolution-Based Drug Discovery: Antifungal Also Disrupts Blood Vessel Formation

    doi: 10.1371/journal.pbio.1001379

    Figure Lengend Snippet: TBZ impedes migration of HUVECs in a wound scratch assay, but treatment with the Rho Kinase inhibitor Y27632 reverses TBZ's effects. (A) The effects of 1% DMSO-treated control versus 1% DMSO, 250 µM TBZ, and 1% DMSO, 250 µM TBZ, 10 µM Y27632. Scale bar, 200 µm. (B) quantifies the dose-dependent suppression of TBZ inhibition by Y27632. Error bars represent the mean ± 1 s.d. across 3 wells (1 of 3 trials). TBZ results in disorganization of actin stress fibers, as shown in (C) for 1% DMSO-treated control versus 1% DMSO, 250 µM TBZ-treated cells. Scale bar, 20 µm.

    Article Snippet: Mass Spectrometry HUVECs were treated with 1% DMSO or 1% DMSO, 250 µM TBZ for 24 h, and lysed by Dounce homogenization in low salt buffer (10 mM Tris-HCl, pH 8.8, 10 mM KCl, 1.5 mM MgCl2 ) with 0.5 mM DTT and protease inhibitor mixture (Calbiochem).

    Techniques: Migration, Wound Healing Assay, Inhibition

    26S proteasomes and 19S proteasome caps are chaperones for denatured RTA ( A ) 26S proteasomes maintain the solubility of denatured RTA. Top panels: GdnHCl-denatured RTA (40 nM) was diluted in the absence (−) or presence (+) of 26S proteasomes (25 nM, equivalent to 40 nM 19S RP) and, after centrifugation, aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. Bottom panels: samples were treated in the same way as above, but were incubated at 37°C for 2 h before SDS/PAGE. ( B ) The solubilizing activity of the 26S proteasome is independent of its proteolytic activity. GdnHCl-denatured RTA (40 nM) was diluted in the absence (−) or presence (+) of 40 nM 26S proteasomes pretreated with the vehicle DMSO (top panel) or the proteasome inhibitor cLβ-l (bottom panel), and aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. ( C ) Efficacy of cLβ-l was confirmed in vitro by its ability to block degradation of casein (arrowhead) in the presence of mammalian 26S proteasomes. Casein (40 nM) was incubated in the presence (+) or absence (−) of 26S proteasomes that had been pretreated with vehicle DMSO or cLβ-l. ( D ) 20S proteasome cores do not maintain the solubility of denatured RTA. GdnHCl-denatured RTA (40 nM) was diluted in the absence (−) or presence of 40 nM 20S proteasomes (+ 20S) or 40 nM BSA (+ BSA) and, after centrifugation, aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. ( E ) 19S proteasome RP maintain the solubility of denatured RTA. Heat (45°C)-denatured and GdnHCl-denatured RTA (40 nM) were diluted in the absence (−) or presence (+) of 40 nM 19S RP and, after centrifugation, aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. ( F ) Catalytic activity can be recovered from proteasome-solubilized RTA. Native RTA (RTA), GdnHCl-denatured RTA (denRTA) and a mixture of denRTA and 19S RP were centrifuged to remove aggregates and the soluble fractions were incubated with 20 μg of yeast ribosomes for 2 h at 30°C. After cleavage of any depurinated 28S rRNA with acetic-aniline, rRNAs were extracted, electrophoresed in denaturing conditions (1.2% agarose/50% formamide), and the gel was stained with ethidium bromide for visualization. Aniline fragment, grey arrowhead; 5.8S rRNA, white arrowhead.

    Journal: Biochemical Journal

    Article Title: The proteasome cap RPT5/Rpt5p subunit prevents aggregation of unfolded ricin A chain

    doi: 10.1042/BJ20130133

    Figure Lengend Snippet: 26S proteasomes and 19S proteasome caps are chaperones for denatured RTA ( A ) 26S proteasomes maintain the solubility of denatured RTA. Top panels: GdnHCl-denatured RTA (40 nM) was diluted in the absence (−) or presence (+) of 26S proteasomes (25 nM, equivalent to 40 nM 19S RP) and, after centrifugation, aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. Bottom panels: samples were treated in the same way as above, but were incubated at 37°C for 2 h before SDS/PAGE. ( B ) The solubilizing activity of the 26S proteasome is independent of its proteolytic activity. GdnHCl-denatured RTA (40 nM) was diluted in the absence (−) or presence (+) of 40 nM 26S proteasomes pretreated with the vehicle DMSO (top panel) or the proteasome inhibitor cLβ-l (bottom panel), and aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. ( C ) Efficacy of cLβ-l was confirmed in vitro by its ability to block degradation of casein (arrowhead) in the presence of mammalian 26S proteasomes. Casein (40 nM) was incubated in the presence (+) or absence (−) of 26S proteasomes that had been pretreated with vehicle DMSO or cLβ-l. ( D ) 20S proteasome cores do not maintain the solubility of denatured RTA. GdnHCl-denatured RTA (40 nM) was diluted in the absence (−) or presence of 40 nM 20S proteasomes (+ 20S) or 40 nM BSA (+ BSA) and, after centrifugation, aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. ( E ) 19S proteasome RP maintain the solubility of denatured RTA. Heat (45°C)-denatured and GdnHCl-denatured RTA (40 nM) were diluted in the absence (−) or presence (+) of 40 nM 19S RP and, after centrifugation, aggregated (P) and soluble (S) fractions were examined by anti-RTA immunoblot after SDS/PAGE. ( F ) Catalytic activity can be recovered from proteasome-solubilized RTA. Native RTA (RTA), GdnHCl-denatured RTA (denRTA) and a mixture of denRTA and 19S RP were centrifuged to remove aggregates and the soluble fractions were incubated with 20 μg of yeast ribosomes for 2 h at 30°C. After cleavage of any depurinated 28S rRNA with acetic-aniline, rRNAs were extracted, electrophoresed in denaturing conditions (1.2% agarose/50% formamide), and the gel was stained with ethidium bromide for visualization. Aniline fragment, grey arrowhead; 5.8S rRNA, white arrowhead.

    Article Snippet: Agents, their final concentrations and vehicles were Pi1 [proteasome inhibitor 1 (Calbiochem); used at a final concentration of 50 μM with DMSO as a vehicle], the proteasome inhibitor ALLN [N -acetyl-L -leucyl-L -leucylnorleucinal (Calbiochem); used at a final concentration of 20 μM with DMSO as a vehicle], the proteasome inhibitor cLβ-l (clasto lactacystin β-lactone; used at a final concentration of 20 μM with DMSO as a vehicle), a mixture of the cathepsin inhibitors pepstatin and leupeptin (used at final concentrations of 100 μM and 1 μM respectively with water as a vehicle) and the secretion inhibitor BFA (brefeldin A; used at a final concentration of 10 μg/ml with ethanol as a vehicle).

    Techniques: Solubility, Centrifugation, SDS Page, Incubation, Activity Assay, In Vitro, Blocking Assay, Staining

    Inhibition of proteasome proteolytic activities sensitizes HeLa cells to ricin only after long incubations ( A ) HeLa cells were treated for 1, 2, 4, 6 or 8 h with graded doses of ricin in growth medium containing 1 μM Pi1 (●) or vehicle (DMSO, ○), and their subsequent ability to synthesize proteins was determined by measuring incorporation of [ 35 S]methionine into acid-precipitable material. Typical single assays are shown. ( B ) Top panel: cells were treated as described in ( A ), sensitivities to toxin (IC 50 , toxin concentration required to reduce protein synthesis to 50% that of non-toxin treated controls) were determined, and fold protection (IC 50 Pi1-treated cells/IC 50 DMSO-treated cells) is displayed. Middle and bottom panels: cells were treated as described above, substituting the proteasome inhibitor ALLN (middle panel) or a mixture of the cathepsin inhibitors leupeptin and pepstatin (L+P, bottom panel) for Pi1, and substituting water for the vehicle for L+P treatment. Values are means±S.D. for three independent experiments. Broken line, no protective effect over that of treatment with vehicle only; n.d., not determined. ( C ) Cells were treated with a saturating dose of ricin for 4 h in the presence (+) or absence (−) of Pi1 (top panel), L+P (middle panel) or the secretion inhibitor BFA (bottom panel). Detergent-soluble extracts were separated by SDS/PAGE, and RTA (black arrowhead) and a proteolytically clipped RTA (white arrowhead) were revealed by immunoblotting. ( D ) HeLa cells were treated for 2, 4 or 6 h as described in ( A ), substituting the proteasome inhibitor cLβ-l for Pi1. Values are means±S.D. for three independent experiments. Broken line, no protective effect over that of treatment with DMSO only. ( E ) Cells were treated with a single low dose of ricin (1 ng/ml , top panel) for increasing times in the presence of cLβ-l (●) or vehicle (DMSO, ○) and in parallel with cLβ-l (●) or vehicle (DMSO, ○) in the absence of ricin (bottom panel) and their subsequent ability to synthesize proteins was determined by measuring incorporation of [ 35 S]methionine into acid-precipitable material. Typical single assays are shown. Inset: toxin trafficking times, determined as in the top panel, in the presence of DMSO (D, ▼) or cLβ-l (c, ▽). Values are means±S.D. for three independent experiments.

    Journal: Biochemical Journal

    Article Title: The proteasome cap RPT5/Rpt5p subunit prevents aggregation of unfolded ricin A chain

    doi: 10.1042/BJ20130133

    Figure Lengend Snippet: Inhibition of proteasome proteolytic activities sensitizes HeLa cells to ricin only after long incubations ( A ) HeLa cells were treated for 1, 2, 4, 6 or 8 h with graded doses of ricin in growth medium containing 1 μM Pi1 (●) or vehicle (DMSO, ○), and their subsequent ability to synthesize proteins was determined by measuring incorporation of [ 35 S]methionine into acid-precipitable material. Typical single assays are shown. ( B ) Top panel: cells were treated as described in ( A ), sensitivities to toxin (IC 50 , toxin concentration required to reduce protein synthesis to 50% that of non-toxin treated controls) were determined, and fold protection (IC 50 Pi1-treated cells/IC 50 DMSO-treated cells) is displayed. Middle and bottom panels: cells were treated as described above, substituting the proteasome inhibitor ALLN (middle panel) or a mixture of the cathepsin inhibitors leupeptin and pepstatin (L+P, bottom panel) for Pi1, and substituting water for the vehicle for L+P treatment. Values are means±S.D. for three independent experiments. Broken line, no protective effect over that of treatment with vehicle only; n.d., not determined. ( C ) Cells were treated with a saturating dose of ricin for 4 h in the presence (+) or absence (−) of Pi1 (top panel), L+P (middle panel) or the secretion inhibitor BFA (bottom panel). Detergent-soluble extracts were separated by SDS/PAGE, and RTA (black arrowhead) and a proteolytically clipped RTA (white arrowhead) were revealed by immunoblotting. ( D ) HeLa cells were treated for 2, 4 or 6 h as described in ( A ), substituting the proteasome inhibitor cLβ-l for Pi1. Values are means±S.D. for three independent experiments. Broken line, no protective effect over that of treatment with DMSO only. ( E ) Cells were treated with a single low dose of ricin (1 ng/ml , top panel) for increasing times in the presence of cLβ-l (●) or vehicle (DMSO, ○) and in parallel with cLβ-l (●) or vehicle (DMSO, ○) in the absence of ricin (bottom panel) and their subsequent ability to synthesize proteins was determined by measuring incorporation of [ 35 S]methionine into acid-precipitable material. Typical single assays are shown. Inset: toxin trafficking times, determined as in the top panel, in the presence of DMSO (D, ▼) or cLβ-l (c, ▽). Values are means±S.D. for three independent experiments.

    Article Snippet: Agents, their final concentrations and vehicles were Pi1 [proteasome inhibitor 1 (Calbiochem); used at a final concentration of 50 μM with DMSO as a vehicle], the proteasome inhibitor ALLN [N -acetyl-L -leucyl-L -leucylnorleucinal (Calbiochem); used at a final concentration of 20 μM with DMSO as a vehicle], the proteasome inhibitor cLβ-l (clasto lactacystin β-lactone; used at a final concentration of 20 μM with DMSO as a vehicle), a mixture of the cathepsin inhibitors pepstatin and leupeptin (used at final concentrations of 100 μM and 1 μM respectively with water as a vehicle) and the secretion inhibitor BFA (brefeldin A; used at a final concentration of 10 μg/ml with ethanol as a vehicle).

    Techniques: Inhibition, Concentration Assay, SDS Page

    Mef2d is necessary for the activating effect of Fgf on Myod expression at the neurula stage. (A) Embryos were injected unilaterally either with Fgf8b synthetic mRNA alone (5 pg), or with moMyod1 or with moMef2d1, fixed at the gastrula stage and submitted to in situ hybridization for Xbra , Myod or Mef2d . Co-injection of Mef2dF mRNA with moMef2d1 was able to rescue the phenotype of moMef2d1 embryos (+Mef2dF). (B) Unilateral injection of Fgf8b mRNA induced the lateral expansion of Mef2d expression domain at stage 16. (C) Embryos were injected unilaterally with 20 ng of XIMOF8 and fixed at stage 16. Co-injection with Mef2dF mRNA was able to rescue the phenotype of XIMOF8 embryos (+Mef2dF). (D) Embryos were injected at stage 11.5/12 with DMSO or 5 mM SU5402, an inhibitor of Fgf signaling, and fixed at stage 16. A first unilateral injection of Mef2dF mRNA at the two-cell stage was able to rescue the phenotype of treated embryos (+Mef2dF). Injected side (*) at the bottom except in (A), on the left. Bracket indicates the position of lateral myogenic cells. Probes are in a framed box and indicated for each panel. For complete statistical data, see supporting information, figure S2 .

    Journal: PLoS ONE

    Article Title: Mef2d Acts Upstream of Muscle Identity Genes and Couples Lateral Myogenesis to Dermomyotome Formation in Xenopus laevis

    doi: 10.1371/journal.pone.0052359

    Figure Lengend Snippet: Mef2d is necessary for the activating effect of Fgf on Myod expression at the neurula stage. (A) Embryos were injected unilaterally either with Fgf8b synthetic mRNA alone (5 pg), or with moMyod1 or with moMef2d1, fixed at the gastrula stage and submitted to in situ hybridization for Xbra , Myod or Mef2d . Co-injection of Mef2dF mRNA with moMef2d1 was able to rescue the phenotype of moMef2d1 embryos (+Mef2dF). (B) Unilateral injection of Fgf8b mRNA induced the lateral expansion of Mef2d expression domain at stage 16. (C) Embryos were injected unilaterally with 20 ng of XIMOF8 and fixed at stage 16. Co-injection with Mef2dF mRNA was able to rescue the phenotype of XIMOF8 embryos (+Mef2dF). (D) Embryos were injected at stage 11.5/12 with DMSO or 5 mM SU5402, an inhibitor of Fgf signaling, and fixed at stage 16. A first unilateral injection of Mef2dF mRNA at the two-cell stage was able to rescue the phenotype of treated embryos (+Mef2dF). Injected side (*) at the bottom except in (A), on the left. Bracket indicates the position of lateral myogenic cells. Probes are in a framed box and indicated for each panel. For complete statistical data, see supporting information, figure S2 .

    Article Snippet: 20 nanoliters of DMSO solution or a 5 mM DMSO stock of the Fgf receptor kinase inhibitor SU5402 (Calbiochem) were injected into the archenteron.

    Techniques: Expressing, Injection, In Situ Hybridization