dimethyl sulfoxide dmso Millipore Search Results


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  • 99
    Millipore dimethyl sulfoxide dmso
    Suppressing Smad7 protein expression with Notch inhibitor N-(N-[3,5-difluorophenacetyl]-l-alanyl)-S-phenylglycine t-butyl ester facilitates epithelial–mesenchymal transition and growth arrest induced by transforming growth factor-beta. A : Western blot analysis of Smad 2/3 phosphorylation. Left planes: Transforming growth factor-beta (TGFβ1) time-dependently induces Smad2/3 phosphorylation in P1 cells. Right planes: P1 cells were treated with 10 ng/ml TGFβ1 for 30 and 40 min as positive control. P0 and P1 cells were pretreated with 10 µM N-(N-[3,5-difluorophenacetyl]-l-alanyl)-S-phenylglycine t-butyl ester <t>(DAPT)</t> or dimethyl sulfoxide <t>(DMSO)</t> vehicle for 48 h and then stimulated with TGFβ1 for 30 and 40 min, respectively. Cells were harvested and subjected to western blot analysis with phosphospecific antibodies of Smad2 and Smad3. The total protein levels of Smad2 and Smad3 were estimated by reprobing the membranes with total Smad2 and Smad3 antibody shown in the panels below. B : P0 LSCs pretreated with DAPT facilitates mesenchymal marker expression induced by TGFβ1. Representative blots (left panels) and densitometric analysis with standard deviation (SD; right columns) of three independent experiments are shown. *p
    Dimethyl Sulfoxide Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 20463 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore dmso
    Artificial diet assays with Myzus persicae to establish the specificity of aphid responses to NAM. Aphid feeding behavior, survival, and fecundity were observed at: ( a ) 6 hours, ( b ) 12 hours, ( c ) 24 hours, and ( d ) 48 hours in response to feeding on artificial diets without supplementation (Control) or supplemented with 10 mM NAM, 10 mM nicotinic acid (NA), 1.38 mM <t>pymetrozine</t> insecticide solubilized in 0.7% <t>DMSO,</t> or 0.7% DMSO alone. The average number of aphids in each behavioral response group were analyzed by one-way ANOVA. Means for each behavioral response that do not share a letter are significantly different at the 95% confidence level.
    Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 55308 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore dimethyl sulphoxide dmso
    Inhibitory effect of <t>17β-estradiol</t> ( E ), genistein ( G ) and daidzein ( D ) on post-injury neointima development in concentrations of 20/30/40 μM, compared to endothelium balloon denuded ( BD ) controls ( C ) (mean ± SD). The medium of all groups contained 1 % isopropanol ( 1 % iso ) and 1 % <t>DMSO</t> . Compared with controls 17β-estradiol, genistein, and daidzein (*) reduced neointima formation significantly (p = 0.05) in a concentration dependent manner. Data from Finking et al., 2000 [ 25 ].
    Dimethyl Sulphoxide Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1540 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore mtt
    Decreased survival of <t>CSA-13-treated</t> B. subtilis spores. Decline in the survival of B. subtilis culture after incubation with various concentrations of CSA-13 (panels A–C) evaluated using a killing assay method. Level of metabolic activity observed in ceragenin-treated spore samples exposed to appropriate environmental conditions when compared to untreated control (panels D–F) assessed using the <t>MTT</t> assay. Samples were incubated at RT (panels A, D for vegetative form and panels B, E for spore form, respectively) and at 70 °C (panels C, F) for 15 (white squares), 30 (grey circles), 60 (black circles), 120 (white triangles) and 240 (grey diamonds) minutes. Data from experiments performed in triplicate are shown.
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    Millipore 3 4 5 dimethylthiazol 2 yl 2 5 diphenyltetrazolium bromide
    Cytotoxic effect of cycloartobiloxanthone on H460, H292, A549 and H23 cancer cells. Each cell lines were treated with 0-100 μM of cycloartobiloxanthone for 24 h. Cell viability was analyzed by <t>3-[4,5-dimethylthiazol-2-yl]-2,5</t> <t>diphenyltetrazolium</t> bromide assay. Data was presented as mean±SD (n=3). The half maximal inhibitory concentration (IC50) of each cell line was calculated.
    3 4 5 Dimethylthiazol 2 Yl 2 5 Diphenyltetrazolium Bromide, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 13080 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore triton x 100
    Effects of BAK on Cx43 distribution. In untreated cornea, Cx43 exhibited a great uniformity in appearance with numerous big gap junction plaque. The distribution of Cx43 became disorganized and large gap junction plaques were rarely noticeable in 0.05% and 0.1% BAK-treated group (A). Western blot analysis for 1% Triton X-100 insoluble Cx43 (B) and quantitative analysis of insoluble Cx43 abundance were shown (C). Data are means ± SE from three independent experiments. **P value
    Triton X 100, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 65173 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore propidium iodide pi
    Cell cycle distribution in MCF12A ( a ), MCF7 ( b ), HaCaT ( c ) and A431 ( d ) cell lines after 24 h of incubation with resveratrol, 3,4,5-MTS and 2,4,5-MTS followed by <t>propidium</t> iodide labeling and flow cytometry analysis. Camptothecin at final concentration of 50 nM was used as a positive control. Results of three independent experiments are presented as mean ± SEM. * p
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    Millipore bovine serum albumin bsa
    Extracellularly applied oleic acid, an inducer of lipid droplet formation, rescues intracellular DENV-2 RNA abundance in PF-429242-treated Huh-7.5.1 cells. (A–D) Huh-7.5.1 cells were treated either with 0.01% <t>DMSO</t> (control) or 10 μM PF-429242 for 24 hours. The inhibitor was removed and the cells were then infected with DENV-2 (MOI = 0.01) (A–B) or mock-infected (C–D) with or without addition of 0.6 mM oleic acid (with <t>BSA</t> in molar ratio 6:1) for 24 hours. (A) Total RNA was harvested and DENV-2 RNA levels, normalized to β-actin transcript levels, were relatively quantified in cell extracts using qRT-PCR. (B) Collected supernatant was cultured with naïve Huh-7.5.1 cells for 48 hours, and DENV-2 RNA levels were quantified. (C) Representative images of the effect of PF-429242 and oleic acid on lipid droplets are shown. Fixed cells were permeabilized with Triton X-100 and stained for cell nuclei using Hoechst dye and for lipid droplets using Nile red. Images were obtained using a Leica SP8 confocal microscope with a 100x objective. (D) Abundance of LDs was quantified by measuring the average LD-positive area (μm 2 )/cell and the average number of LDs/cell based on Nile red fluorescence in 0.01% DMSO-treated (control), PF-429242-treated (10 μM), OA/BSA-treated, and PF-429242 (10 μM) + OA/BSA-treated cells using Fiji software ( > 50 cells analyzed). Statistical significance was calculated with a two-way (A, D) and one-way (B) ANOVA with a Bonferroni’s post-test (*, p
    Bovine Serum Albumin Bsa, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 32017 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore phosphate buffered saline pbs
    The effect of increasing concentrations of <t>TBBPA</t> (0.1, 1, 10, 50, and 100 μM) on the DCF fluorescence in cell-free <t>PBS</t> without Ca 2 + and Mg 2 + buffer and with addition of NAC after 30 min ( A ) and 60 min ( B ). Medium with 0.3 % hydrogen peroxide (H 2 O 2 ) was used as a positive control. The data is expressed as the means ± SEM of four independent experiments, each of which consisted of eight replicates per treatment group. * p
    Phosphate Buffered Saline Pbs, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 41313 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore fetal bovine serum fbs
    QODG inhibits VEGF–induced chemotactic motility of endothelial cells. QODG inhibited the migration of HUVECs. HUVECs were allowed to grow to full confluence in 6-well plates pre-coated with 0.1% gelatin and then starved with ECGM containing 0.5% <t>FBS</t> to inactivate cell proliferation. After that, cells were wounded with pipette and washed with <t>PBS,</t> then treated with or without VEGF (10 ng/mL) and DMSO (0.1%) or different concentrations of QODG (5, 20, 60, 180 µM) in ECGM containing 0.5% FBS. Images were taken using an inverted microscope (Olympus, Center Valley, PA, USA) (at 100×magnification) after 8 h of incubation, and migrated cells were quantified by manual counting. (A) Migration assay of HUVECs treated with only DMSO (0.1%). (B) Migration assay of HUVECs treated with VEGF (10 ng/mL) and DMSO (0.1%). (C–F) Migration assay of HUVECs treated with VEGF (10 ng/mL) and various concentrations of QODG (5, 20, 60, 180 µM). (G) Quantitative comparison of the numbers of migrated cells in different groups. Cells receiving only DMSO (0.1%) served as a vehicle control. Data are expressed as percentages of the vehicle control (100%) in mean ± SD from three independent experiments. **, P
    Fetal Bovine Serum Fbs, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 16849 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore mg132
    Assessment of senescence in U397 human leukemic cells treated with <t>MG132</t> (1 μM), Doxorubicin (DOX) 1 μM, or MG132 + DOX. U937 cells (1 × 10 6 ) were incubated alone or with different treatments for 24 h at 37°C in a humid atmosphere containing 5% CO2 and 95% air in RPMI-S medium. After incubation, the cells were washed in the same medium, and senescence was determined by measuring SA-β-gal utilizing flow cytometry. For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. Results are shown as % and represent the mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis, the Mann–Whitney U test. *p
    Mg132, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 20851 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore doxorubicin
    The impact of curcumin on stress induced premature senescence of VSMC A. Analysis of DNA damage in VSMC upon treatment with curcumin (0.1 or 1 μM) together with <t>doxorubicin</t> (10, 25 or 50 nM) expressed as a number of 53BP1 foci, after 3 and 7 days. 0 - cells without DNA damage, 1 - with only one focus, 2-5 - with the number of foci between 2 and 5, > 5 - cells with more than five foci. B. Western blot analysis of ATM, p53 total and phosphorylated, and p21 level in VSMC treated with doxorubicin (50 nM) and doxorubicin together with curcumin (0.1 or 1 μM). GAPDH served as a loading control. C. SA-β-gal activity in cells treated with curcumin (0.1 or 1 μM) or curcumin together with doxorubicin (10, 20 or 50 nM). The graph with the percentage of SA-β-gal-positive cells is shown. D. Western blot analysis of sirtuin 1, 3, 5 and 6 level and phosphorylation of sirtuin 1 after treatment with doxorubicin (50 nM) and doxorubicin together with curcumin (0.1 or 1 μM). GAPDH served as a loading control. All treated cells were in the phase of intensive growth (proliferating, young, passage 7-9). c - control; cur 0.1, cur 1 - 0.1 or 1 μM curcumin; dox 10, 25, 50 - 10, 25 or 50 nM doxorubicin. Error bars indicate SD, n = 3 or more.
    Doxorubicin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 8784 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore phalloidin
    Effect of wortmannin on SK channel activity in the presence or absence of 10 μM <t>phalloidin.</t> The experiments were performed in cell-attached patches.
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    Millipore nocodazole
    Time-lapse SHGM of the RNFL during continuous treatment with <t>nocodazole.</t> (a) Thickness of the RNFL at T = 0 min. (b) Axial cross section corresponding to the dashed line in (a). From top to bottom, at 0, 20, 40, and 60 min in treatment, respectively. The length of axis labels is 30 μ m. (c) Superposed margins of the retinal nerve fibers.
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    Millipore rapamycin
    Figure 10. <t>Rapamycin</t> (Rap) enhanced high glucose toxic effect on adult mouse cardiomyocytes, whereas 3-methyladenine (3-MA) attenuated it. Cardiomyocytes were isolated from adult C57B/6 mice, cultured under the indicated glucose conditions for
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    Millipore cisplatin
    <t>Cisplatin</t> concentration–dependent induction of p53, p53Ser15P, XPC, and DDB2. Cells were treated with the indicated concentrations of cisplatin (CP) at 37°C for 1 h. After treatment, cells were washed twice with PBS and incubated in drug-free
    Cisplatin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 10270 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore penicillin streptomycin
    <t>Cisplatin</t> concentration–dependent induction of p53, p53Ser15P, XPC, and DDB2. Cells were treated with the indicated concentrations of cisplatin (CP) at 37°C for 1 h. After treatment, cells were washed twice with PBS and incubated in drug-free
    Penicillin Streptomycin, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 36709 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore paclitaxel
    Downregulation of Mcl-1 is necessary for <t>paclitaxel-induced</t> apoptosis of bladder cancer cells. ( a , b ) 5637 and HT1197 cells were treated with dimethyl sulfoxide (DMSO) or 0.1 µM paclitaxel for 24 and 48 h. Western blot analyses of PARP, cleaved caspase-3, cleaved caspase-9, Mcl-1, Bcl-xL, Bax, Bak, cyclin B1, and p-histone H3 Ser10 are shown. β-actin was used as a loading control. ( c ) Cells were subjected to FISH with a centromeric probe specific for chromosome 17 (Spectrum green). DNA was stained with DAPI (blue). At least 100 cells were counted for each condition, and the percentage of cells with normal ploidy or higher ploidy is presented in the histogram. Representative images are shown. Photomicrographs were taken using a 40× objective. ( d ) The expression of Mcl-1 was evaluated in samples of 72 patients with muscle-invasive or non-muscle-invasive bladder carcinoma by immunohistochemistry and quantified in the histogram. The relationship between the levels of expression of Mcl-1 and tumor infiltration in patients with bladder carcinoma was statistically significant. p -values were obtained using Fisher’s exact test. NMIBC: non-muscle-invasive bladder cancer; MIBC: muscle-invasive bladder cancer. Bars represent 100 μm. ( e ) Kaplan–Meier analysis of recurrence-free survival in patients with low (blue) and high (red) Mcl-1 expression. Tick marks represent censored patients. p -value
    Paclitaxel, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 5901 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore hepes
    Outer membrane permeabilization assays of the engineered peptides. A 2 mL volume of E. coli UB1005 cells, diluted to 10 5 CFU/mL in 5 mM sodium <t>HEPES</t> buffer, was added to a quartz cuvette containing <t>NPN</t> to give a final concentration of 10 μM. Aliquots of peptides were added to the cuvette, and the fluorescence was recorded until there was no further increase in fluorescence. Data shown are three independent experiments.
    Hepes, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 50050 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore hydrogen peroxide
    Outer membrane permeabilization assays of the engineered peptides. A 2 mL volume of E. coli UB1005 cells, diluted to 10 5 CFU/mL in 5 mM sodium <t>HEPES</t> buffer, was added to a quartz cuvette containing <t>NPN</t> to give a final concentration of 10 μM. Aliquots of peptides were added to the cuvette, and the fluorescence was recorded until there was no further increase in fluorescence. Data shown are three independent experiments.
    Hydrogen Peroxide, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 16428 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore trypan blue
    Outer membrane permeabilization assays of the engineered peptides. A 2 mL volume of E. coli UB1005 cells, diluted to 10 5 CFU/mL in 5 mM sodium <t>HEPES</t> buffer, was added to a quartz cuvette containing <t>NPN</t> to give a final concentration of 10 μM. Aliquots of peptides were added to the cuvette, and the fluorescence was recorded until there was no further increase in fluorescence. Data shown are three independent experiments.
    Trypan Blue, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 13493 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore rpmi 1640 medium
    Effect of sophorolipid on C. albicans mature hyphae. C. albicans cells were grown in <t>RPMI-1640</t> containing 10% FBS for 5 hrs. After that mature hypha were treated with indicated concentrations of SL for time point zero (upper panel) and 5 hrs (lower panel) at 37 °C. At the end of incubation an aliquot was withdrawn from each sample and photographed at 60× magnification.
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    Millipore l glutamine
    Effect of sophorolipid on C. albicans mature hyphae. C. albicans cells were grown in <t>RPMI-1640</t> containing 10% FBS for 5 hrs. After that mature hypha were treated with indicated concentrations of SL for time point zero (upper panel) and 5 hrs (lower panel) at 37 °C. At the end of incubation an aliquot was withdrawn from each sample and photographed at 60× magnification.
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    Suppressing Smad7 protein expression with Notch inhibitor N-(N-[3,5-difluorophenacetyl]-l-alanyl)-S-phenylglycine t-butyl ester facilitates epithelial–mesenchymal transition and growth arrest induced by transforming growth factor-beta. A : Western blot analysis of Smad 2/3 phosphorylation. Left planes: Transforming growth factor-beta (TGFβ1) time-dependently induces Smad2/3 phosphorylation in P1 cells. Right planes: P1 cells were treated with 10 ng/ml TGFβ1 for 30 and 40 min as positive control. P0 and P1 cells were pretreated with 10 µM N-(N-[3,5-difluorophenacetyl]-l-alanyl)-S-phenylglycine t-butyl ester (DAPT) or dimethyl sulfoxide (DMSO) vehicle for 48 h and then stimulated with TGFβ1 for 30 and 40 min, respectively. Cells were harvested and subjected to western blot analysis with phosphospecific antibodies of Smad2 and Smad3. The total protein levels of Smad2 and Smad3 were estimated by reprobing the membranes with total Smad2 and Smad3 antibody shown in the panels below. B : P0 LSCs pretreated with DAPT facilitates mesenchymal marker expression induced by TGFβ1. Representative blots (left panels) and densitometric analysis with standard deviation (SD; right columns) of three independent experiments are shown. *p

    Journal: Molecular Vision

    Article Title: Notch prevents transforming growth factor-beta-assisted epithelial-mesenchymal transition in cultured limbal progenitor cells through the induction of Smad7

    doi:

    Figure Lengend Snippet: Suppressing Smad7 protein expression with Notch inhibitor N-(N-[3,5-difluorophenacetyl]-l-alanyl)-S-phenylglycine t-butyl ester facilitates epithelial–mesenchymal transition and growth arrest induced by transforming growth factor-beta. A : Western blot analysis of Smad 2/3 phosphorylation. Left planes: Transforming growth factor-beta (TGFβ1) time-dependently induces Smad2/3 phosphorylation in P1 cells. Right planes: P1 cells were treated with 10 ng/ml TGFβ1 for 30 and 40 min as positive control. P0 and P1 cells were pretreated with 10 µM N-(N-[3,5-difluorophenacetyl]-l-alanyl)-S-phenylglycine t-butyl ester (DAPT) or dimethyl sulfoxide (DMSO) vehicle for 48 h and then stimulated with TGFβ1 for 30 and 40 min, respectively. Cells were harvested and subjected to western blot analysis with phosphospecific antibodies of Smad2 and Smad3. The total protein levels of Smad2 and Smad3 were estimated by reprobing the membranes with total Smad2 and Smad3 antibody shown in the panels below. B : P0 LSCs pretreated with DAPT facilitates mesenchymal marker expression induced by TGFβ1. Representative blots (left panels) and densitometric analysis with standard deviation (SD; right columns) of three independent experiments are shown. *p

    Article Snippet: DAPT, IWR-1, cyclopamine, hydrocortisone, dimethyl sulfoxide (DMSO), insulin-transferrin-sodium selenite (ITSE) media supplement, mitomycin C (MMC), bovine serum albumin (BSA), 5-bromo-2'-deoxyuridine (BrdU), Triton X-100, Hoechst 33,258 dye, and formalin were all from Sigma-Aldrich (St. Louis, MO).

    Techniques: Expressing, Western Blot, Positive Control, Marker, Standard Deviation

    Artificial diet assays with Myzus persicae to establish the specificity of aphid responses to NAM. Aphid feeding behavior, survival, and fecundity were observed at: ( a ) 6 hours, ( b ) 12 hours, ( c ) 24 hours, and ( d ) 48 hours in response to feeding on artificial diets without supplementation (Control) or supplemented with 10 mM NAM, 10 mM nicotinic acid (NA), 1.38 mM pymetrozine insecticide solubilized in 0.7% DMSO, or 0.7% DMSO alone. The average number of aphids in each behavioral response group were analyzed by one-way ANOVA. Means for each behavioral response that do not share a letter are significantly different at the 95% confidence level.

    Journal: Scientific Reports

    Article Title: Nicotinamide Inhibits Aphid Fecundity and Impacts Survival

    doi: 10.1038/s41598-019-55931-z

    Figure Lengend Snippet: Artificial diet assays with Myzus persicae to establish the specificity of aphid responses to NAM. Aphid feeding behavior, survival, and fecundity were observed at: ( a ) 6 hours, ( b ) 12 hours, ( c ) 24 hours, and ( d ) 48 hours in response to feeding on artificial diets without supplementation (Control) or supplemented with 10 mM NAM, 10 mM nicotinic acid (NA), 1.38 mM pymetrozine insecticide solubilized in 0.7% DMSO, or 0.7% DMSO alone. The average number of aphids in each behavioral response group were analyzed by one-way ANOVA. Means for each behavioral response that do not share a letter are significantly different at the 95% confidence level.

    Article Snippet: To establish the specificity of the M. persicae response to NAM, diets were supplemented with 10 mM NAM, 10 mM nicotinic acid (NA, negative control), 1.38 µM pymetrozine solubilized in 0.7% DMSO (Millipore-Sigma, positive control), 0.7% DMSO (solvent control) or with diet alone (buffer control).

    Techniques:

    Inhibitory effect of 17β-estradiol ( E ), genistein ( G ) and daidzein ( D ) on post-injury neointima development in concentrations of 20/30/40 μM, compared to endothelium balloon denuded ( BD ) controls ( C ) (mean ± SD). The medium of all groups contained 1 % isopropanol ( 1 % iso ) and 1 % DMSO . Compared with controls 17β-estradiol, genistein, and daidzein (*) reduced neointima formation significantly (p = 0.05) in a concentration dependent manner. Data from Finking et al., 2000 [ 25 ].

    Journal: BMC Cardiovascular Disorders

    Article Title: Reduction of post injury neointima formation due to 17?-estradiol and phytoestrogen treatment is not influenced by the pure synthetic estrogen receptor antagonist ICI 182,780 in vitro

    doi:

    Figure Lengend Snippet: Inhibitory effect of 17β-estradiol ( E ), genistein ( G ) and daidzein ( D ) on post-injury neointima development in concentrations of 20/30/40 μM, compared to endothelium balloon denuded ( BD ) controls ( C ) (mean ± SD). The medium of all groups contained 1 % isopropanol ( 1 % iso ) and 1 % DMSO . Compared with controls 17β-estradiol, genistein, and daidzein (*) reduced neointima formation significantly (p = 0.05) in a concentration dependent manner. Data from Finking et al., 2000 [ 25 ].

    Article Snippet: Control groups were held in medium containing 1 % isopropanol (Roth, Karlsruhe, Germany) and 1% dimethyl sulphoxide (DMSO) (Sigma, Deisenhofen, Germany) because 17β-estradiol, Genistein and Daidzein were dissolved in isopropanol and DMSO of the same concentration.

    Techniques: Concentration Assay

    Decreased survival of CSA-13-treated B. subtilis spores. Decline in the survival of B. subtilis culture after incubation with various concentrations of CSA-13 (panels A–C) evaluated using a killing assay method. Level of metabolic activity observed in ceragenin-treated spore samples exposed to appropriate environmental conditions when compared to untreated control (panels D–F) assessed using the MTT assay. Samples were incubated at RT (panels A, D for vegetative form and panels B, E for spore form, respectively) and at 70 °C (panels C, F) for 15 (white squares), 30 (grey circles), 60 (black circles), 120 (white triangles) and 240 (grey diamonds) minutes. Data from experiments performed in triplicate are shown.

    Journal: Scientific Reports

    Article Title: Sporicidal activity of ceragenin CSA-13 against Bacillus subtilis

    doi: 10.1038/srep44452

    Figure Lengend Snippet: Decreased survival of CSA-13-treated B. subtilis spores. Decline in the survival of B. subtilis culture after incubation with various concentrations of CSA-13 (panels A–C) evaluated using a killing assay method. Level of metabolic activity observed in ceragenin-treated spore samples exposed to appropriate environmental conditions when compared to untreated control (panels D–F) assessed using the MTT assay. Samples were incubated at RT (panels A, D for vegetative form and panels B, E for spore form, respectively) and at 70 °C (panels C, F) for 15 (white squares), 30 (grey circles), 60 (black circles), 120 (white triangles) and 240 (grey diamonds) minutes. Data from experiments performed in triplicate are shown.

    Article Snippet: Briefly, after incubation of spores with various concentrations of CSA-13 in a non-growing medium (PBS), 20 μL of MTT solution (thiazolyl blue tetrazolium bromide, Sigma-Aldrich, St Louis, MO, USA, 5 mg/mL) and 100 μL of LB medium broth were added.

    Techniques: Incubation, Activity Assay, MTT Assay

    Cytotoxic effect of cycloartobiloxanthone on H460, H292, A549 and H23 cancer cells. Each cell lines were treated with 0-100 μM of cycloartobiloxanthone for 24 h. Cell viability was analyzed by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide assay. Data was presented as mean±SD (n=3). The half maximal inhibitory concentration (IC50) of each cell line was calculated.

    Journal: In Vivo

    Article Title: Cycloartobiloxanthone Induces Human Lung Cancer Cell Apoptosis via Mitochondria-dependent Apoptotic Pathway

    doi: 10.21873/invivo.11206

    Figure Lengend Snippet: Cytotoxic effect of cycloartobiloxanthone on H460, H292, A549 and H23 cancer cells. Each cell lines were treated with 0-100 μM of cycloartobiloxanthone for 24 h. Cell viability was analyzed by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide assay. Data was presented as mean±SD (n=3). The half maximal inhibitory concentration (IC50) of each cell line was calculated.

    Article Snippet: 3-(4,5-Dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT), dimethyl sulfoxide (DMSO), Hoechst 33342, propidium iodide (PI), and bovine serum albumin (BSA) were purchased from Sigma Chemical, Inc. (St. Louis, MO, USA).

    Techniques: Concentration Assay

    Effects of BAK on Cx43 distribution. In untreated cornea, Cx43 exhibited a great uniformity in appearance with numerous big gap junction plaque. The distribution of Cx43 became disorganized and large gap junction plaques were rarely noticeable in 0.05% and 0.1% BAK-treated group (A). Western blot analysis for 1% Triton X-100 insoluble Cx43 (B) and quantitative analysis of insoluble Cx43 abundance were shown (C). Data are means ± SE from three independent experiments. **P value

    Journal: PLoS ONE

    Article Title: Benzalkonium Chloride Suppresses Rabbit Corneal Endothelium Intercellular Gap Junction Communication

    doi: 10.1371/journal.pone.0109708

    Figure Lengend Snippet: Effects of BAK on Cx43 distribution. In untreated cornea, Cx43 exhibited a great uniformity in appearance with numerous big gap junction plaque. The distribution of Cx43 became disorganized and large gap junction plaques were rarely noticeable in 0.05% and 0.1% BAK-treated group (A). Western blot analysis for 1% Triton X-100 insoluble Cx43 (B) and quantitative analysis of insoluble Cx43 abundance were shown (C). Data are means ± SE from three independent experiments. **P value

    Article Snippet: Reagents and Antibodies BAK, dimethyl sulfoxide (DMSO), and Triton X-100, collagenase I and Lucifer Yellow dye were purchased from Sigma Aldrich (St. Louis, MO); Protein A/G PLUS-Agarose Immunoprecipitation Reagent was from Santa Cruz Biotechnology (Santa Cruz, CA); PVDF Western Blotting Membrane was from Roche (Basel, Switzerland); pentobarbital sodium was from Abbott Laboratories (North Chicago, IL); Enhanced chemiluminescence (ECL) kit was obtained from GE Healthcare UK (Chalfont, UK); Mounting medium with 4, 6-diamidino-2-phenylindole (DAPI) and bovine serum albumin (BSA) were from Vector Laboratories (Burlingame CA); Dulbecco's modified Eagle's medium (DMEM) and fetal bovine serum (FBS) were purchased from Life Technologies (Carlsbad, CA); mouse-anti-rabbit ZO-1 antibody, Alexa488-conjugated donkey-anti-mouse IgG, and Alexa555-conjugated donkey-anti-goat IgG were from Life Technologies (Carlsbad, CA); goat polyclonal antibody for Cx43 and P-Cx43, Horseradish peroxidase (HRP)-conjugated donkey anti- goat IgG were from Santa Cruz Biotechnology (Santa Cruz, CA); mouse anti-rabbit β-actin antibody from Sigma Aldrich (St. Louis, MO); Horseradish peroxidase (HRP)-conjugated goat anti- mouse IgG from Merck (Darmstadt, Germany).

    Techniques: Western Blot

    Cell cycle distribution in MCF12A ( a ), MCF7 ( b ), HaCaT ( c ) and A431 ( d ) cell lines after 24 h of incubation with resveratrol, 3,4,5-MTS and 2,4,5-MTS followed by propidium iodide labeling and flow cytometry analysis. Camptothecin at final concentration of 50 nM was used as a positive control. Results of three independent experiments are presented as mean ± SEM. * p

    Journal: Cytotechnology

    Article Title: Cytotoxic, tubulin-interfering and proapoptotic activities of 4′-methylthio-trans-stilbene derivatives, analogues of trans-resveratrol

    doi: 10.1007/s10616-018-0227-3

    Figure Lengend Snippet: Cell cycle distribution in MCF12A ( a ), MCF7 ( b ), HaCaT ( c ) and A431 ( d ) cell lines after 24 h of incubation with resveratrol, 3,4,5-MTS and 2,4,5-MTS followed by propidium iodide labeling and flow cytometry analysis. Camptothecin at final concentration of 50 nM was used as a positive control. Results of three independent experiments are presented as mean ± SEM. * p

    Article Snippet: Chemicals Resveratrol (purity 99%), antibiotic solution (10,000 units’ penicillin, 10 mg streptomycin and 25 μg amphotericin B per mL), camptothecin, dimethyl sulfoxide (DMSO), fetal bovine serum (FBS), Dulbecco’s Modified Eagle’s Medium (DMEM), Ham’s F12 mixture, propidium iodide (PI), ribonuclease A (RNase A) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and all other compounds were provided by Sigma-Aldrich Co. (St Louis, MO, USA).

    Techniques: Incubation, Labeling, Flow Cytometry, Cytometry, Concentration Assay, Positive Control

    Effect of resveratrol, 3,4,5-MTS and 2,4,5-MTS on phosphatidylserine externalization and propidium iodide staining in MCF12A ( a ), MCF7 ( b ), HaCaT ( c ) and A431 ( d ) cell lines after 24 h of incubation subjected to Annexin-V binding and propidium iodide labeling followed by flow cytometry analysis. The cell populations of AV−/PI−, AV+/PI+ and AV+/PI− representing viable, dead and apoptotic cells, respectively, were estimated. Camptothecin at final concentration of 50 nM was used as a positive control. Results of three independent experiments are presented as mean ± SEM. * p

    Journal: Cytotechnology

    Article Title: Cytotoxic, tubulin-interfering and proapoptotic activities of 4′-methylthio-trans-stilbene derivatives, analogues of trans-resveratrol

    doi: 10.1007/s10616-018-0227-3

    Figure Lengend Snippet: Effect of resveratrol, 3,4,5-MTS and 2,4,5-MTS on phosphatidylserine externalization and propidium iodide staining in MCF12A ( a ), MCF7 ( b ), HaCaT ( c ) and A431 ( d ) cell lines after 24 h of incubation subjected to Annexin-V binding and propidium iodide labeling followed by flow cytometry analysis. The cell populations of AV−/PI−, AV+/PI+ and AV+/PI− representing viable, dead and apoptotic cells, respectively, were estimated. Camptothecin at final concentration of 50 nM was used as a positive control. Results of three independent experiments are presented as mean ± SEM. * p

    Article Snippet: Chemicals Resveratrol (purity 99%), antibiotic solution (10,000 units’ penicillin, 10 mg streptomycin and 25 μg amphotericin B per mL), camptothecin, dimethyl sulfoxide (DMSO), fetal bovine serum (FBS), Dulbecco’s Modified Eagle’s Medium (DMEM), Ham’s F12 mixture, propidium iodide (PI), ribonuclease A (RNase A) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and all other compounds were provided by Sigma-Aldrich Co. (St Louis, MO, USA).

    Techniques: Staining, Incubation, Binding Assay, Labeling, Flow Cytometry, Cytometry, Concentration Assay, Positive Control

    Extracellularly applied oleic acid, an inducer of lipid droplet formation, rescues intracellular DENV-2 RNA abundance in PF-429242-treated Huh-7.5.1 cells. (A–D) Huh-7.5.1 cells were treated either with 0.01% DMSO (control) or 10 μM PF-429242 for 24 hours. The inhibitor was removed and the cells were then infected with DENV-2 (MOI = 0.01) (A–B) or mock-infected (C–D) with or without addition of 0.6 mM oleic acid (with BSA in molar ratio 6:1) for 24 hours. (A) Total RNA was harvested and DENV-2 RNA levels, normalized to β-actin transcript levels, were relatively quantified in cell extracts using qRT-PCR. (B) Collected supernatant was cultured with naïve Huh-7.5.1 cells for 48 hours, and DENV-2 RNA levels were quantified. (C) Representative images of the effect of PF-429242 and oleic acid on lipid droplets are shown. Fixed cells were permeabilized with Triton X-100 and stained for cell nuclei using Hoechst dye and for lipid droplets using Nile red. Images were obtained using a Leica SP8 confocal microscope with a 100x objective. (D) Abundance of LDs was quantified by measuring the average LD-positive area (μm 2 )/cell and the average number of LDs/cell based on Nile red fluorescence in 0.01% DMSO-treated (control), PF-429242-treated (10 μM), OA/BSA-treated, and PF-429242 (10 μM) + OA/BSA-treated cells using Fiji software ( > 50 cells analyzed). Statistical significance was calculated with a two-way (A, D) and one-way (B) ANOVA with a Bonferroni’s post-test (*, p

    Journal: PLoS ONE

    Article Title: Human Subtilisin Kexin Isozyme-1 (SKI-1)/Site-1 Protease (S1P) regulates cytoplasmic lipid droplet abundance: A potential target for indirect-acting anti-dengue virus agents

    doi: 10.1371/journal.pone.0174483

    Figure Lengend Snippet: Extracellularly applied oleic acid, an inducer of lipid droplet formation, rescues intracellular DENV-2 RNA abundance in PF-429242-treated Huh-7.5.1 cells. (A–D) Huh-7.5.1 cells were treated either with 0.01% DMSO (control) or 10 μM PF-429242 for 24 hours. The inhibitor was removed and the cells were then infected with DENV-2 (MOI = 0.01) (A–B) or mock-infected (C–D) with or without addition of 0.6 mM oleic acid (with BSA in molar ratio 6:1) for 24 hours. (A) Total RNA was harvested and DENV-2 RNA levels, normalized to β-actin transcript levels, were relatively quantified in cell extracts using qRT-PCR. (B) Collected supernatant was cultured with naïve Huh-7.5.1 cells for 48 hours, and DENV-2 RNA levels were quantified. (C) Representative images of the effect of PF-429242 and oleic acid on lipid droplets are shown. Fixed cells were permeabilized with Triton X-100 and stained for cell nuclei using Hoechst dye and for lipid droplets using Nile red. Images were obtained using a Leica SP8 confocal microscope with a 100x objective. (D) Abundance of LDs was quantified by measuring the average LD-positive area (μm 2 )/cell and the average number of LDs/cell based on Nile red fluorescence in 0.01% DMSO-treated (control), PF-429242-treated (10 μM), OA/BSA-treated, and PF-429242 (10 μM) + OA/BSA-treated cells using Fiji software ( > 50 cells analyzed). Statistical significance was calculated with a two-way (A, D) and one-way (B) ANOVA with a Bonferroni’s post-test (*, p

    Article Snippet: Bovine serum albumin (BSA), oleic acid (OA), and dimethyl sulfoxide (DMSO) were obtained from Sigma-Aldrich Corp.

    Techniques: Infection, Quantitative RT-PCR, Cell Culture, Staining, Microscopy, Fluorescence, Software

    The effect of increasing concentrations of TBBPA (0.1, 1, 10, 50, and 100 μM) on the DCF fluorescence in cell-free PBS without Ca 2 + and Mg 2 + buffer and with addition of NAC after 30 min ( A ) and 60 min ( B ). Medium with 0.3 % hydrogen peroxide (H 2 O 2 ) was used as a positive control. The data is expressed as the means ± SEM of four independent experiments, each of which consisted of eight replicates per treatment group. * p

    Journal: Environmental Science and Pollution Research International

    Article Title: Tetrabromobisphenol A (TBBPA)-stimulated reactive oxygen species (ROS) production in cell-free model using the 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA) assay—limitations of method

    doi: 10.1007/s11356-016-6450-6

    Figure Lengend Snippet: The effect of increasing concentrations of TBBPA (0.1, 1, 10, 50, and 100 μM) on the DCF fluorescence in cell-free PBS without Ca 2 + and Mg 2 + buffer and with addition of NAC after 30 min ( A ) and 60 min ( B ). Medium with 0.3 % hydrogen peroxide (H 2 O 2 ) was used as a positive control. The data is expressed as the means ± SEM of four independent experiments, each of which consisted of eight replicates per treatment group. * p

    Article Snippet: Reagents TBBPA, H2 DCFDA, phosphate-buffered saline (PBS) without Ca2+ and Mg2+ , 2,2-diphenyl-1-picrylhydrazyl (DPPH·), fetal bovine serum (FBS), hydrogen peroxide (H2 O2 ), ethanol, and dimethyl sulfoxide (DMSO) were purchased from Sigma-Aldrich (St. Louis, MO, USA).

    Techniques: Fluorescence, Positive Control

    QODG inhibits VEGF–induced chemotactic motility of endothelial cells. QODG inhibited the migration of HUVECs. HUVECs were allowed to grow to full confluence in 6-well plates pre-coated with 0.1% gelatin and then starved with ECGM containing 0.5% FBS to inactivate cell proliferation. After that, cells were wounded with pipette and washed with PBS, then treated with or without VEGF (10 ng/mL) and DMSO (0.1%) or different concentrations of QODG (5, 20, 60, 180 µM) in ECGM containing 0.5% FBS. Images were taken using an inverted microscope (Olympus, Center Valley, PA, USA) (at 100×magnification) after 8 h of incubation, and migrated cells were quantified by manual counting. (A) Migration assay of HUVECs treated with only DMSO (0.1%). (B) Migration assay of HUVECs treated with VEGF (10 ng/mL) and DMSO (0.1%). (C–F) Migration assay of HUVECs treated with VEGF (10 ng/mL) and various concentrations of QODG (5, 20, 60, 180 µM). (G) Quantitative comparison of the numbers of migrated cells in different groups. Cells receiving only DMSO (0.1%) served as a vehicle control. Data are expressed as percentages of the vehicle control (100%) in mean ± SD from three independent experiments. **, P

    Journal: PLoS ONE

    Article Title: Quercetin-4?-O-?-D-glucopyranoside (QODG) Inhibits Angiogenesis by Suppressing VEGFR2-Mediated Signaling in Zebrafish and Endothelial Cells

    doi: 10.1371/journal.pone.0031708

    Figure Lengend Snippet: QODG inhibits VEGF–induced chemotactic motility of endothelial cells. QODG inhibited the migration of HUVECs. HUVECs were allowed to grow to full confluence in 6-well plates pre-coated with 0.1% gelatin and then starved with ECGM containing 0.5% FBS to inactivate cell proliferation. After that, cells were wounded with pipette and washed with PBS, then treated with or without VEGF (10 ng/mL) and DMSO (0.1%) or different concentrations of QODG (5, 20, 60, 180 µM) in ECGM containing 0.5% FBS. Images were taken using an inverted microscope (Olympus, Center Valley, PA, USA) (at 100×magnification) after 8 h of incubation, and migrated cells were quantified by manual counting. (A) Migration assay of HUVECs treated with only DMSO (0.1%). (B) Migration assay of HUVECs treated with VEGF (10 ng/mL) and DMSO (0.1%). (C–F) Migration assay of HUVECs treated with VEGF (10 ng/mL) and various concentrations of QODG (5, 20, 60, 180 µM). (G) Quantitative comparison of the numbers of migrated cells in different groups. Cells receiving only DMSO (0.1%) served as a vehicle control. Data are expressed as percentages of the vehicle control (100%) in mean ± SD from three independent experiments. **, P

    Article Snippet: Phosphate-buffered saline (PBS), Tween 20, fetal bovine serum (FBS), bovine serum albumin (BSA), phenylmethanesulfonyl fluoride (PMSF), ethylenediaminetetraacetic acid (EDTA), heparin, HEPES buffer, penicillin, streptomycin, NaHCO3 , amphotericin B, dimethyl sulfoxide (DMSO), gelatin and MS-222 (tricaine methanesulfonate) were obtained from Sigma (St. Louis, MO, USA).

    Techniques: Migration, Transferring, Inverted Microscopy, Incubation

    Assessment of senescence in U397 human leukemic cells treated with MG132 (1 μM), Doxorubicin (DOX) 1 μM, or MG132 + DOX. U937 cells (1 × 10 6 ) were incubated alone or with different treatments for 24 h at 37°C in a humid atmosphere containing 5% CO2 and 95% air in RPMI-S medium. After incubation, the cells were washed in the same medium, and senescence was determined by measuring SA-β-gal utilizing flow cytometry. For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. Results are shown as % and represent the mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis, the Mann–Whitney U test. *p

    Journal: Cancer Cell International

    Article Title: Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss

    doi: 10.1186/1475-2867-14-13

    Figure Lengend Snippet: Assessment of senescence in U397 human leukemic cells treated with MG132 (1 μM), Doxorubicin (DOX) 1 μM, or MG132 + DOX. U937 cells (1 × 10 6 ) were incubated alone or with different treatments for 24 h at 37°C in a humid atmosphere containing 5% CO2 and 95% air in RPMI-S medium. After incubation, the cells were washed in the same medium, and senescence was determined by measuring SA-β-gal utilizing flow cytometry. For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. Results are shown as % and represent the mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis, the Mann–Whitney U test. *p

    Article Snippet: With regard to the proteasome inhibitor, we utilized MG132 (N-CBZ-LEU-LEU-AL) (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Incubation, Flow Cytometry, Cytometry, Software, Standard Deviation, MANN-WHITNEY

    Expression of pro- and antiapoptotic genes in U937 human leukemic cells treated with Doxorubicin (DOX), MG132, and MG132 + DOX. U937 human leukemia cells were incubated, alone or in combination, with DOX 1 μM, MG132 (1 μM), or MG132 + DOX for 3 h. Gene expression was assessed by quantitative real-time PCR. Data are expressed as relative mRNA fold-change. RPL32 were used for normalization, and the value of untreated cells, as calibrator. Variations were considered significant at ≥30% compared with the constitutive gene. In all cases, the Standard deviation (SD) was not > 0.08.

    Journal: Cancer Cell International

    Article Title: Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss

    doi: 10.1186/1475-2867-14-13

    Figure Lengend Snippet: Expression of pro- and antiapoptotic genes in U937 human leukemic cells treated with Doxorubicin (DOX), MG132, and MG132 + DOX. U937 human leukemia cells were incubated, alone or in combination, with DOX 1 μM, MG132 (1 μM), or MG132 + DOX for 3 h. Gene expression was assessed by quantitative real-time PCR. Data are expressed as relative mRNA fold-change. RPL32 were used for normalization, and the value of untreated cells, as calibrator. Variations were considered significant at ≥30% compared with the constitutive gene. In all cases, the Standard deviation (SD) was not > 0.08.

    Article Snippet: With regard to the proteasome inhibitor, we utilized MG132 (N-CBZ-LEU-LEU-AL) (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Expressing, Incubation, Real-time Polymerase Chain Reaction, Standard Deviation

    Apoptosis and caspase activity in U937 cells treated with Doxorubicin (DOX), MG132, or MG132 + DOX. U937 cells were incubated alone or in combination with MG132 (1 μM), DOX 1 μM, or MG132 + DOX for 24 h at 37°C in a humid atmosphere containing 5% CO 2 in RPMI-S culture medium. After incubation, the cells were washed and apoptosis was assessed by flow cytometry using Annexin V-fluorescein isothiocyanate (FITC) (a) . For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. The activity of caspase-3 (b) , -8 (c) , and −9 (d) was evaluated utilizing a caspase colorimetric staining kit. The results represent the mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis was performed by means of the Mann–Whitney U test. ●p

    Journal: Cancer Cell International

    Article Title: Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss

    doi: 10.1186/1475-2867-14-13

    Figure Lengend Snippet: Apoptosis and caspase activity in U937 cells treated with Doxorubicin (DOX), MG132, or MG132 + DOX. U937 cells were incubated alone or in combination with MG132 (1 μM), DOX 1 μM, or MG132 + DOX for 24 h at 37°C in a humid atmosphere containing 5% CO 2 in RPMI-S culture medium. After incubation, the cells were washed and apoptosis was assessed by flow cytometry using Annexin V-fluorescein isothiocyanate (FITC) (a) . For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. The activity of caspase-3 (b) , -8 (c) , and −9 (d) was evaluated utilizing a caspase colorimetric staining kit. The results represent the mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis was performed by means of the Mann–Whitney U test. ●p

    Article Snippet: With regard to the proteasome inhibitor, we utilized MG132 (N-CBZ-LEU-LEU-AL) (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Activity Assay, Incubation, Flow Cytometry, Cytometry, Software, Staining, Standard Deviation, MANN-WHITNEY

    Western blot analysis of caspases −3, -8, and −9 and cytochrome c in U937 cells treated with MG132, Doxorubicin (DOX) or MG132 + DOX. U937 cells were cultured and treated with MG132 (1 μM), DOX 1 μM, or with MG132 + DOX. After 24 h, the cells were harvested and lysed. Equivalent amounts of individual lysates were placed onto 10% SDS gradient polyacrylamide gels for electrophoresis and then were electrotransferred onto Immobilon-P PVDF membranes. A representative study is shown. Relative density was calculated using IMAGEJ ver.1.46r software.

    Journal: Cancer Cell International

    Article Title: Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss

    doi: 10.1186/1475-2867-14-13

    Figure Lengend Snippet: Western blot analysis of caspases −3, -8, and −9 and cytochrome c in U937 cells treated with MG132, Doxorubicin (DOX) or MG132 + DOX. U937 cells were cultured and treated with MG132 (1 μM), DOX 1 μM, or with MG132 + DOX. After 24 h, the cells were harvested and lysed. Equivalent amounts of individual lysates were placed onto 10% SDS gradient polyacrylamide gels for electrophoresis and then were electrotransferred onto Immobilon-P PVDF membranes. A representative study is shown. Relative density was calculated using IMAGEJ ver.1.46r software.

    Article Snippet: With regard to the proteasome inhibitor, we utilized MG132 (N-CBZ-LEU-LEU-AL) (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Western Blot, Cell Culture, Electrophoresis, Software

    MG132 + DOX induces potential mitochondrial membrane loss. U937 cells were cultured and treated with MG132 (1 μM), DOX 1 μM, or MG132 + DOX. After 24 h, the cells were harvested and the ΔΨm was assessed by flow cytometry using DIOC 6 staining. Protonophore Cyanide m-chorophenylhydrazone (CCCP) was used as a positive control. Representative histograms of each treatment are displayed. For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. The graph shows the results as mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis, the Mann–Whitney U test. *p

    Journal: Cancer Cell International

    Article Title: Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss

    doi: 10.1186/1475-2867-14-13

    Figure Lengend Snippet: MG132 + DOX induces potential mitochondrial membrane loss. U937 cells were cultured and treated with MG132 (1 μM), DOX 1 μM, or MG132 + DOX. After 24 h, the cells were harvested and the ΔΨm was assessed by flow cytometry using DIOC 6 staining. Protonophore Cyanide m-chorophenylhydrazone (CCCP) was used as a positive control. Representative histograms of each treatment are displayed. For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. The graph shows the results as mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis, the Mann–Whitney U test. *p

    Article Snippet: With regard to the proteasome inhibitor, we utilized MG132 (N-CBZ-LEU-LEU-AL) (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Cell Culture, Flow Cytometry, Cytometry, Staining, Positive Control, Software, Standard Deviation, MANN-WHITNEY

    Determination of phosphorylated p65 (NF-кB subunit), Bcl-2 and bcl-XL in U937 human leukemic cells treated in vitro with Doxorubicin (DOX), the MG132 proteasome inhibitor, and MG132 + DOX. U937 cells were incubated, alone or in combination, with DOX 1 μM, MG132 (1 μM), or MG132 + DOX. After 1 h, the cells were harvested and the phosphorylated p65 protein was determined by flow cytometry (a) . U937 were treated with MG132, DOX or it´s combination after that the cells were harvested and the Bcl-2 (b) and Bcl-XL (c) antiapoptotic proteins were determined by flow cytometry. An appropriate isotype control was utilized to adjust background fluorescence. The graphs show the Mean fluorescence intensity (MFI) of p65, Bcl-2 or Bcl-XL. For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. Results represent the mean ± the Standard deviation (SD) of three independent experiments carried out in triplicate. ♦ p

    Journal: Cancer Cell International

    Article Title: Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss

    doi: 10.1186/1475-2867-14-13

    Figure Lengend Snippet: Determination of phosphorylated p65 (NF-кB subunit), Bcl-2 and bcl-XL in U937 human leukemic cells treated in vitro with Doxorubicin (DOX), the MG132 proteasome inhibitor, and MG132 + DOX. U937 cells were incubated, alone or in combination, with DOX 1 μM, MG132 (1 μM), or MG132 + DOX. After 1 h, the cells were harvested and the phosphorylated p65 protein was determined by flow cytometry (a) . U937 were treated with MG132, DOX or it´s combination after that the cells were harvested and the Bcl-2 (b) and Bcl-XL (c) antiapoptotic proteins were determined by flow cytometry. An appropriate isotype control was utilized to adjust background fluorescence. The graphs show the Mean fluorescence intensity (MFI) of p65, Bcl-2 or Bcl-XL. For each sample, at least 20,000 events were acquired in a FACSAria-I cell sorter and the data were analyzed with FACSDiva software. Results represent the mean ± the Standard deviation (SD) of three independent experiments carried out in triplicate. ♦ p

    Article Snippet: With regard to the proteasome inhibitor, we utilized MG132 (N-CBZ-LEU-LEU-AL) (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: In Vitro, Incubation, Flow Cytometry, Cytometry, Fluorescence, Software, Standard Deviation

    MG132 + DOX induces a decrease in viability and proliferation in U937 cells. U937 cells were treated with MG132 proteasome inhibitor (1 μM), Doxorubicin (DOX) 1 μM, and MG132 + DOX. U937 cells (2 × 10 4 ) were incubated in the presence of different treatments for 18, 24, 36, and 48 h at 37°C in a humid atmosphere containing 5% CO 2 and 95% air in RPMI-S culture medium. Subsequently, WST-1 was added and the cells were incubated for 3 h; then, viability was assessed by measuring the Optical density (OD) at 490 nm. The OD value of the Untreated control group (UCG) was taken as 100% cell viability (a) . After 24 h, morphological changes were observed, U937 cells were stained with blue trypan or fixed and stained with Wright on cover-glass slides and observed under a light microscope with zoom lens of 4X to 40X using a Leica DMLB microscope (b) . After 72 h, proliferation was assessed using BrdU (c) . The results represent the mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis, the Mann–Whitney U test. *p

    Journal: Cancer Cell International

    Article Title: Sensitization of U937 leukemia cells to doxorubicin by the MG132 proteasome inhibitor induces an increase in apoptosis by suppressing NF-kappa B and mitochondrial membrane potential loss

    doi: 10.1186/1475-2867-14-13

    Figure Lengend Snippet: MG132 + DOX induces a decrease in viability and proliferation in U937 cells. U937 cells were treated with MG132 proteasome inhibitor (1 μM), Doxorubicin (DOX) 1 μM, and MG132 + DOX. U937 cells (2 × 10 4 ) were incubated in the presence of different treatments for 18, 24, 36, and 48 h at 37°C in a humid atmosphere containing 5% CO 2 and 95% air in RPMI-S culture medium. Subsequently, WST-1 was added and the cells were incubated for 3 h; then, viability was assessed by measuring the Optical density (OD) at 490 nm. The OD value of the Untreated control group (UCG) was taken as 100% cell viability (a) . After 24 h, morphological changes were observed, U937 cells were stained with blue trypan or fixed and stained with Wright on cover-glass slides and observed under a light microscope with zoom lens of 4X to 40X using a Leica DMLB microscope (b) . After 72 h, proliferation was assessed using BrdU (c) . The results represent the mean ± the Standard deviation (SD) of three independent experiments performed in triplicate. Statistical analysis, the Mann–Whitney U test. *p

    Article Snippet: With regard to the proteasome inhibitor, we utilized MG132 (N-CBZ-LEU-LEU-AL) (Sigma-Aldrich, St. Louis, MO, USA).

    Techniques: Incubation, Staining, Light Microscopy, Microscopy, Standard Deviation, MANN-WHITNEY

    The impact of curcumin on stress induced premature senescence of VSMC A. Analysis of DNA damage in VSMC upon treatment with curcumin (0.1 or 1 μM) together with doxorubicin (10, 25 or 50 nM) expressed as a number of 53BP1 foci, after 3 and 7 days. 0 - cells without DNA damage, 1 - with only one focus, 2-5 - with the number of foci between 2 and 5, > 5 - cells with more than five foci. B. Western blot analysis of ATM, p53 total and phosphorylated, and p21 level in VSMC treated with doxorubicin (50 nM) and doxorubicin together with curcumin (0.1 or 1 μM). GAPDH served as a loading control. C. SA-β-gal activity in cells treated with curcumin (0.1 or 1 μM) or curcumin together with doxorubicin (10, 20 or 50 nM). The graph with the percentage of SA-β-gal-positive cells is shown. D. Western blot analysis of sirtuin 1, 3, 5 and 6 level and phosphorylation of sirtuin 1 after treatment with doxorubicin (50 nM) and doxorubicin together with curcumin (0.1 or 1 μM). GAPDH served as a loading control. All treated cells were in the phase of intensive growth (proliferating, young, passage 7-9). c - control; cur 0.1, cur 1 - 0.1 or 1 μM curcumin; dox 10, 25, 50 - 10, 25 or 50 nM doxorubicin. Error bars indicate SD, n = 3 or more.

    Journal: Oncotarget

    Article Title: Curcumin elevates sirtuin level but does not postpone in vitro senescence of human cells building the vasculature

    doi: 10.18632/oncotarget.8450

    Figure Lengend Snippet: The impact of curcumin on stress induced premature senescence of VSMC A. Analysis of DNA damage in VSMC upon treatment with curcumin (0.1 or 1 μM) together with doxorubicin (10, 25 or 50 nM) expressed as a number of 53BP1 foci, after 3 and 7 days. 0 - cells without DNA damage, 1 - with only one focus, 2-5 - with the number of foci between 2 and 5, > 5 - cells with more than five foci. B. Western blot analysis of ATM, p53 total and phosphorylated, and p21 level in VSMC treated with doxorubicin (50 nM) and doxorubicin together with curcumin (0.1 or 1 μM). GAPDH served as a loading control. C. SA-β-gal activity in cells treated with curcumin (0.1 or 1 μM) or curcumin together with doxorubicin (10, 20 or 50 nM). The graph with the percentage of SA-β-gal-positive cells is shown. D. Western blot analysis of sirtuin 1, 3, 5 and 6 level and phosphorylation of sirtuin 1 after treatment with doxorubicin (50 nM) and doxorubicin together with curcumin (0.1 or 1 μM). GAPDH served as a loading control. All treated cells were in the phase of intensive growth (proliferating, young, passage 7-9). c - control; cur 0.1, cur 1 - 0.1 or 1 μM curcumin; dox 10, 25, 50 - 10, 25 or 50 nM doxorubicin. Error bars indicate SD, n = 3 or more.

    Article Snippet: Reagents Curcumin (C1386) was from Cayman (Ann Arbor, USA); dimethyl sulfoxide (DMSO) (D4540), doxorubicin and DAPI were purchased from Sigma-Aldrich (St. Louis, USA); BSA was from BioShop (Burlington, Canada).

    Techniques: Western Blot, Activity Assay

    Effect of wortmannin on SK channel activity in the presence or absence of 10 μM phalloidin. The experiments were performed in cell-attached patches.

    Journal: American journal of physiology. Renal physiology

    Article Title: Inhibition of phosphatidylinositol 3-kinase stimulates activity of the small-conductance K channel in the CCD

    doi: 10.1152/ajprenal.00352.2005

    Figure Lengend Snippet: Effect of wortmannin on SK channel activity in the presence or absence of 10 μM phalloidin. The experiments were performed in cell-attached patches.

    Article Snippet: Phalloidin was obtained from Sigma and dissolved in methanol.

    Techniques: Activity Assay

    Time-lapse SHGM of the RNFL during continuous treatment with nocodazole. (a) Thickness of the RNFL at T = 0 min. (b) Axial cross section corresponding to the dashed line in (a). From top to bottom, at 0, 20, 40, and 60 min in treatment, respectively. The length of axis labels is 30 μ m. (c) Superposed margins of the retinal nerve fibers.

    Journal: Journal of Biomedical Optics

    Article Title: Effect of axonal micro-tubules on the morphology of retinal nerve fibers studied by second-harmonic generation

    doi: 10.1117/1.JBO.17.11.110502

    Figure Lengend Snippet: Time-lapse SHGM of the RNFL during continuous treatment with nocodazole. (a) Thickness of the RNFL at T = 0 min. (b) Axial cross section corresponding to the dashed line in (a). From top to bottom, at 0, 20, 40, and 60 min in treatment, respectively. The length of axis labels is 30 μ m. (c) Superposed margins of the retinal nerve fibers.

    Article Snippet: Initially the fresh retina received only Ames’ Medium, then perfusion was switched at time zero to the second medium containing 25-µM nocodazole and 0.4% dimethyl sulfoxide (DMSO, Sigma-Aldrich).

    Techniques:

    RNFL thickness and SHG intensity during nocodazole treatment ( N = 4).

    Journal: Journal of Biomedical Optics

    Article Title: Effect of axonal micro-tubules on the morphology of retinal nerve fibers studied by second-harmonic generation

    doi: 10.1117/1.JBO.17.11.110502

    Figure Lengend Snippet: RNFL thickness and SHG intensity during nocodazole treatment ( N = 4).

    Article Snippet: Initially the fresh retina received only Ames’ Medium, then perfusion was switched at time zero to the second medium containing 25-µM nocodazole and 0.4% dimethyl sulfoxide (DMSO, Sigma-Aldrich).

    Techniques:

    Figure 10. Rapamycin (Rap) enhanced high glucose toxic effect on adult mouse cardiomyocytes, whereas 3-methyladenine (3-MA) attenuated it. Cardiomyocytes were isolated from adult C57B/6 mice, cultured under the indicated glucose conditions for

    Journal: Autophagy

    Article Title: Suppression of autophagy is protective in high glucose-induced cardiomyocyte injury

    doi: 10.4161/auto.18980

    Figure Lengend Snippet: Figure 10. Rapamycin (Rap) enhanced high glucose toxic effect on adult mouse cardiomyocytes, whereas 3-methyladenine (3-MA) attenuated it. Cardiomyocytes were isolated from adult C57B/6 mice, cultured under the indicated glucose conditions for

    Article Snippet: Rapamycin (Sigma, R0395) was dissolved in absolute ethanol (Fisher Scientific, AC61508), 3-methyladenine (Sigma, M9281) in DMEM (GIBCO, 11966), and Bafilomycin A1 (LC Laboratories, B-1080) in dimethyl sulfoxide (DMSO; Sigma, 472301).

    Techniques: Isolation, Mouse Assay, Cell Culture

    Figure 9. mTORC1 mediated high glucose-induced phosphorylation of ULK1. Cardiomyocytes were cultured under different doses of glucose for 72 h. (A) The phosphorylation of ULK1 at Ser467 was determined by western blot analysis. (B) Rapamycin (Rap)

    Journal: Autophagy

    Article Title: Suppression of autophagy is protective in high glucose-induced cardiomyocyte injury

    doi: 10.4161/auto.18980

    Figure Lengend Snippet: Figure 9. mTORC1 mediated high glucose-induced phosphorylation of ULK1. Cardiomyocytes were cultured under different doses of glucose for 72 h. (A) The phosphorylation of ULK1 at Ser467 was determined by western blot analysis. (B) Rapamycin (Rap)

    Article Snippet: Rapamycin (Sigma, R0395) was dissolved in absolute ethanol (Fisher Scientific, AC61508), 3-methyladenine (Sigma, M9281) in DMEM (GIBCO, 11966), and Bafilomycin A1 (LC Laboratories, B-1080) in dimethyl sulfoxide (DMSO; Sigma, 472301).

    Techniques: Cell Culture, Western Blot

    Figure 3. Rapamycin (Rap) enhanced high-glucose-induced cardiomyocyte death, whereas 3-methyladenine (3-MA) attenuated it. Cardiomyocytes were treated with Rap (50 nM) or 3-MA (2 mM) for 24 h under the indicated glucose conditions. Cell death

    Journal: Autophagy

    Article Title: Suppression of autophagy is protective in high glucose-induced cardiomyocyte injury

    doi: 10.4161/auto.18980

    Figure Lengend Snippet: Figure 3. Rapamycin (Rap) enhanced high-glucose-induced cardiomyocyte death, whereas 3-methyladenine (3-MA) attenuated it. Cardiomyocytes were treated with Rap (50 nM) or 3-MA (2 mM) for 24 h under the indicated glucose conditions. Cell death

    Article Snippet: Rapamycin (Sigma, R0395) was dissolved in absolute ethanol (Fisher Scientific, AC61508), 3-methyladenine (Sigma, M9281) in DMEM (GIBCO, 11966), and Bafilomycin A1 (LC Laboratories, B-1080) in dimethyl sulfoxide (DMSO; Sigma, 472301).

    Techniques:

    Cisplatin concentration–dependent induction of p53, p53Ser15P, XPC, and DDB2. Cells were treated with the indicated concentrations of cisplatin (CP) at 37°C for 1 h. After treatment, cells were washed twice with PBS and incubated in drug-free

    Journal: Toxicological Sciences

    Article Title: Sodium Arsenite ? Hyperthermia Sensitizes p53-Expressing Human Ovarian Cancer Cells to Cisplatin by Modulating Platinum-DNA Damage Responses

    doi: 10.1093/toxsci/kfs085

    Figure Lengend Snippet: Cisplatin concentration–dependent induction of p53, p53Ser15P, XPC, and DDB2. Cells were treated with the indicated concentrations of cisplatin (CP) at 37°C for 1 h. After treatment, cells were washed twice with PBS and incubated in drug-free

    Article Snippet: Bovine serum albumin (BSA), MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide), RNase A, cisplatin, and sodium arsenite were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Concentration Assay, Incubation

    Suppression of cisplatin-induced XPC by sodium arsenite and hyperthermia and the effect of XPC siRNA transfection on cisplatin cytotoxicity. (A and B) Western blot analyses of XPC and p53Ser15P in A2780 and A2780/CP70 cells. Cells were treated with their

    Journal: Toxicological Sciences

    Article Title: Sodium Arsenite ? Hyperthermia Sensitizes p53-Expressing Human Ovarian Cancer Cells to Cisplatin by Modulating Platinum-DNA Damage Responses

    doi: 10.1093/toxsci/kfs085

    Figure Lengend Snippet: Suppression of cisplatin-induced XPC by sodium arsenite and hyperthermia and the effect of XPC siRNA transfection on cisplatin cytotoxicity. (A and B) Western blot analyses of XPC and p53Ser15P in A2780 and A2780/CP70 cells. Cells were treated with their

    Article Snippet: Bovine serum albumin (BSA), MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide), RNase A, cisplatin, and sodium arsenite were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Transfection, Western Blot

    Cellular platinum accumulation and effect of sodium arsenite and hyperthermia. (A) Cisplatin concentration–dependent accumulation of platinum in A2780 (▴), A2780/CP70 (▪), and SKOV-3 (•) cells. The regression equation for

    Journal: Toxicological Sciences

    Article Title: Sodium Arsenite ? Hyperthermia Sensitizes p53-Expressing Human Ovarian Cancer Cells to Cisplatin by Modulating Platinum-DNA Damage Responses

    doi: 10.1093/toxsci/kfs085

    Figure Lengend Snippet: Cellular platinum accumulation and effect of sodium arsenite and hyperthermia. (A) Cisplatin concentration–dependent accumulation of platinum in A2780 (▴), A2780/CP70 (▪), and SKOV-3 (•) cells. The regression equation for

    Article Snippet: Bovine serum albumin (BSA), MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide), RNase A, cisplatin, and sodium arsenite were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Concentration Assay

    Alteration of platinum bound to DNA by sodium arsenite and hyperthermia. A2780, A2780/CP70, and SKOV-3 cells were treated with 40μM cisplatin for 1 h at 37°C or 39°C. Total genomic DNA was isolated at 0 (immediately) and 24 h after

    Journal: Toxicological Sciences

    Article Title: Sodium Arsenite ? Hyperthermia Sensitizes p53-Expressing Human Ovarian Cancer Cells to Cisplatin by Modulating Platinum-DNA Damage Responses

    doi: 10.1093/toxsci/kfs085

    Figure Lengend Snippet: Alteration of platinum bound to DNA by sodium arsenite and hyperthermia. A2780, A2780/CP70, and SKOV-3 cells were treated with 40μM cisplatin for 1 h at 37°C or 39°C. Total genomic DNA was isolated at 0 (immediately) and 24 h after

    Article Snippet: Bovine serum albumin (BSA), MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide), RNase A, cisplatin, and sodium arsenite were purchased from Sigma-Aldrich (St Louis, MO).

    Techniques: Isolation

    Downregulation of Mcl-1 is necessary for paclitaxel-induced apoptosis of bladder cancer cells. ( a , b ) 5637 and HT1197 cells were treated with dimethyl sulfoxide (DMSO) or 0.1 µM paclitaxel for 24 and 48 h. Western blot analyses of PARP, cleaved caspase-3, cleaved caspase-9, Mcl-1, Bcl-xL, Bax, Bak, cyclin B1, and p-histone H3 Ser10 are shown. β-actin was used as a loading control. ( c ) Cells were subjected to FISH with a centromeric probe specific for chromosome 17 (Spectrum green). DNA was stained with DAPI (blue). At least 100 cells were counted for each condition, and the percentage of cells with normal ploidy or higher ploidy is presented in the histogram. Representative images are shown. Photomicrographs were taken using a 40× objective. ( d ) The expression of Mcl-1 was evaluated in samples of 72 patients with muscle-invasive or non-muscle-invasive bladder carcinoma by immunohistochemistry and quantified in the histogram. The relationship between the levels of expression of Mcl-1 and tumor infiltration in patients with bladder carcinoma was statistically significant. p -values were obtained using Fisher’s exact test. NMIBC: non-muscle-invasive bladder cancer; MIBC: muscle-invasive bladder cancer. Bars represent 100 μm. ( e ) Kaplan–Meier analysis of recurrence-free survival in patients with low (blue) and high (red) Mcl-1 expression. Tick marks represent censored patients. p -value

    Journal: Cancers

    Article Title: Obatoclax and Paclitaxel Synergistically Induce Apoptosis and Overcome Paclitaxel Resistance in Urothelial Cancer Cells

    doi: 10.3390/cancers10120490

    Figure Lengend Snippet: Downregulation of Mcl-1 is necessary for paclitaxel-induced apoptosis of bladder cancer cells. ( a , b ) 5637 and HT1197 cells were treated with dimethyl sulfoxide (DMSO) or 0.1 µM paclitaxel for 24 and 48 h. Western blot analyses of PARP, cleaved caspase-3, cleaved caspase-9, Mcl-1, Bcl-xL, Bax, Bak, cyclin B1, and p-histone H3 Ser10 are shown. β-actin was used as a loading control. ( c ) Cells were subjected to FISH with a centromeric probe specific for chromosome 17 (Spectrum green). DNA was stained with DAPI (blue). At least 100 cells were counted for each condition, and the percentage of cells with normal ploidy or higher ploidy is presented in the histogram. Representative images are shown. Photomicrographs were taken using a 40× objective. ( d ) The expression of Mcl-1 was evaluated in samples of 72 patients with muscle-invasive or non-muscle-invasive bladder carcinoma by immunohistochemistry and quantified in the histogram. The relationship between the levels of expression of Mcl-1 and tumor infiltration in patients with bladder carcinoma was statistically significant. p -values were obtained using Fisher’s exact test. NMIBC: non-muscle-invasive bladder cancer; MIBC: muscle-invasive bladder cancer. Bars represent 100 μm. ( e ) Kaplan–Meier analysis of recurrence-free survival in patients with low (blue) and high (red) Mcl-1 expression. Tick marks represent censored patients. p -value

    Article Snippet: The stock solutions of paclitaxel (Calbiochem, San Diego, CA, USA) and obatoclax (Selleck, Houston, TX, USA) were prepared at 10 mM in dimethyl sulfoxide (DMSO, Sigma) and stored at −20 °C.

    Techniques: Western Blot, Fluorescence In Situ Hybridization, Staining, Expressing, Immunohistochemistry

    The combination of paclitaxel and obatoclax in HT1376 and MDA-MB-231 cells overcomes paclitaxel resistance. HT1376 and MDA-MB-231 cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. ( a ) Western blot analyses of PARP, cleaved caspase-3, Mcl-1, p62, and LC3-B are shown. β-actin was used as a loading control. ( b ) Cell cycle analysis of propidium iodide-stained cells by flow cytometry. The quantification of each phase is shown in the histograms. ( c ) Cells were subjected to FISH with a centromeric probe specific for chromosome 17 (Spectrum green). DNA was stained with DAPI (blue). At least 100 cells were counted for each condition, and the percentage of cells with normal ploidy or higher ploidy is presented in the histogram.

    Journal: Cancers

    Article Title: Obatoclax and Paclitaxel Synergistically Induce Apoptosis and Overcome Paclitaxel Resistance in Urothelial Cancer Cells

    doi: 10.3390/cancers10120490

    Figure Lengend Snippet: The combination of paclitaxel and obatoclax in HT1376 and MDA-MB-231 cells overcomes paclitaxel resistance. HT1376 and MDA-MB-231 cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. ( a ) Western blot analyses of PARP, cleaved caspase-3, Mcl-1, p62, and LC3-B are shown. β-actin was used as a loading control. ( b ) Cell cycle analysis of propidium iodide-stained cells by flow cytometry. The quantification of each phase is shown in the histograms. ( c ) Cells were subjected to FISH with a centromeric probe specific for chromosome 17 (Spectrum green). DNA was stained with DAPI (blue). At least 100 cells were counted for each condition, and the percentage of cells with normal ploidy or higher ploidy is presented in the histogram.

    Article Snippet: The stock solutions of paclitaxel (Calbiochem, San Diego, CA, USA) and obatoclax (Selleck, Houston, TX, USA) were prepared at 10 mM in dimethyl sulfoxide (DMSO, Sigma) and stored at −20 °C.

    Techniques: Multiple Displacement Amplification, Western Blot, Cell Cycle Assay, Staining, Flow Cytometry, Cytometry, Fluorescence In Situ Hybridization

    Obatoclax induces S arrest and a delayed cell cycle progression when combined with paclitaxel. Cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. ( a ) Cell cycle analysis of propidium iodide-stained cells by flow cytometry. Quantification of each phase is shown in the histograms. ( b ) Ploidy analysis for chromosome 17 by FISH. At least 100 cells were counted for each condition, and the percentage of cells with normal ploidy or higher ploidy is presented in the histogram. OBT: obatoclax; PTX: paclitaxel.

    Journal: Cancers

    Article Title: Obatoclax and Paclitaxel Synergistically Induce Apoptosis and Overcome Paclitaxel Resistance in Urothelial Cancer Cells

    doi: 10.3390/cancers10120490

    Figure Lengend Snippet: Obatoclax induces S arrest and a delayed cell cycle progression when combined with paclitaxel. Cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. ( a ) Cell cycle analysis of propidium iodide-stained cells by flow cytometry. Quantification of each phase is shown in the histograms. ( b ) Ploidy analysis for chromosome 17 by FISH. At least 100 cells were counted for each condition, and the percentage of cells with normal ploidy or higher ploidy is presented in the histogram. OBT: obatoclax; PTX: paclitaxel.

    Article Snippet: The stock solutions of paclitaxel (Calbiochem, San Diego, CA, USA) and obatoclax (Selleck, Houston, TX, USA) were prepared at 10 mM in dimethyl sulfoxide (DMSO, Sigma) and stored at −20 °C.

    Techniques: Cell Cycle Assay, Staining, Flow Cytometry, Cytometry, Fluorescence In Situ Hybridization

    Autophagic flux blockade correlates with apoptotic cell death in obatoclax plus paclitaxel-treated HT1197 cells. Cells were treated with 1 µM obatoclax or 1 µM obatoclax and 0.1 µM paclitaxel for 48 h. DMSO, 800 nM rapamycin, and 50 µM chloroquine were used as controls. ( a ) Obatoclax-blocked autophagic flux at 48 h was measured by flow cytometry-based profiling of Cyto-ID TM Autophagy Detection in 5637 and HT1197 cell lines. Histograms show the mean intensity of each treatment. Data were presented as the mean ± SD. * p -value

    Journal: Cancers

    Article Title: Obatoclax and Paclitaxel Synergistically Induce Apoptosis and Overcome Paclitaxel Resistance in Urothelial Cancer Cells

    doi: 10.3390/cancers10120490

    Figure Lengend Snippet: Autophagic flux blockade correlates with apoptotic cell death in obatoclax plus paclitaxel-treated HT1197 cells. Cells were treated with 1 µM obatoclax or 1 µM obatoclax and 0.1 µM paclitaxel for 48 h. DMSO, 800 nM rapamycin, and 50 µM chloroquine were used as controls. ( a ) Obatoclax-blocked autophagic flux at 48 h was measured by flow cytometry-based profiling of Cyto-ID TM Autophagy Detection in 5637 and HT1197 cell lines. Histograms show the mean intensity of each treatment. Data were presented as the mean ± SD. * p -value

    Article Snippet: The stock solutions of paclitaxel (Calbiochem, San Diego, CA, USA) and obatoclax (Selleck, Houston, TX, USA) were prepared at 10 mM in dimethyl sulfoxide (DMSO, Sigma) and stored at −20 °C.

    Techniques: Flow Cytometry, Cytometry

    The combined treatment of Mcl-1 antagonist obatoclax with paclitaxel induces apoptosis in HT1197 cells. Cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. The combinations were: obatoclax followed by paclitaxel after 8 h, paclitaxel followed by obatoclax after 8 h, and both at the same time. In the 5637 and HT1197 cell lines, Western blot analyses of PARP, cleaved caspase 3, Mcl-1, and Bak are shown. β-actin is shown as a loading control. Histograms show the densitometric analysis of indicated proteins. Data are presented as the mean ± SD, and each treatment was compared with DMSO as the control. * p -value

    Journal: Cancers

    Article Title: Obatoclax and Paclitaxel Synergistically Induce Apoptosis and Overcome Paclitaxel Resistance in Urothelial Cancer Cells

    doi: 10.3390/cancers10120490

    Figure Lengend Snippet: The combined treatment of Mcl-1 antagonist obatoclax with paclitaxel induces apoptosis in HT1197 cells. Cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. The combinations were: obatoclax followed by paclitaxel after 8 h, paclitaxel followed by obatoclax after 8 h, and both at the same time. In the 5637 and HT1197 cell lines, Western blot analyses of PARP, cleaved caspase 3, Mcl-1, and Bak are shown. β-actin is shown as a loading control. Histograms show the densitometric analysis of indicated proteins. Data are presented as the mean ± SD, and each treatment was compared with DMSO as the control. * p -value

    Article Snippet: The stock solutions of paclitaxel (Calbiochem, San Diego, CA, USA) and obatoclax (Selleck, Houston, TX, USA) were prepared at 10 mM in dimethyl sulfoxide (DMSO, Sigma) and stored at −20 °C.

    Techniques: Western Blot

    LC3-II and p62 accumulate in obatoclax-treated 5637 cells and HT1197 cells treated with obatoclax plus paclitaxel. The cleavage of beclin-1 by caspase 3 induces apoptosis as a mechanism of sensitization. ( a ) Cells were treated with DMSO or 1 µM obatoclax for 48 h, in the absence or presence of the autophagy inhibitor bafilomycin A1 400 nM for 4 h. ( b ) Cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. Western blot analyses of LC3-B and p62 are shown. β-actin is shown as a loading control. OBT: obatoclax; PTX: paclitaxel. ( c ) The 5637 cell line was treated with DMSO or 1 µM obatoclax for 48 h in the presence or absence of pre-incubation with pan-caspase inhibitor Z-VAD-fmk 20 µM for 1 h. ( d ) 5637 and HT1197 cells were treated with DMSO, 1 μM obatoclax, or 1 µM obatoclax plus 0.1 µM paclitaxel for 48 h. Western blot analyses of beclin-1 and cleaved caspase-3 are shown. Symbol ◄ indicates an unspecific band of 50-kDa beclin-1. β-actin is shown as a loading control.

    Journal: Cancers

    Article Title: Obatoclax and Paclitaxel Synergistically Induce Apoptosis and Overcome Paclitaxel Resistance in Urothelial Cancer Cells

    doi: 10.3390/cancers10120490

    Figure Lengend Snippet: LC3-II and p62 accumulate in obatoclax-treated 5637 cells and HT1197 cells treated with obatoclax plus paclitaxel. The cleavage of beclin-1 by caspase 3 induces apoptosis as a mechanism of sensitization. ( a ) Cells were treated with DMSO or 1 µM obatoclax for 48 h, in the absence or presence of the autophagy inhibitor bafilomycin A1 400 nM for 4 h. ( b ) Cells were treated with 1 μM obatoclax, 0.1 µM paclitaxel, and combinations of both for 48 h. Western blot analyses of LC3-B and p62 are shown. β-actin is shown as a loading control. OBT: obatoclax; PTX: paclitaxel. ( c ) The 5637 cell line was treated with DMSO or 1 µM obatoclax for 48 h in the presence or absence of pre-incubation with pan-caspase inhibitor Z-VAD-fmk 20 µM for 1 h. ( d ) 5637 and HT1197 cells were treated with DMSO, 1 μM obatoclax, or 1 µM obatoclax plus 0.1 µM paclitaxel for 48 h. Western blot analyses of beclin-1 and cleaved caspase-3 are shown. Symbol ◄ indicates an unspecific band of 50-kDa beclin-1. β-actin is shown as a loading control.

    Article Snippet: The stock solutions of paclitaxel (Calbiochem, San Diego, CA, USA) and obatoclax (Selleck, Houston, TX, USA) were prepared at 10 mM in dimethyl sulfoxide (DMSO, Sigma) and stored at −20 °C.

    Techniques: Western Blot, Incubation

    Outer membrane permeabilization assays of the engineered peptides. A 2 mL volume of E. coli UB1005 cells, diluted to 10 5 CFU/mL in 5 mM sodium HEPES buffer, was added to a quartz cuvette containing NPN to give a final concentration of 10 μM. Aliquots of peptides were added to the cuvette, and the fluorescence was recorded until there was no further increase in fluorescence. Data shown are three independent experiments.

    Journal: International Journal of Molecular Sciences

    Article Title: Simplified Head-to-Tail Cyclic Polypeptides as Biomaterial-Associated Antimicrobials with Endotoxin Neutralizing and Anti-Inflammatory Capabilities

    doi: 10.3390/ijms20235904

    Figure Lengend Snippet: Outer membrane permeabilization assays of the engineered peptides. A 2 mL volume of E. coli UB1005 cells, diluted to 10 5 CFU/mL in 5 mM sodium HEPES buffer, was added to a quartz cuvette containing NPN to give a final concentration of 10 μM. Aliquots of peptides were added to the cuvette, and the fluorescence was recorded until there was no further increase in fluorescence. Data shown are three independent experiments.

    Article Snippet: TritonX-100, propidium iodide (PI), Polymyxin B, o-nitrophenyl-b-D-galactopyranoside (ONPG), N-phenyl-1-napthylamine (NPN), 3.3’-depropylthiadicarbocyanine iodide (DiSC3-5), HEPES, ethanol (analytical grade, 99%), acetone (analytical grade, 99%), lipopolysaccharide (LPS, derived from E. coli 055: B5) and BODIPY-TR-cadaverine were all obtained from Sigma (Shanghai, China).

    Techniques: Concentration Assay, Fluorescence

    Effect of sophorolipid on C. albicans mature hyphae. C. albicans cells were grown in RPMI-1640 containing 10% FBS for 5 hrs. After that mature hypha were treated with indicated concentrations of SL for time point zero (upper panel) and 5 hrs (lower panel) at 37 °C. At the end of incubation an aliquot was withdrawn from each sample and photographed at 60× magnification.

    Journal: Scientific Reports

    Article Title: Inhibitory Effect of Sophorolipid on Candida albicans Biofilm Formation and Hyphal Growth

    doi: 10.1038/srep23575

    Figure Lengend Snippet: Effect of sophorolipid on C. albicans mature hyphae. C. albicans cells were grown in RPMI-1640 containing 10% FBS for 5 hrs. After that mature hypha were treated with indicated concentrations of SL for time point zero (upper panel) and 5 hrs (lower panel) at 37 °C. At the end of incubation an aliquot was withdrawn from each sample and photographed at 60× magnification.

    Article Snippet: RPMI-1640 medium with L-glutamine without sodium bicarbonate (Sigma) was buffered with 0.165 M morpholinepropanesulfonic acid (Sigma) to a pH of 7.

    Techniques: Incubation

    Effect of sophorolipid on C. albicans hyphal growth. C. albicans cells were grown in RPMI-1640 medium (upper panel) and RPMI-1640 containing 10% FBS (lower panel) at the indicated concentration of SL at 37 °C for 5 hrs. At the end of incubation an aliquot was withdrawn from each sample and photographed at 100× magnification.

    Journal: Scientific Reports

    Article Title: Inhibitory Effect of Sophorolipid on Candida albicans Biofilm Formation and Hyphal Growth

    doi: 10.1038/srep23575

    Figure Lengend Snippet: Effect of sophorolipid on C. albicans hyphal growth. C. albicans cells were grown in RPMI-1640 medium (upper panel) and RPMI-1640 containing 10% FBS (lower panel) at the indicated concentration of SL at 37 °C for 5 hrs. At the end of incubation an aliquot was withdrawn from each sample and photographed at 100× magnification.

    Article Snippet: RPMI-1640 medium with L-glutamine without sodium bicarbonate (Sigma) was buffered with 0.165 M morpholinepropanesulfonic acid (Sigma) to a pH of 7.

    Techniques: Concentration Assay, Incubation

    Effect of sophorolipid on the expression of C. albicans hypha specific genes. C. albicans cells were incubated in the absence (control) or presence (15 μg/ml) of SL in RPMI-1640 medium at 37 °C for 5 hrs. Following incubation expression of the indicated genes were determined by qRT-PCR. Expression level of each gene is displayed after normalization with internal control housekeeping gene ACT1 . The histogram shows the relative expression fold change of genes by SL treatment with respect to the control. Results represent the average of three independent experiments ± SD. * p

    Journal: Scientific Reports

    Article Title: Inhibitory Effect of Sophorolipid on Candida albicans Biofilm Formation and Hyphal Growth

    doi: 10.1038/srep23575

    Figure Lengend Snippet: Effect of sophorolipid on the expression of C. albicans hypha specific genes. C. albicans cells were incubated in the absence (control) or presence (15 μg/ml) of SL in RPMI-1640 medium at 37 °C for 5 hrs. Following incubation expression of the indicated genes were determined by qRT-PCR. Expression level of each gene is displayed after normalization with internal control housekeeping gene ACT1 . The histogram shows the relative expression fold change of genes by SL treatment with respect to the control. Results represent the average of three independent experiments ± SD. * p

    Article Snippet: RPMI-1640 medium with L-glutamine without sodium bicarbonate (Sigma) was buffered with 0.165 M morpholinepropanesulfonic acid (Sigma) to a pH of 7.

    Techniques: Expressing, Incubation, Quantitative RT-PCR