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  • 99
    ATCC dimethyl sulfoxide dmso
    Dimethyl Sulfoxide Dmso, supplied by ATCC, used in various techniques. Bioz Stars score: 99/100, based on 51 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore dimethyl sulfoxide dmso
    Phenotypic analysis of zebrafish after morpholino-mediated knockdown of vcp or ML240 treatment. ( a to d ) Lateral view of embryos at 60 <t>hpf.</t> Zebrafish embryos were either injected with 5 base pair (bp) mismatch vcp start morpholino (MO1- control ; a ) and vcp start MO (MO1- vcp ; b ) or treated with <t>DMSO</t> ( c ) and VCP-Inhibitor ML240 (VCP-I; d ). scale bar: 500 μm ( a, c ); scale bar: 650 μm ( b, d ). ( e ) Statistical analysis of affected embryos after MO1/2- control or MO1/2- vcp injection and DMSO or ML240 treatment. N = 3/4 Experiments with in total n = 259 (MO1- control ), n = 161 (MO2- control ), n = 242 (MO1- vcp ), n = 212 (MO2- vcp ), n = 152 (DMSO), n = 148 (VCP-I); N two-sided Fisher exact tests combined with a modified Fisher method. Ratios of N experiments are displayed. MO1- vcp -injected embryos reveal reduced levels of Vcp by western blot analysis. Actb1/β-actin was used as loading control. RT-PCR after MO2- vcp injection shows specific effect on mRNA splicing. ( f ) Ventricular fractional shortening (FS) measurements of MO1- control and MO1- vcp -injected embryos at 48, 72 and 96 hpf, n = 18 to 30 (MO1- control ), n = 19 to 32 (MO1- vcp ). The individual measurements are shown (two-sided Wilcoxon rank-sum test). ( g ) Heart beat per minute at 48, 72 and 96 hpf of vcp morphants and control embryos. n = 30 (MO1- control ), n = 29 or 30 (MO1- vcp ). The individual measurements are shown (two-sided unpaired Wilcoxon rank-sum test. ( h and i ) False-colored superimposed overviews of spontaneous movement assay with MO1- control - ( h ) and MO1- vcp -injected ( i ) embryos at 24 hpf; red pictures = 0 s; green pictures = 10 s; yellow = merge. scale bar: 1 mm. ( j ) Quantification of spontaneous movement assay; N = 3 experiments with n = 30 (MO1- control ), n = 29 or 30 (MO1- vcp ) per experiment (N two-sided Fisher exact tests combined with modified Fisher method). The ratios of N experiments are shown. ( k ) Statistical analysis of touch-evoke flight response of control and vcp morphants at 72 hpf. n = 30, N = 3 (MO1- control ), n = 29 or 30, N = 3 (MO1- vcp ); data represent means.
    Dimethyl Sulfoxide Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 20430 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Thermo Fisher dimethyl sulfoxide dmso
    Heat map of <t>panobinostat</t> -induced gene expression changes in MDA-MB-231, MDA-MB-468, and MCF-7 cells . MDA-MB-231, MDA-MB-468, or MCF-7 cells were treated for 24 hours with panobinostat (100 nM) or vehicle <t>(DMSO)</t> and then assayed via the Human Breast Cancer and Estrogen Receptor Signaling RT 2 Profiler™ PCR Array. Red signifies up-regulation and green signifies down-regulation by panobinostat compared to MDA-MB-231 vehicle treated controls. Data are representative of three independent experiments. Genes regulated at least 2-fold are also summarized in Additional file 1 (MDA-MB-231), 2 (MDA-MB-468) and 3 (MCF-7). DMSO, dimethyl sulfoxide.
    Dimethyl Sulfoxide Dmso, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 92/100, based on 2578 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Merck KGaA dimethyl sulfoxide dmso
    In vivo activity of <t>EN4</t> in a mouse tissue cage model. The numbers of planktonic SA113 (A) and the viability of leukocytes (B) present in tissue cage fluid from cages treated with 100 or 250 μg of EN4 or 30 μg of daptomycin (DAP) and infected with 4 × 10 3 CFU of SA113 were investigated. Open circles indicate control mice injected with 1% <t>DMSO-PBS.</t> The values shown are means ± the SDs.
    Dimethyl Sulfoxide Dmso, supplied by Merck KGaA, used in various techniques. Bioz Stars score: 94/100, based on 813 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Fisher Scientific dimethyl sulfoxide dmso
    Analogs of <t>honokiol</t> do not bind or inhibit FOXM1 a C3-luc cells were induced with doxycycline and treated with the indicated analogs of honokiol at the concentration of 40 µM for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b DU145 prostate cancer cells were treated as indicated. Following treatment cells were collected and immunoblotting was performed for FOXM1, cleaved caspase-3 and β-actin as the loading control. c STD NMR spectra of 2 mM of: (I) 2 mM eugenol, (II) 2 mM O -eugenol, (III) 2 mM 2-allylphenol and (IV) 2 mM 2,2 dihydroxybiphenyl. The chemical structure of each small molecule is illustrated. Signals from vehicle <t>(DMSO)</t> and water are labeled. d STD NMR spectra of 150 ng of recombinant FOXM1 plus: (I) 2 mM eugenol, (II) 2 mM O-eugenol, (III) 2 mM 2,2 dihydroxybiphenyl and (IV) 2 mM 2-allylphenol. The chemical structure of each small molecule is illustrated. STD signals arising from the aryl groups are annotated and signals from vehicle (DMSO) and water are labeled
    Dimethyl Sulfoxide Dmso, supplied by Fisher Scientific, used in various techniques. Bioz Stars score: 93/100, based on 730 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore dimethyl sulfoxide
    Analogs of <t>honokiol</t> do not bind or inhibit FOXM1 a C3-luc cells were induced with doxycycline and treated with the indicated analogs of honokiol at the concentration of 40 µM for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b DU145 prostate cancer cells were treated as indicated. Following treatment cells were collected and immunoblotting was performed for FOXM1, cleaved caspase-3 and β-actin as the loading control. c STD NMR spectra of 2 mM of: (I) 2 mM eugenol, (II) 2 mM O -eugenol, (III) 2 mM 2-allylphenol and (IV) 2 mM 2,2 dihydroxybiphenyl. The chemical structure of each small molecule is illustrated. Signals from vehicle <t>(DMSO)</t> and water are labeled. d STD NMR spectra of 150 ng of recombinant FOXM1 plus: (I) 2 mM eugenol, (II) 2 mM O-eugenol, (III) 2 mM 2,2 dihydroxybiphenyl and (IV) 2 mM 2-allylphenol. The chemical structure of each small molecule is illustrated. STD signals arising from the aryl groups are annotated and signals from vehicle (DMSO) and water are labeled
    Dimethyl Sulfoxide, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 27840 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    FUJIFILM dimethyl sulfoxide dmso
    Supervised cluster analysis and GSEA between PD-L1 -high and -low human lung cancer cell lines suggested mRNA upregulation of several molecules related to cell adhesion, motility, cell proliferation, cell signaling, and pathways related to actin cytoskeleton regulation and integrin-mediated cell adhesion. A, Supervised cluster analysis of 10 PD-L1 -high versus 10 PD-L1 -low human lung cancer cell lines. Depicted by the CCLE Analysis Tools: Differential Expression ( http://www.broadinstitute.org/ccle/data/browseAnalyses ). B, qRT-PCR of 11 selected mRNAs in PD-L1 -high and -low cell lines. The ΔΔCt method was used for the target mRNA, which was normalized by endogenous β-actin mRNA and reference BEAS-2B cells. RQ data are the mean of 10 cell lines. C, RAC2 and CDA mRNA levels were compared between <t>DMSO</t> and <t>U0126</t> treatments in the three KRAS -mutant lung adenocarcinoma cell lines. The target mRNAs were normalized with endogenous β-actin mRNAs and DMSO-treated cells as a reference (ΔΔCt method). Data are the mean of three independent experiments. D, GSEA showed significant results for pathways related to actin cytoskeleton regulation by Rho GTPases and to integrin-mediated cell adhesion.
    Dimethyl Sulfoxide Dmso, supplied by FUJIFILM, used in various techniques. Bioz Stars score: 93/100, based on 323 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Merck & Co dimethyl sulfoxide dmso
    The combination of <t>LCRF‐0006</t> and bortezomib synergistically increases MM tumor response in vivo. A, Schematic representation of treatment regimen. C57BL/KaLwRij mice with established MM (day 14 post‐5TGM1 cell injection) were treated with LCRF‐0006 (100 mg/kg) and bortezomib (0.5 mg/kg) (LCRF‐0006 + bortezomib), LCRF‐0006 and <t>DMSO</t> vehicle (LCRF‐0006), 2‐HP‐b‐CD vehicle and bortezomib (bortezomib) or 2‐HP‐b‐CD and DMSO vehicles (vehicles) i.p., as per treatment regimen. B, Tumor burden was assessed at day 14, 21 and 28 by BLI. Graph depicts mean ± SEM. n = 5‐6 mice/treatment group. *** P
    Dimethyl Sulfoxide Dmso, supplied by Merck & Co, used in various techniques. Bioz Stars score: 92/100, based on 381 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Amresco dimethyl sulfoxide dmso
    An mTOR inhibitor <t>AZD8055</t> preferentially inhibited proliferation and phosphorylation levels in NCI N87 T-R cells (A) T-S and T-R cells were cultured for 24 h, and then treated with or without AZD8055 at the indicated concentrations (0, 25, 50, 100, 200, 400 and 800 nM) or <t>DMSO</t> as control, respectively. Cell viability was determined by Cell Count kit-8 assay 48 h later. Results expressed as % control represent the mean of three experiments. * p
    Dimethyl Sulfoxide Dmso, supplied by Amresco, used in various techniques. Bioz Stars score: 94/100, based on 428 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Sinopharm dimethyl sulfoxide dmso
    The UV spectra of PBDE hapten, protein and conjugates: ( a ) with <t>BSA</t> and ( b ) with OVA; absorbance value at the characteristic peak, 292 nm: OD BSA-PBDE = 1.860, OD PBDE-hapten = 1.893, OD BSA = 0.074; 272 nm: OD OVA-PBDE = 1.767, OD PBDE - hapten = 1.893, OD OVA = 0.401; C BSA : 0.20 g/L, C OVA : 0.20 g/L, and C hapten : 0.05 g/L; protein and conjugate were dissolved in PBS buffer; hapten was dissolved in <t>DMSO.</t>
    Dimethyl Sulfoxide Dmso, supplied by Sinopharm, used in various techniques. Bioz Stars score: 93/100, based on 268 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore anhydrous dimethyl sulfoxide dmso
    The UV spectra of PBDE hapten, protein and conjugates: ( a ) with <t>BSA</t> and ( b ) with OVA; absorbance value at the characteristic peak, 292 nm: OD BSA-PBDE = 1.860, OD PBDE-hapten = 1.893, OD BSA = 0.074; 272 nm: OD OVA-PBDE = 1.767, OD PBDE - hapten = 1.893, OD OVA = 0.401; C BSA : 0.20 g/L, C OVA : 0.20 g/L, and C hapten : 0.05 g/L; protein and conjugate were dissolved in PBS buffer; hapten was dissolved in <t>DMSO.</t>
    Anhydrous Dimethyl Sulfoxide Dmso, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 298 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Avantor dimethyl sulfoxide dmso
    Controlling TC-83 induced ROS response in microglial cells decreases inflammatory burden. (a) Experimental workflow indicating HMC3 cells were pre-treated for 2h with 0.1% <t>DMSO,</t> 1μM BAY 11–7082 (BAY-82), or 100pM <t>mitoquinone</t> mesylate (MitoQ) prior to inoculation with TC-83 (MOI:2). Levels of the pro-inflammatory cytokines (b) IL-1α, (c) IL-1β, (d), IL-6 and (e) IL-8 produced by HMC3 cells as the direct result of TC-83 infection were measured at 1, 2, and 6hpi. The quantitative data are depicted as the means of three biologically independent experiments ± SD. * p
    Dimethyl Sulfoxide Dmso, supplied by Avantor, used in various techniques. Bioz Stars score: 93/100, based on 174 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Beyotime dimethyl sulfoxide dmso
    Inhibition of the Piezo1 channel in bladder ICC-LCs by <t>GsMTx4</t> significantly alleviated bladder hyperactivity in CYP-8d rats. a – c Compared with the vehicle <t>(DMSO),</t> 2.5 μM GsMTx4 significantly decreased the MBP and prolonged the ICI in naive and CYP-8d rats, and naive rats showed lower sensitivity to GsMTx4 than CYP-8d rats (the data represent the mean ± S.D., n = 8; * P
    Dimethyl Sulfoxide Dmso, supplied by Beyotime, used in various techniques. Bioz Stars score: 92/100, based on 191 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Applichem dimethyl sulfoxide dmso
    Infliximab prevents <t>Zaprinast-induced</t> glial fibrillary acidic protein (GFAP) overexpression in cultured porcine retina. Retinal explants were incubated with dimethyl sulfoxide <t>(DMSO),</t> Zaprinast and Infliximab alone or combined with Zaprinast as described in Methods. (A) Confocal laser scanning micrographs of retinal sections showing GFAP content. Scale bar: 50 μm. (B) Bar graphs showing the quantification of GFAP content. Values are the mean ±SEM of six different cultures. Values that are significantly different are indicated by asterisks * P
    Dimethyl Sulfoxide Dmso, supplied by Applichem, used in various techniques. Bioz Stars score: 92/100, based on 126 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Valiant dimethyl sulfoxide dmso
    Inhibition of ALDH1A2 expression by GSK3 inhibitor <t>CHIR99021</t> is via β-catenin signaling. (A) Effect of CHIR99021 on β-catenin nuclear translocation in WiT49 cells. WiT49 cells were treated with 5 μM CHIR99021 or 0.1% <t>DMSO</t> for 48 h, and then immunofluorescence was performed with β-catenin antibody and DAPI staining. Negative control represents no primary antibody control. Bar length equals 50 μm. (B) Effect of CHIR99021 on the TCF reporter in WiT49 cells. A 500-ng sample of TOPFlash or FOPFlash constructs, together with 50 ng of the pRL-TK renilla control vector, was transiently transfected into WiT49 cells. Cells were treated with 0.1% DMSO or 5 μM CHIR99021 after 24 h of transfection. Luciferase reporter activities relative to renilla (pRL-TK) are shown in percentage terms. Values are the mean ± SD of three determinations from separate experiments. ∗∗∗ P
    Dimethyl Sulfoxide Dmso, supplied by Valiant, used in various techniques. Bioz Stars score: 93/100, based on 153 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Nacalai dimethyl sulfoxide dmso
    Increases in Mcl-1 and Bcl-xL expression were mediated by YAP1. a Relative gene expression of MCL1 and BCLXL detected by qRT-PCR in 4 ALK-rearranged lung cancer cell lines ( n = 4 independent experiments). Significance was evaluated using a one-way ANOVA followed by Sidak’s test. b Evaluation of Mcl-1 expression, Bcl-xL expression, and YAP phosphorylation (pS127). Cell lysates were blotted using an immunoblotting assay with the indicated antibody. c Initial survival rate (top) and apoptosis activity (bottom) of KTOR1 (left) and KTOR2 (right) when cells were exposed to <t>ALC</t> or vehicle in combination with the knockdown of MCL1 and BCLXL . Summarized results are shown in the panels. The number of independent experiments is shown in each panel. Complete results are presented in Supplementary Fig. 6c, d . Significance was evaluated using a one-way ANOVA followed by Sidak’s test. d – e Increases in Mcl-1 and Bcl-xL expression were canceled by the inhibition of YAP1. YAP1 activity was inhibited using siRNA ( d ) or verteporfin ( e ). Traced experiments using H2228 are shown in Supplementary Fig. 6f, g . f Sequence analysis of putative YAP1/TEAD binding sites (labeled as A, B) in the upstream regions of MCL1 and BCLXL . The ChIP-PCR primer pairs designed were shown as arrows. g ChIP-qPCR on KTOR1 and KTOR2, which confirmed the presence of YAP1/TEAD binding sites in the MCL1 and BCLXL upstream regions and increased binding induced by the ALC treatment. BCLXL , a known YAP target, was also used as a positive control. Results are expressed as Percent Input (Two biologically independent cells, two-independent primers, two-independent experiment in each cell lines and two-independent qRT-PCR quantification in each experiment). The titration of DNA digestion, electrophoresis of input samples, and specificity of ChIP-qRT-PCR primers were shown in Supplementary Fig. 8 . Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. h Schematic explanation of Fig. 6. The inhibition of YAP1 down-regulated Mcl-1 and Bcl-xL expression. The inhibition of both Mcl-1 and Bcl-xL induced apoptosis and inhibited initial survival. ALC alectinib, <t>DMSO</t> dimethyl sulfoxide, VER verteporfin, siSC si scramble control. Error bars indicate ± S.D. * P
    Dimethyl Sulfoxide Dmso, supplied by Nacalai, used in various techniques. Bioz Stars score: 92/100, based on 99 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc dimethyl sulfoxide dmso
    Increases in Mcl-1 and Bcl-xL expression were mediated by YAP1. a Relative gene expression of MCL1 and BCLXL detected by qRT-PCR in 4 ALK-rearranged lung cancer cell lines ( n = 4 independent experiments). Significance was evaluated using a one-way ANOVA followed by Sidak’s test. b Evaluation of Mcl-1 expression, Bcl-xL expression, and YAP phosphorylation (pS127). Cell lysates were blotted using an immunoblotting assay with the indicated antibody. c Initial survival rate (top) and apoptosis activity (bottom) of KTOR1 (left) and KTOR2 (right) when cells were exposed to <t>ALC</t> or vehicle in combination with the knockdown of MCL1 and BCLXL . Summarized results are shown in the panels. The number of independent experiments is shown in each panel. Complete results are presented in Supplementary Fig. 6c, d . Significance was evaluated using a one-way ANOVA followed by Sidak’s test. d – e Increases in Mcl-1 and Bcl-xL expression were canceled by the inhibition of YAP1. YAP1 activity was inhibited using siRNA ( d ) or verteporfin ( e ). Traced experiments using H2228 are shown in Supplementary Fig. 6f, g . f Sequence analysis of putative YAP1/TEAD binding sites (labeled as A, B) in the upstream regions of MCL1 and BCLXL . The ChIP-PCR primer pairs designed were shown as arrows. g ChIP-qPCR on KTOR1 and KTOR2, which confirmed the presence of YAP1/TEAD binding sites in the MCL1 and BCLXL upstream regions and increased binding induced by the ALC treatment. BCLXL , a known YAP target, was also used as a positive control. Results are expressed as Percent Input (Two biologically independent cells, two-independent primers, two-independent experiment in each cell lines and two-independent qRT-PCR quantification in each experiment). The titration of DNA digestion, electrophoresis of input samples, and specificity of ChIP-qRT-PCR primers were shown in Supplementary Fig. 8 . Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. h Schematic explanation of Fig. 6. The inhibition of YAP1 down-regulated Mcl-1 and Bcl-xL expression. The inhibition of both Mcl-1 and Bcl-xL induced apoptosis and inhibited initial survival. ALC alectinib, <t>DMSO</t> dimethyl sulfoxide, VER verteporfin, siSC si scramble control. Error bars indicate ± S.D. * P
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    Carl Roth GmbH dimethyl sulfoxide dmso
    ( A ) SEC elugram of <t>AMCA-PLGA;</t> the black line indicates the refractive index of the PLGA expressed as relative units (RIU), and the gray line indicates the ultraviolet absorption by AMCA at 350 nm expressed as relative milli-units (mAU). ( B ) Stability studies via fluorescence intensity measurements of AMCA-PLGA in <t>DMSO</t> for > 120 hours at 4°C (solid line, crosses), 20°C (dotted line, triangles) and 37°C (broken line, circles). Data are expressed as ratios of I/I 0 and shown as the mean ± SD of three independent experiments. Abbreviations: AMCA-PLGA, 7-amino-4-methyl-3-coumarinylacetic acid-poly( d,l -lactide-co-glycolide); DMSO, dimethyl sulfoxide; mAU, milli-absorption unit; PLGA, poly( d,l -lactide-co-glycolide); RIU, refractive index unit; SD, standard deviation; SEC, size exclusion chromatography.
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    Millipore dmso solution
    ( A ) SEC elugram of <t>AMCA-PLGA;</t> the black line indicates the refractive index of the PLGA expressed as relative units (RIU), and the gray line indicates the ultraviolet absorption by AMCA at 350 nm expressed as relative milli-units (mAU). ( B ) Stability studies via fluorescence intensity measurements of AMCA-PLGA in <t>DMSO</t> for > 120 hours at 4°C (solid line, crosses), 20°C (dotted line, triangles) and 37°C (broken line, circles). Data are expressed as ratios of I/I 0 and shown as the mean ± SD of three independent experiments. Abbreviations: AMCA-PLGA, 7-amino-4-methyl-3-coumarinylacetic acid-poly( d,l -lactide-co-glycolide); DMSO, dimethyl sulfoxide; mAU, milli-absorption unit; PLGA, poly( d,l -lactide-co-glycolide); RIU, refractive index unit; SD, standard deviation; SEC, size exclusion chromatography.
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    Junsei Chemical dimethyl sulfoxide dmso
    ( A ) SEC elugram of <t>AMCA-PLGA;</t> the black line indicates the refractive index of the PLGA expressed as relative units (RIU), and the gray line indicates the ultraviolet absorption by AMCA at 350 nm expressed as relative milli-units (mAU). ( B ) Stability studies via fluorescence intensity measurements of AMCA-PLGA in <t>DMSO</t> for > 120 hours at 4°C (solid line, crosses), 20°C (dotted line, triangles) and 37°C (broken line, circles). Data are expressed as ratios of I/I 0 and shown as the mean ± SD of three independent experiments. Abbreviations: AMCA-PLGA, 7-amino-4-methyl-3-coumarinylacetic acid-poly( d,l -lactide-co-glycolide); DMSO, dimethyl sulfoxide; mAU, milli-absorption unit; PLGA, poly( d,l -lactide-co-glycolide); RIU, refractive index unit; SD, standard deviation; SEC, size exclusion chromatography.
    Dimethyl Sulfoxide Dmso, supplied by Junsei Chemical, used in various techniques. Bioz Stars score: 93/100, based on 77 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Toxicity of the four selected small molecules <t>(SMs)</t> on several eukaryotic models. ( A ) Cell toxicity and hemolytic activity of the SMs after 24 hrs and 1 hr treatment with 200 μM, respectively. Data were normalized with a 0.1% triton-100X control. Bar: standard deviation; n = 8 replicates per group. ( B ) Galleria mellonella larva survival rate after a single treatment with 12.5 μg of SM per larva. Larva death was monitored every 12 hrs for three days after treatment. Both untreated larvae (NC) and larvae treated with 1% dimethyl sulfoxide <t>(DMSO)</t> had 100% survival (blue line). RBCs: red blood cells; n = 15 replicates per group.
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    Image Search Results


    Phenotypic analysis of zebrafish after morpholino-mediated knockdown of vcp or ML240 treatment. ( a to d ) Lateral view of embryos at 60 hpf. Zebrafish embryos were either injected with 5 base pair (bp) mismatch vcp start morpholino (MO1- control ; a ) and vcp start MO (MO1- vcp ; b ) or treated with DMSO ( c ) and VCP-Inhibitor ML240 (VCP-I; d ). scale bar: 500 μm ( a, c ); scale bar: 650 μm ( b, d ). ( e ) Statistical analysis of affected embryos after MO1/2- control or MO1/2- vcp injection and DMSO or ML240 treatment. N = 3/4 Experiments with in total n = 259 (MO1- control ), n = 161 (MO2- control ), n = 242 (MO1- vcp ), n = 212 (MO2- vcp ), n = 152 (DMSO), n = 148 (VCP-I); N two-sided Fisher exact tests combined with a modified Fisher method. Ratios of N experiments are displayed. MO1- vcp -injected embryos reveal reduced levels of Vcp by western blot analysis. Actb1/β-actin was used as loading control. RT-PCR after MO2- vcp injection shows specific effect on mRNA splicing. ( f ) Ventricular fractional shortening (FS) measurements of MO1- control and MO1- vcp -injected embryos at 48, 72 and 96 hpf, n = 18 to 30 (MO1- control ), n = 19 to 32 (MO1- vcp ). The individual measurements are shown (two-sided Wilcoxon rank-sum test). ( g ) Heart beat per minute at 48, 72 and 96 hpf of vcp morphants and control embryos. n = 30 (MO1- control ), n = 29 or 30 (MO1- vcp ). The individual measurements are shown (two-sided unpaired Wilcoxon rank-sum test. ( h and i ) False-colored superimposed overviews of spontaneous movement assay with MO1- control - ( h ) and MO1- vcp -injected ( i ) embryos at 24 hpf; red pictures = 0 s; green pictures = 10 s; yellow = merge. scale bar: 1 mm. ( j ) Quantification of spontaneous movement assay; N = 3 experiments with n = 30 (MO1- control ), n = 29 or 30 (MO1- vcp ) per experiment (N two-sided Fisher exact tests combined with modified Fisher method). The ratios of N experiments are shown. ( k ) Statistical analysis of touch-evoke flight response of control and vcp morphants at 72 hpf. n = 30, N = 3 (MO1- control ), n = 29 or 30, N = 3 (MO1- vcp ); data represent means.

    Journal: Autophagy

    Article Title: Loss of the novel Vcp (valosin containing protein) interactor Washc4 interferes with autophagy-mediated proteostasis in striated muscle and leads to myopathy in vivo

    doi: 10.1080/15548627.2018.1491491

    Figure Lengend Snippet: Phenotypic analysis of zebrafish after morpholino-mediated knockdown of vcp or ML240 treatment. ( a to d ) Lateral view of embryos at 60 hpf. Zebrafish embryos were either injected with 5 base pair (bp) mismatch vcp start morpholino (MO1- control ; a ) and vcp start MO (MO1- vcp ; b ) or treated with DMSO ( c ) and VCP-Inhibitor ML240 (VCP-I; d ). scale bar: 500 μm ( a, c ); scale bar: 650 μm ( b, d ). ( e ) Statistical analysis of affected embryos after MO1/2- control or MO1/2- vcp injection and DMSO or ML240 treatment. N = 3/4 Experiments with in total n = 259 (MO1- control ), n = 161 (MO2- control ), n = 242 (MO1- vcp ), n = 212 (MO2- vcp ), n = 152 (DMSO), n = 148 (VCP-I); N two-sided Fisher exact tests combined with a modified Fisher method. Ratios of N experiments are displayed. MO1- vcp -injected embryos reveal reduced levels of Vcp by western blot analysis. Actb1/β-actin was used as loading control. RT-PCR after MO2- vcp injection shows specific effect on mRNA splicing. ( f ) Ventricular fractional shortening (FS) measurements of MO1- control and MO1- vcp -injected embryos at 48, 72 and 96 hpf, n = 18 to 30 (MO1- control ), n = 19 to 32 (MO1- vcp ). The individual measurements are shown (two-sided Wilcoxon rank-sum test). ( g ) Heart beat per minute at 48, 72 and 96 hpf of vcp morphants and control embryos. n = 30 (MO1- control ), n = 29 or 30 (MO1- vcp ). The individual measurements are shown (two-sided unpaired Wilcoxon rank-sum test. ( h and i ) False-colored superimposed overviews of spontaneous movement assay with MO1- control - ( h ) and MO1- vcp -injected ( i ) embryos at 24 hpf; red pictures = 0 s; green pictures = 10 s; yellow = merge. scale bar: 1 mm. ( j ) Quantification of spontaneous movement assay; N = 3 experiments with n = 30 (MO1- control ), n = 29 or 30 (MO1- vcp ) per experiment (N two-sided Fisher exact tests combined with modified Fisher method). The ratios of N experiments are shown. ( k ) Statistical analysis of touch-evoke flight response of control and vcp morphants at 72 hpf. n = 30, N = 3 (MO1- control ), n = 29 or 30, N = 3 (MO1- vcp ); data represent means.

    Article Snippet: At 10 hpf wild-type zebrafish embryos were treated with 1% DMSO (Sigma-Aldrich, D8418) or 15 µM ML240 (Sigma-Aldrich, SML1071) in E3 (4 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl2 , 0.33 mM MgSO4 ) for 38 h. At 48 hpf, control zebrafish embryos or vcp and washc4 morphants were treated with DMSO or ammonium chloride (100 mM NH4 Cl; Sigma-Aldrich, A9434) in E3 for 4 h or MG132 (100 µM) (Tocris Bioscience, 1748) [ ].

    Techniques: Injection, Modification, Western Blot, Reverse Transcription Polymerase Chain Reaction, Motility Assay

    Loss of Washc4 leads to an upregulation of ER stress markers and to an impaired autophagy. ( a ) Western blot analysis of ubiquitinated proteins after MO1- control and MO1- washc4 injection. ( b ) qRT-PCR of ER stress markers ( hspa5, atf4 and ppp1r15a ) of control embryos or washc4 morphants at 3 dpf. The individual measurements are shown (log10). The 3 markers were analyzed via Wilcoxon rank-sum tests. A combined P value (comb. p) was determined via a modified Fisher method. slc25a5 was used as housekeeping gene. ( c ) Western blot analysis using anti-Sqstm1 antibody after MO1- control and MO1- washc4 injection (upper panel); quantification of gray values (lower panel; n = 6). ( d ) Representative western blotting of Lc3 of control embryos or washc4 morphants (upper panel); quantification of gray values (lower panel; n = 6). Individual measurements are shown (Wilcoxon rank-sum test). ( e ) MO1- control -or MO1- washc4 -injected embryos treated with DMSO or ammonium chloride (NH 4 Cl). Control embryos show a significant increase of Lc3-II levels after NH 4 Cl treatment in comparison to DMSO controls. Ammonium chloride treatment of Washc4 morphants did not significantly enhance the Lc3-II level in comparison to DMSO treated embryos (n = 6). Actb1/β-actin was used as loading control. The individual measurements are shown (Wilcoxon rank-sum test).

    Journal: Autophagy

    Article Title: Loss of the novel Vcp (valosin containing protein) interactor Washc4 interferes with autophagy-mediated proteostasis in striated muscle and leads to myopathy in vivo

    doi: 10.1080/15548627.2018.1491491

    Figure Lengend Snippet: Loss of Washc4 leads to an upregulation of ER stress markers and to an impaired autophagy. ( a ) Western blot analysis of ubiquitinated proteins after MO1- control and MO1- washc4 injection. ( b ) qRT-PCR of ER stress markers ( hspa5, atf4 and ppp1r15a ) of control embryos or washc4 morphants at 3 dpf. The individual measurements are shown (log10). The 3 markers were analyzed via Wilcoxon rank-sum tests. A combined P value (comb. p) was determined via a modified Fisher method. slc25a5 was used as housekeeping gene. ( c ) Western blot analysis using anti-Sqstm1 antibody after MO1- control and MO1- washc4 injection (upper panel); quantification of gray values (lower panel; n = 6). ( d ) Representative western blotting of Lc3 of control embryos or washc4 morphants (upper panel); quantification of gray values (lower panel; n = 6). Individual measurements are shown (Wilcoxon rank-sum test). ( e ) MO1- control -or MO1- washc4 -injected embryos treated with DMSO or ammonium chloride (NH 4 Cl). Control embryos show a significant increase of Lc3-II levels after NH 4 Cl treatment in comparison to DMSO controls. Ammonium chloride treatment of Washc4 morphants did not significantly enhance the Lc3-II level in comparison to DMSO treated embryos (n = 6). Actb1/β-actin was used as loading control. The individual measurements are shown (Wilcoxon rank-sum test).

    Article Snippet: At 10 hpf wild-type zebrafish embryos were treated with 1% DMSO (Sigma-Aldrich, D8418) or 15 µM ML240 (Sigma-Aldrich, SML1071) in E3 (4 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl2 , 0.33 mM MgSO4 ) for 38 h. At 48 hpf, control zebrafish embryos or vcp and washc4 morphants were treated with DMSO or ammonium chloride (100 mM NH4 Cl; Sigma-Aldrich, A9434) in E3 for 4 h or MG132 (100 µM) (Tocris Bioscience, 1748) [ ].

    Techniques: Western Blot, Injection, Quantitative RT-PCR, Modification

    Inactivation of Vcp leads to an accumulation of ubiquitinated proteins and Sqstm1. ( a ) Western blot analysis using an anti-K48-linkage specific ubiquitin antibody after MO1- control and MO1- vcp injection and DMSO or MG132 treatment (n = 4). ( b ) Vcp morphants show a significant increase of Sqstm1 by western blot analysis (n = 3); quantification of gray values. Actb1/β-actin was used as loading control. Data represent means ± SD, unpaired Student t test, * P value

    Journal: Autophagy

    Article Title: Loss of the novel Vcp (valosin containing protein) interactor Washc4 interferes with autophagy-mediated proteostasis in striated muscle and leads to myopathy in vivo

    doi: 10.1080/15548627.2018.1491491

    Figure Lengend Snippet: Inactivation of Vcp leads to an accumulation of ubiquitinated proteins and Sqstm1. ( a ) Western blot analysis using an anti-K48-linkage specific ubiquitin antibody after MO1- control and MO1- vcp injection and DMSO or MG132 treatment (n = 4). ( b ) Vcp morphants show a significant increase of Sqstm1 by western blot analysis (n = 3); quantification of gray values. Actb1/β-actin was used as loading control. Data represent means ± SD, unpaired Student t test, * P value

    Article Snippet: At 10 hpf wild-type zebrafish embryos were treated with 1% DMSO (Sigma-Aldrich, D8418) or 15 µM ML240 (Sigma-Aldrich, SML1071) in E3 (4 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl2 , 0.33 mM MgSO4 ) for 38 h. At 48 hpf, control zebrafish embryos or vcp and washc4 morphants were treated with DMSO or ammonium chloride (100 mM NH4 Cl; Sigma-Aldrich, A9434) in E3 for 4 h or MG132 (100 µM) (Tocris Bioscience, 1748) [ ].

    Techniques: Western Blot, Injection

    The effect of atosiban and nolasiban on spontaneous and OT-induced myometrial contractions. Pre-labour lower segment myometrial biopsies were subjected to stretch force of 4 g to attain spontaneous contractions. After 20 min of basal reading, vehicle control (DMSO), atosiban (Ato) or nolasiban (Nol) (6, 60, or 600 nM) was added and its effect on spontaneous contractions was measured for 10 min. The effect of the atosiban ( a ) or nolasiban ( b ) upon OT was then measured by adding increasing concentrations of the agonist (1, 10, and 100 nM) at 10 minute intervals. Total work (area under all contractions) was measured for each experimental time point and re-expressed as a ratio to the baseline period measurements (n = 6, *** p

    Journal: Scientific Reports

    Article Title: Oxytocin Receptor Antagonists, Atosiban and Nolasiban, Inhibit Prostaglandin F2α-induced Contractions and Inflammatory Responses in Human Myometrium

    doi: 10.1038/s41598-019-42181-2

    Figure Lengend Snippet: The effect of atosiban and nolasiban on spontaneous and OT-induced myometrial contractions. Pre-labour lower segment myometrial biopsies were subjected to stretch force of 4 g to attain spontaneous contractions. After 20 min of basal reading, vehicle control (DMSO), atosiban (Ato) or nolasiban (Nol) (6, 60, or 600 nM) was added and its effect on spontaneous contractions was measured for 10 min. The effect of the atosiban ( a ) or nolasiban ( b ) upon OT was then measured by adding increasing concentrations of the agonist (1, 10, and 100 nM) at 10 minute intervals. Total work (area under all contractions) was measured for each experimental time point and re-expressed as a ratio to the baseline period measurements (n = 6, *** p

    Article Snippet: Drugs and reagents Dimethyl sulfoxide (DMSO) and atosiban were purchased from Sigma-Aldrich (Dorset, UK).

    Techniques:

    The effect of atosiban and nolasiban on PGF 2α -induced myometrial contractions. Pre-labour lower segment myometrial biopsies were subjected to stretch force of 4 g to attain spontaneous contractions. After 20 min of basal reading, vehicle control (DMSO), atosiban (Ato) or nolasiban (Nol) (6, 60, or 600 nM) was added and its effect on spontaneous contractions was measured for 10 min. The effect of the atosiban ( a ) or nolasiban ( b ) upon PGF 2α was then measured by adding increasing concentrations of agonist (10, 100, and 1000 nM) at 10 minute intervals. Total work (area under all contractions) was measured for each experimental time point and re-expressed as a ratio to the baseline period measurements (n = 6, ** p

    Journal: Scientific Reports

    Article Title: Oxytocin Receptor Antagonists, Atosiban and Nolasiban, Inhibit Prostaglandin F2α-induced Contractions and Inflammatory Responses in Human Myometrium

    doi: 10.1038/s41598-019-42181-2

    Figure Lengend Snippet: The effect of atosiban and nolasiban on PGF 2α -induced myometrial contractions. Pre-labour lower segment myometrial biopsies were subjected to stretch force of 4 g to attain spontaneous contractions. After 20 min of basal reading, vehicle control (DMSO), atosiban (Ato) or nolasiban (Nol) (6, 60, or 600 nM) was added and its effect on spontaneous contractions was measured for 10 min. The effect of the atosiban ( a ) or nolasiban ( b ) upon PGF 2α was then measured by adding increasing concentrations of agonist (10, 100, and 1000 nM) at 10 minute intervals. Total work (area under all contractions) was measured for each experimental time point and re-expressed as a ratio to the baseline period measurements (n = 6, ** p

    Article Snippet: Drugs and reagents Dimethyl sulfoxide (DMSO) and atosiban were purchased from Sigma-Aldrich (Dorset, UK).

    Techniques:

    Heat map of panobinostat -induced gene expression changes in MDA-MB-231, MDA-MB-468, and MCF-7 cells . MDA-MB-231, MDA-MB-468, or MCF-7 cells were treated for 24 hours with panobinostat (100 nM) or vehicle (DMSO) and then assayed via the Human Breast Cancer and Estrogen Receptor Signaling RT 2 Profiler™ PCR Array. Red signifies up-regulation and green signifies down-regulation by panobinostat compared to MDA-MB-231 vehicle treated controls. Data are representative of three independent experiments. Genes regulated at least 2-fold are also summarized in Additional file 1 (MDA-MB-231), 2 (MDA-MB-468) and 3 (MCF-7). DMSO, dimethyl sulfoxide.

    Journal: Breast Cancer Research : BCR

    Article Title: Targeting triple-negative breast cancer cells with the histone deacetylase inhibitor panobinostat

    doi: 10.1186/bcr3192

    Figure Lengend Snippet: Heat map of panobinostat -induced gene expression changes in MDA-MB-231, MDA-MB-468, and MCF-7 cells . MDA-MB-231, MDA-MB-468, or MCF-7 cells were treated for 24 hours with panobinostat (100 nM) or vehicle (DMSO) and then assayed via the Human Breast Cancer and Estrogen Receptor Signaling RT 2 Profiler™ PCR Array. Red signifies up-regulation and green signifies down-regulation by panobinostat compared to MDA-MB-231 vehicle treated controls. Data are representative of three independent experiments. Genes regulated at least 2-fold are also summarized in Additional file 1 (MDA-MB-231), 2 (MDA-MB-468) and 3 (MCF-7). DMSO, dimethyl sulfoxide.

    Article Snippet: Panobinostat was dissolved in dimethyl sulfoxide (DMSO) (Invitrogen) as a 1 mM stock solution and kept at -20°C.

    Techniques: Expressing, Multiple Displacement Amplification, Polymerase Chain Reaction

    Panobinostat decreases TNBC cell proliferation and viability . Cells from four TNBC cell lines (MDA-MB-157, MDA-MB-231, MDA-MB-468, BT549) and three ER-positive cell lines (MCF-7, MDA-MB-361, ZR-75) were treated with panobinostat (50, 100, 200 nM) or vehicle (DMSO) for 24 hours and assayed by (A) MTT proliferation and (B) trypan blue exclusion assays. Data are represented as percent control (mean ± SEM) of three independent experiments, (**, P

    Journal: Breast Cancer Research : BCR

    Article Title: Targeting triple-negative breast cancer cells with the histone deacetylase inhibitor panobinostat

    doi: 10.1186/bcr3192

    Figure Lengend Snippet: Panobinostat decreases TNBC cell proliferation and viability . Cells from four TNBC cell lines (MDA-MB-157, MDA-MB-231, MDA-MB-468, BT549) and three ER-positive cell lines (MCF-7, MDA-MB-361, ZR-75) were treated with panobinostat (50, 100, 200 nM) or vehicle (DMSO) for 24 hours and assayed by (A) MTT proliferation and (B) trypan blue exclusion assays. Data are represented as percent control (mean ± SEM) of three independent experiments, (**, P

    Article Snippet: Panobinostat was dissolved in dimethyl sulfoxide (DMSO) (Invitrogen) as a 1 mM stock solution and kept at -20°C.

    Techniques: Multiple Displacement Amplification, MTT Assay

    Panobinostat induces apoptosis in TNBC cells . (A) TNBC cells treated with panobinostat (100, 200 nM) or vehicle (DMSO) for 24 hours were assayed by DNA fragmentation (Cell Death ELISA) assay to assess changes in apoptosis. Data are presented as enrichment (mean ± SEM) versus control of two independent experiments (***, P

    Journal: Breast Cancer Research : BCR

    Article Title: Targeting triple-negative breast cancer cells with the histone deacetylase inhibitor panobinostat

    doi: 10.1186/bcr3192

    Figure Lengend Snippet: Panobinostat induces apoptosis in TNBC cells . (A) TNBC cells treated with panobinostat (100, 200 nM) or vehicle (DMSO) for 24 hours were assayed by DNA fragmentation (Cell Death ELISA) assay to assess changes in apoptosis. Data are presented as enrichment (mean ± SEM) versus control of two independent experiments (***, P

    Article Snippet: Panobinostat was dissolved in dimethyl sulfoxide (DMSO) (Invitrogen) as a 1 mM stock solution and kept at -20°C.

    Techniques: Enzyme-linked Immunosorbent Assay

    Effect of panobinostat on tumor growth in MDA-MB-231 and BT549 xenograft models . Female, CB-17/SCID mice ( n = 5/group) were injected with (A) MDA-MB-231-tRFP or (B) BT-549-tRFP cells (5 × 10 6 cells/injection) bilaterally into the inguinal mammary fat pad. On day 14, mice were treated intraperitoneally (i.p.) with panobinostat (10 mg/kg) or vehicle (1:20 DMSO in normal saline) five days/week for 28 days. Data points represent average tumor volume ± SEM, (***, P

    Journal: Breast Cancer Research : BCR

    Article Title: Targeting triple-negative breast cancer cells with the histone deacetylase inhibitor panobinostat

    doi: 10.1186/bcr3192

    Figure Lengend Snippet: Effect of panobinostat on tumor growth in MDA-MB-231 and BT549 xenograft models . Female, CB-17/SCID mice ( n = 5/group) were injected with (A) MDA-MB-231-tRFP or (B) BT-549-tRFP cells (5 × 10 6 cells/injection) bilaterally into the inguinal mammary fat pad. On day 14, mice were treated intraperitoneally (i.p.) with panobinostat (10 mg/kg) or vehicle (1:20 DMSO in normal saline) five days/week for 28 days. Data points represent average tumor volume ± SEM, (***, P

    Article Snippet: Panobinostat was dissolved in dimethyl sulfoxide (DMSO) (Invitrogen) as a 1 mM stock solution and kept at -20°C.

    Techniques: Multiple Displacement Amplification, Mouse Assay, Injection

    Panobinostat up-regulates CDH1 expression and initiates EMT reversal in MDA-MB-231 Cells . MDA-MB-231 cells were plated overnight and treated with panobinostat (100 nM) or vehicle (DMSO) for 24 hours. The expression of CDH1 was examined by (A) flow cytometry and (B) ELISA. Data are represented as mean ± SEM of two independent experiments, (***, P

    Journal: Breast Cancer Research : BCR

    Article Title: Targeting triple-negative breast cancer cells with the histone deacetylase inhibitor panobinostat

    doi: 10.1186/bcr3192

    Figure Lengend Snippet: Panobinostat up-regulates CDH1 expression and initiates EMT reversal in MDA-MB-231 Cells . MDA-MB-231 cells were plated overnight and treated with panobinostat (100 nM) or vehicle (DMSO) for 24 hours. The expression of CDH1 was examined by (A) flow cytometry and (B) ELISA. Data are represented as mean ± SEM of two independent experiments, (***, P

    Article Snippet: Panobinostat was dissolved in dimethyl sulfoxide (DMSO) (Invitrogen) as a 1 mM stock solution and kept at -20°C.

    Techniques: Expressing, Multiple Displacement Amplification, Flow Cytometry, Cytometry, Enzyme-linked Immunosorbent Assay

    Panobinostat increases histone H3 (Lys9) and H4 (Lys8) acetylation in TNBC cell lines . Cells were treated with panobinostat (100, 200 nM) or vehicle (DMSO) for 18 hours, fixed, permeabilized and stained for acetyl-histones (A) H3 (Lys9) or (B) H4 (Lys8) and subjected to flow cytometry. Data are presented as mean fluorescence intensity (mean ± SEM) of two independent experiments, (***, P

    Journal: Breast Cancer Research : BCR

    Article Title: Targeting triple-negative breast cancer cells with the histone deacetylase inhibitor panobinostat

    doi: 10.1186/bcr3192

    Figure Lengend Snippet: Panobinostat increases histone H3 (Lys9) and H4 (Lys8) acetylation in TNBC cell lines . Cells were treated with panobinostat (100, 200 nM) or vehicle (DMSO) for 18 hours, fixed, permeabilized and stained for acetyl-histones (A) H3 (Lys9) or (B) H4 (Lys8) and subjected to flow cytometry. Data are presented as mean fluorescence intensity (mean ± SEM) of two independent experiments, (***, P

    Article Snippet: Panobinostat was dissolved in dimethyl sulfoxide (DMSO) (Invitrogen) as a 1 mM stock solution and kept at -20°C.

    Techniques: Staining, Flow Cytometry, Cytometry, Fluorescence

    In vivo activity of EN4 in a mouse tissue cage model. The numbers of planktonic SA113 (A) and the viability of leukocytes (B) present in tissue cage fluid from cages treated with 100 or 250 μg of EN4 or 30 μg of daptomycin (DAP) and infected with 4 × 10 3 CFU of SA113 were investigated. Open circles indicate control mice injected with 1% DMSO-PBS. The values shown are means ± the SDs.

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Antimicrobial Properties of 8-Hydroxyserrulat-14-en-19-oic Acid for Treatment of Implant-Associated Infections

    doi: 10.1128/AAC.01735-12

    Figure Lengend Snippet: In vivo activity of EN4 in a mouse tissue cage model. The numbers of planktonic SA113 (A) and the viability of leukocytes (B) present in tissue cage fluid from cages treated with 100 or 250 μg of EN4 or 30 μg of daptomycin (DAP) and infected with 4 × 10 3 CFU of SA113 were investigated. Open circles indicate control mice injected with 1% DMSO-PBS. The values shown are means ± the SDs.

    Article Snippet: EN4 was dissolved in 1% dimethyl sulfoxide (DMSO) (Merck, Darmstadt, Germany) in phosphate-buffered saline (PBS) (Reagens, Basel, Switzerland) (DMSO-PBS) up to a concentration of 400 μg/ml.

    Techniques: In Vivo, Activity Assay, Infection, Mouse Assay, Injection

    Effect of EN4 on bacterial membrane integrity. (A) S. epidermidis 1457 was incubated for 10 min with EN4 or CHX at 100 μg/ml or 1% DMSO-PBS and subsequently double stained with propidium iodide (PI) and SYTO9 and analyzed by flow cytometry. The results of one representative experiment of three performed are shown. (B) Transmission electron microscopy images of ultrathin sections of WSPPA treated for 1 h with 1% DMSO-PBS as a control, EN4 at 100 or 200 μg/ml, or 250 μg of CHX/ml. Arrowheads indicate ultrastructural changes; the bars represent 200 nm. (C) WSPPA was treated for 10 min with EN4 or CHX at 100 μg/ml, followed by centrifugation and investigation of supernatants for the presence of ATP using luciferase reaction. Nisin (NIS) and ciprofloxacin (CIP) were used as a positive and a negative control, respectively, and 1% DMSO-PBS was used as an untreated control (Ctrl). The values shown are means ± the SDs of areas under the concentration-time curve calculated for the first 30 min of luciferase reaction from at least three independent experiments prepared in duplicates. Significant ATP leakage compared to results for the untreated control is indicated (*, P

    Journal: Antimicrobial Agents and Chemotherapy

    Article Title: Antimicrobial Properties of 8-Hydroxyserrulat-14-en-19-oic Acid for Treatment of Implant-Associated Infections

    doi: 10.1128/AAC.01735-12

    Figure Lengend Snippet: Effect of EN4 on bacterial membrane integrity. (A) S. epidermidis 1457 was incubated for 10 min with EN4 or CHX at 100 μg/ml or 1% DMSO-PBS and subsequently double stained with propidium iodide (PI) and SYTO9 and analyzed by flow cytometry. The results of one representative experiment of three performed are shown. (B) Transmission electron microscopy images of ultrathin sections of WSPPA treated for 1 h with 1% DMSO-PBS as a control, EN4 at 100 or 200 μg/ml, or 250 μg of CHX/ml. Arrowheads indicate ultrastructural changes; the bars represent 200 nm. (C) WSPPA was treated for 10 min with EN4 or CHX at 100 μg/ml, followed by centrifugation and investigation of supernatants for the presence of ATP using luciferase reaction. Nisin (NIS) and ciprofloxacin (CIP) were used as a positive and a negative control, respectively, and 1% DMSO-PBS was used as an untreated control (Ctrl). The values shown are means ± the SDs of areas under the concentration-time curve calculated for the first 30 min of luciferase reaction from at least three independent experiments prepared in duplicates. Significant ATP leakage compared to results for the untreated control is indicated (*, P

    Article Snippet: EN4 was dissolved in 1% dimethyl sulfoxide (DMSO) (Merck, Darmstadt, Germany) in phosphate-buffered saline (PBS) (Reagens, Basel, Switzerland) (DMSO-PBS) up to a concentration of 400 μg/ml.

    Techniques: Incubation, Staining, Flow Cytometry, Cytometry, Transmission Assay, Electron Microscopy, Centrifugation, Luciferase, Negative Control, Concentration Assay

    Analogs of honokiol do not bind or inhibit FOXM1 a C3-luc cells were induced with doxycycline and treated with the indicated analogs of honokiol at the concentration of 40 µM for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b DU145 prostate cancer cells were treated as indicated. Following treatment cells were collected and immunoblotting was performed for FOXM1, cleaved caspase-3 and β-actin as the loading control. c STD NMR spectra of 2 mM of: (I) 2 mM eugenol, (II) 2 mM O -eugenol, (III) 2 mM 2-allylphenol and (IV) 2 mM 2,2 dihydroxybiphenyl. The chemical structure of each small molecule is illustrated. Signals from vehicle (DMSO) and water are labeled. d STD NMR spectra of 150 ng of recombinant FOXM1 plus: (I) 2 mM eugenol, (II) 2 mM O-eugenol, (III) 2 mM 2,2 dihydroxybiphenyl and (IV) 2 mM 2-allylphenol. The chemical structure of each small molecule is illustrated. STD signals arising from the aryl groups are annotated and signals from vehicle (DMSO) and water are labeled

    Journal: Cell Death & Disease

    Article Title: Honokiol is a FOXM1 antagonist

    doi: 10.1038/s41419-017-0156-7

    Figure Lengend Snippet: Analogs of honokiol do not bind or inhibit FOXM1 a C3-luc cells were induced with doxycycline and treated with the indicated analogs of honokiol at the concentration of 40 µM for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b DU145 prostate cancer cells were treated as indicated. Following treatment cells were collected and immunoblotting was performed for FOXM1, cleaved caspase-3 and β-actin as the loading control. c STD NMR spectra of 2 mM of: (I) 2 mM eugenol, (II) 2 mM O -eugenol, (III) 2 mM 2-allylphenol and (IV) 2 mM 2,2 dihydroxybiphenyl. The chemical structure of each small molecule is illustrated. Signals from vehicle (DMSO) and water are labeled. d STD NMR spectra of 150 ng of recombinant FOXM1 plus: (I) 2 mM eugenol, (II) 2 mM O-eugenol, (III) 2 mM 2,2 dihydroxybiphenyl and (IV) 2 mM 2-allylphenol. The chemical structure of each small molecule is illustrated. STD signals arising from the aryl groups are annotated and signals from vehicle (DMSO) and water are labeled

    Article Snippet: MG132 (EMD Millipore), thiostrepton (Sigma), honokiol, 2,2 dihydroxybiphenyl, 2 allylphenol, eugenol, and O -eugenol were dissolved in dimethyl sulfoxide (DMSO) (Fisher Scientific), N-acetyl-l-cysteine (NAC) (Sigma) in deionized water, and doxycycline (LKT Laboratories) in phosphate buffered saline (PBS).

    Techniques: Concentration Assay, Luciferase, Activity Assay, Nuclear Magnetic Resonance, Labeling, Recombinant

    Honokiol inhibits FOXM1 transactivation via binding a C3-luc cells were induced with doxycycline and treated with honokiol for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b The C3 cell line was treated with doxycycline and honokiol in the indicated concentrations for 24 h. Cells were collected and immunoblotting was performed with a FOXM1 specific antibody. β-actin was used as the loading control. c Representative EMSA image shows the inhibitory effect of honokiol on the formation of the FOXM1 DBD protein–DNA complex. d The C3 cell line was treated with doxycycline and honokiol as indicated for 24 h. Then, cells were processed for the ChIP experiments, as described in “Materials and methods”. Graph shows mean ± SEM of two independent ChIP experiments. e Saturation transfer difference (STD) NMR spectra to assess the binding of honokiol to FOXM1: (I) 2 mM of honokiol alone, (II) 150 ng of recombinant FOXM1 alone, (III) 2 mM honokiol with 150 ng of recombinant FOXM1. The chemical structure of honokiol is illustrated. STD signals arising from the aryl groups in honokiol are annotated, and signals from vehicle (DMSO) and water are labeled

    Journal: Cell Death & Disease

    Article Title: Honokiol is a FOXM1 antagonist

    doi: 10.1038/s41419-017-0156-7

    Figure Lengend Snippet: Honokiol inhibits FOXM1 transactivation via binding a C3-luc cells were induced with doxycycline and treated with honokiol for 24 h. The luciferase activity was determined by using the Luciferase Assay System (Promega). Graph shows quantification as fold induction of firefly luciferase activity compared to control cells, mean ± SD of a representative triplicate experiment. b The C3 cell line was treated with doxycycline and honokiol in the indicated concentrations for 24 h. Cells were collected and immunoblotting was performed with a FOXM1 specific antibody. β-actin was used as the loading control. c Representative EMSA image shows the inhibitory effect of honokiol on the formation of the FOXM1 DBD protein–DNA complex. d The C3 cell line was treated with doxycycline and honokiol as indicated for 24 h. Then, cells were processed for the ChIP experiments, as described in “Materials and methods”. Graph shows mean ± SEM of two independent ChIP experiments. e Saturation transfer difference (STD) NMR spectra to assess the binding of honokiol to FOXM1: (I) 2 mM of honokiol alone, (II) 150 ng of recombinant FOXM1 alone, (III) 2 mM honokiol with 150 ng of recombinant FOXM1. The chemical structure of honokiol is illustrated. STD signals arising from the aryl groups in honokiol are annotated, and signals from vehicle (DMSO) and water are labeled

    Article Snippet: MG132 (EMD Millipore), thiostrepton (Sigma), honokiol, 2,2 dihydroxybiphenyl, 2 allylphenol, eugenol, and O -eugenol were dissolved in dimethyl sulfoxide (DMSO) (Fisher Scientific), N-acetyl-l-cysteine (NAC) (Sigma) in deionized water, and doxycycline (LKT Laboratories) in phosphate buffered saline (PBS).

    Techniques: Binding Assay, Luciferase, Activity Assay, Chromatin Immunoprecipitation, Nuclear Magnetic Resonance, Recombinant, Labeling

    Supervised cluster analysis and GSEA between PD-L1 -high and -low human lung cancer cell lines suggested mRNA upregulation of several molecules related to cell adhesion, motility, cell proliferation, cell signaling, and pathways related to actin cytoskeleton regulation and integrin-mediated cell adhesion. A, Supervised cluster analysis of 10 PD-L1 -high versus 10 PD-L1 -low human lung cancer cell lines. Depicted by the CCLE Analysis Tools: Differential Expression ( http://www.broadinstitute.org/ccle/data/browseAnalyses ). B, qRT-PCR of 11 selected mRNAs in PD-L1 -high and -low cell lines. The ΔΔCt method was used for the target mRNA, which was normalized by endogenous β-actin mRNA and reference BEAS-2B cells. RQ data are the mean of 10 cell lines. C, RAC2 and CDA mRNA levels were compared between DMSO and U0126 treatments in the three KRAS -mutant lung adenocarcinoma cell lines. The target mRNAs were normalized with endogenous β-actin mRNAs and DMSO-treated cells as a reference (ΔΔCt method). Data are the mean of three independent experiments. D, GSEA showed significant results for pathways related to actin cytoskeleton regulation by Rho GTPases and to integrin-mediated cell adhesion.

    Journal: PLoS ONE

    Article Title: RAS–Mitogen-Activated Protein Kinase Signal Is Required for Enhanced PD-L1 Expression in Human Lung Cancers

    doi: 10.1371/journal.pone.0166626

    Figure Lengend Snippet: Supervised cluster analysis and GSEA between PD-L1 -high and -low human lung cancer cell lines suggested mRNA upregulation of several molecules related to cell adhesion, motility, cell proliferation, cell signaling, and pathways related to actin cytoskeleton regulation and integrin-mediated cell adhesion. A, Supervised cluster analysis of 10 PD-L1 -high versus 10 PD-L1 -low human lung cancer cell lines. Depicted by the CCLE Analysis Tools: Differential Expression ( http://www.broadinstitute.org/ccle/data/browseAnalyses ). B, qRT-PCR of 11 selected mRNAs in PD-L1 -high and -low cell lines. The ΔΔCt method was used for the target mRNA, which was normalized by endogenous β-actin mRNA and reference BEAS-2B cells. RQ data are the mean of 10 cell lines. C, RAC2 and CDA mRNA levels were compared between DMSO and U0126 treatments in the three KRAS -mutant lung adenocarcinoma cell lines. The target mRNAs were normalized with endogenous β-actin mRNAs and DMSO-treated cells as a reference (ΔΔCt method). Data are the mean of three independent experiments. D, GSEA showed significant results for pathways related to actin cytoskeleton regulation by Rho GTPases and to integrin-mediated cell adhesion.

    Article Snippet: Inhibitor and RNA interference (RNAi) experiments For in vitro inhibitor assays, U0126 or LY294002 (Cell Signaling Technology; CST) dissolved in dimethyl sulfoxide (DMSO) (Wako) was added to the cells at a final concentration of 20 or 40 μM, respectively, as well as to the same amount of DMSO as a control, and incubated for 24 h. For RNAi experiments, cells plated at 50–60% confluence in a 6-cm dish were transfected with siRNAs specific for ERK2 mRNA (#1, 5′-GACACAACACCUCAGCAAU-3′ and #2, 5′-CUAACGUUCUGCACCGUGA-3′ ), KRAS mRNA (#1, 5′-CUCAGGACUUAGCAAGAAGUU-3′ and #2, 5′-CAGUUGAGACCUUCUAAUUGG-3′ ), or STAT3 mRNA (#1, 5′-GCCUCAAGAUUGACCUAGA-3′ and #4, 5′-AUAGGAAGGUUUAAGGAGA-3′ ) as well as a control siRNA (siCtrl) (directed against firefly luciferase mRNA) ( 5′-GUGCGCUGCUGGUGCCAAC-3′ ) (all obtained from Sigma) at 100 nM with Lipofectamine 2000 (Invitrogen).

    Techniques: Expressing, Quantitative RT-PCR, Mutagenesis

    MEK inhibition significantly decreased PD-L1 expression in human lung cancer cell lines. A, Gating strategy for the determination of PD-L1 levels in lung cancer cell lines stained with FITC anti-PD-L1 mAb. For differentiation between live and dead cells, PI exclusion staining was used. B, (upper) Representative histogram of PD-L1 (shaded histogram) of three KRAS -mutant lung adenocarcinoma cell lines with DMSO or U0126 (20 μM) for 24 h. Empty histogram indicates the isotype control. (upper middle) Relative MFI (PD-L1 MFI / isotype control MFI) of the three cell lines was significantly decreased with U0126. *, p

    Journal: PLoS ONE

    Article Title: RAS–Mitogen-Activated Protein Kinase Signal Is Required for Enhanced PD-L1 Expression in Human Lung Cancers

    doi: 10.1371/journal.pone.0166626

    Figure Lengend Snippet: MEK inhibition significantly decreased PD-L1 expression in human lung cancer cell lines. A, Gating strategy for the determination of PD-L1 levels in lung cancer cell lines stained with FITC anti-PD-L1 mAb. For differentiation between live and dead cells, PI exclusion staining was used. B, (upper) Representative histogram of PD-L1 (shaded histogram) of three KRAS -mutant lung adenocarcinoma cell lines with DMSO or U0126 (20 μM) for 24 h. Empty histogram indicates the isotype control. (upper middle) Relative MFI (PD-L1 MFI / isotype control MFI) of the three cell lines was significantly decreased with U0126. *, p

    Article Snippet: Inhibitor and RNA interference (RNAi) experiments For in vitro inhibitor assays, U0126 or LY294002 (Cell Signaling Technology; CST) dissolved in dimethyl sulfoxide (DMSO) (Wako) was added to the cells at a final concentration of 20 or 40 μM, respectively, as well as to the same amount of DMSO as a control, and incubated for 24 h. For RNAi experiments, cells plated at 50–60% confluence in a 6-cm dish were transfected with siRNAs specific for ERK2 mRNA (#1, 5′-GACACAACACCUCAGCAAU-3′ and #2, 5′-CUAACGUUCUGCACCGUGA-3′ ), KRAS mRNA (#1, 5′-CUCAGGACUUAGCAAGAAGUU-3′ and #2, 5′-CAGUUGAGACCUUCUAAUUGG-3′ ), or STAT3 mRNA (#1, 5′-GCCUCAAGAUUGACCUAGA-3′ and #4, 5′-AUAGGAAGGUUUAAGGAGA-3′ ) as well as a control siRNA (siCtrl) (directed against firefly luciferase mRNA) ( 5′-GUGCGCUGCUGGUGCCAAC-3′ ) (all obtained from Sigma) at 100 nM with Lipofectamine 2000 (Invitrogen).

    Techniques: Inhibition, Expressing, Staining, Mutagenesis

    Regulation of PD-L1 by MAPK signaling is supported by PMA stimulation, ERK2 RNAi, and KRAS RNAi. A, Three KRAS -mutant lung adenocarcinoma cell lines were serum-starved for 48 h, pre-conditioned with DMSO or U0126 (20μM) for 1hr, then stimulated with 100 nM PMA for 15 min, and then evaluated for phospho-ERK and PD-L1 mRNA levels. Immunoblot (upper) and qRT-PCR (lower) showed increases in phospho-ERK and PD-L1 mRNA levels 15 min after PMA stimulation, respectively. *, p

    Journal: PLoS ONE

    Article Title: RAS–Mitogen-Activated Protein Kinase Signal Is Required for Enhanced PD-L1 Expression in Human Lung Cancers

    doi: 10.1371/journal.pone.0166626

    Figure Lengend Snippet: Regulation of PD-L1 by MAPK signaling is supported by PMA stimulation, ERK2 RNAi, and KRAS RNAi. A, Three KRAS -mutant lung adenocarcinoma cell lines were serum-starved for 48 h, pre-conditioned with DMSO or U0126 (20μM) for 1hr, then stimulated with 100 nM PMA for 15 min, and then evaluated for phospho-ERK and PD-L1 mRNA levels. Immunoblot (upper) and qRT-PCR (lower) showed increases in phospho-ERK and PD-L1 mRNA levels 15 min after PMA stimulation, respectively. *, p

    Article Snippet: Inhibitor and RNA interference (RNAi) experiments For in vitro inhibitor assays, U0126 or LY294002 (Cell Signaling Technology; CST) dissolved in dimethyl sulfoxide (DMSO) (Wako) was added to the cells at a final concentration of 20 or 40 μM, respectively, as well as to the same amount of DMSO as a control, and incubated for 24 h. For RNAi experiments, cells plated at 50–60% confluence in a 6-cm dish were transfected with siRNAs specific for ERK2 mRNA (#1, 5′-GACACAACACCUCAGCAAU-3′ and #2, 5′-CUAACGUUCUGCACCGUGA-3′ ), KRAS mRNA (#1, 5′-CUCAGGACUUAGCAAGAAGUU-3′ and #2, 5′-CAGUUGAGACCUUCUAAUUGG-3′ ), or STAT3 mRNA (#1, 5′-GCCUCAAGAUUGACCUAGA-3′ and #4, 5′-AUAGGAAGGUUUAAGGAGA-3′ ) as well as a control siRNA (siCtrl) (directed against firefly luciferase mRNA) ( 5′-GUGCGCUGCUGGUGCCAAC-3′ ) (all obtained from Sigma) at 100 nM with Lipofectamine 2000 (Invitrogen).

    Techniques: Mutagenesis, Quantitative RT-PCR

    STAT3 signal, but not PI3K signal, partially contributes to ectopic PD-L1 expression in human lung cancer cell lines. A, STAT3 RNAi reduced PD-L1 expression at both protein (upper) and mRNA (lower) levels in NCI-H1373 and NCI-H441 cells but not in NCI-H358 cells. The experimental conditions were similar to those in Fig 3B . Data are the mean of three independent experiments. B, Immunoblot (upper) and qRT-PCR (lower) of STAT3 showed the decreases in STAT3 protein and mRNA levels, respectively. One representative immunoblot and the mean of three independent qRT-PCR experiments are shown. C, LY294002, a PI3K inhibitor, did not suppress PD-L1 protein expression. Three KRAS -mutant lung adenocarcinoma cell lines were treated with DMSO or LY294002 (40 μM) for 24 h. PD-L1 protein levels did not show any significant changes with LY294002 with regard to both the surface (upper) and total protein (lower) levels. Data are the mean of three independent experiments.

    Journal: PLoS ONE

    Article Title: RAS–Mitogen-Activated Protein Kinase Signal Is Required for Enhanced PD-L1 Expression in Human Lung Cancers

    doi: 10.1371/journal.pone.0166626

    Figure Lengend Snippet: STAT3 signal, but not PI3K signal, partially contributes to ectopic PD-L1 expression in human lung cancer cell lines. A, STAT3 RNAi reduced PD-L1 expression at both protein (upper) and mRNA (lower) levels in NCI-H1373 and NCI-H441 cells but not in NCI-H358 cells. The experimental conditions were similar to those in Fig 3B . Data are the mean of three independent experiments. B, Immunoblot (upper) and qRT-PCR (lower) of STAT3 showed the decreases in STAT3 protein and mRNA levels, respectively. One representative immunoblot and the mean of three independent qRT-PCR experiments are shown. C, LY294002, a PI3K inhibitor, did not suppress PD-L1 protein expression. Three KRAS -mutant lung adenocarcinoma cell lines were treated with DMSO or LY294002 (40 μM) for 24 h. PD-L1 protein levels did not show any significant changes with LY294002 with regard to both the surface (upper) and total protein (lower) levels. Data are the mean of three independent experiments.

    Article Snippet: Inhibitor and RNA interference (RNAi) experiments For in vitro inhibitor assays, U0126 or LY294002 (Cell Signaling Technology; CST) dissolved in dimethyl sulfoxide (DMSO) (Wako) was added to the cells at a final concentration of 20 or 40 μM, respectively, as well as to the same amount of DMSO as a control, and incubated for 24 h. For RNAi experiments, cells plated at 50–60% confluence in a 6-cm dish were transfected with siRNAs specific for ERK2 mRNA (#1, 5′-GACACAACACCUCAGCAAU-3′ and #2, 5′-CUAACGUUCUGCACCGUGA-3′ ), KRAS mRNA (#1, 5′-CUCAGGACUUAGCAAGAAGUU-3′ and #2, 5′-CAGUUGAGACCUUCUAAUUGG-3′ ), or STAT3 mRNA (#1, 5′-GCCUCAAGAUUGACCUAGA-3′ and #4, 5′-AUAGGAAGGUUUAAGGAGA-3′ ) as well as a control siRNA (siCtrl) (directed against firefly luciferase mRNA) ( 5′-GUGCGCUGCUGGUGCCAAC-3′ ) (all obtained from Sigma) at 100 nM with Lipofectamine 2000 (Invitrogen).

    Techniques: Expressing, Quantitative RT-PCR, Mutagenesis

    Promoter assay experiments of PD-L1 . A, Diagrammatic representation of the PD-L1 regulatory elements, including PD-L1 promoter fragment and the predicted enhancer containing an AP-1 binding site. Fragments cloned into luciferase constructs are annotated below, with positions relative to the PD-L1 TSS. B, Diagram of luciferase vectors. The empty pGL4.17 vector, the GL4.17 vector with the promoter cloned upstream of the luciferase gene alone (PDL-P), the promoter with the enhancer cloned downstream of the luciferase gene (PDL-P+E). C, PD-L1 promoter- and enhancer-driven luciferase activity in NCI-H1373 with or without MEK inhibition. Firefly luciferase activity normalized by Renilla luciferase activity significantly increased with PDL-P+E compared with that with PDL-P after DMSO treatment (left), while no significant difference was found between PDL-P and PDL-P+E after U0126 treatment (right). Data are representative of three independent experiments with the same results and shown as the mean of quadruplicate experiments. Error bars indicate SD. E, Binding of cJUN to the candidate PD-L1 enhancer region following ChIP-coupled qPCR. Data are representative of three independent experiments with similar results.

    Journal: PLoS ONE

    Article Title: RAS–Mitogen-Activated Protein Kinase Signal Is Required for Enhanced PD-L1 Expression in Human Lung Cancers

    doi: 10.1371/journal.pone.0166626

    Figure Lengend Snippet: Promoter assay experiments of PD-L1 . A, Diagrammatic representation of the PD-L1 regulatory elements, including PD-L1 promoter fragment and the predicted enhancer containing an AP-1 binding site. Fragments cloned into luciferase constructs are annotated below, with positions relative to the PD-L1 TSS. B, Diagram of luciferase vectors. The empty pGL4.17 vector, the GL4.17 vector with the promoter cloned upstream of the luciferase gene alone (PDL-P), the promoter with the enhancer cloned downstream of the luciferase gene (PDL-P+E). C, PD-L1 promoter- and enhancer-driven luciferase activity in NCI-H1373 with or without MEK inhibition. Firefly luciferase activity normalized by Renilla luciferase activity significantly increased with PDL-P+E compared with that with PDL-P after DMSO treatment (left), while no significant difference was found between PDL-P and PDL-P+E after U0126 treatment (right). Data are representative of three independent experiments with the same results and shown as the mean of quadruplicate experiments. Error bars indicate SD. E, Binding of cJUN to the candidate PD-L1 enhancer region following ChIP-coupled qPCR. Data are representative of three independent experiments with similar results.

    Article Snippet: Inhibitor and RNA interference (RNAi) experiments For in vitro inhibitor assays, U0126 or LY294002 (Cell Signaling Technology; CST) dissolved in dimethyl sulfoxide (DMSO) (Wako) was added to the cells at a final concentration of 20 or 40 μM, respectively, as well as to the same amount of DMSO as a control, and incubated for 24 h. For RNAi experiments, cells plated at 50–60% confluence in a 6-cm dish were transfected with siRNAs specific for ERK2 mRNA (#1, 5′-GACACAACACCUCAGCAAU-3′ and #2, 5′-CUAACGUUCUGCACCGUGA-3′ ), KRAS mRNA (#1, 5′-CUCAGGACUUAGCAAGAAGUU-3′ and #2, 5′-CAGUUGAGACCUUCUAAUUGG-3′ ), or STAT3 mRNA (#1, 5′-GCCUCAAGAUUGACCUAGA-3′ and #4, 5′-AUAGGAAGGUUUAAGGAGA-3′ ) as well as a control siRNA (siCtrl) (directed against firefly luciferase mRNA) ( 5′-GUGCGCUGCUGGUGCCAAC-3′ ) (all obtained from Sigma) at 100 nM with Lipofectamine 2000 (Invitrogen).

    Techniques: Promoter Assay, Binding Assay, Clone Assay, Luciferase, Construct, Plasmid Preparation, Activity Assay, Inhibition, Chromatin Immunoprecipitation, Real-time Polymerase Chain Reaction

    The combination of LCRF‐0006 and bortezomib synergistically increases MM tumor response in vivo. A, Schematic representation of treatment regimen. C57BL/KaLwRij mice with established MM (day 14 post‐5TGM1 cell injection) were treated with LCRF‐0006 (100 mg/kg) and bortezomib (0.5 mg/kg) (LCRF‐0006 + bortezomib), LCRF‐0006 and DMSO vehicle (LCRF‐0006), 2‐HP‐b‐CD vehicle and bortezomib (bortezomib) or 2‐HP‐b‐CD and DMSO vehicles (vehicles) i.p., as per treatment regimen. B, Tumor burden was assessed at day 14, 21 and 28 by BLI. Graph depicts mean ± SEM. n = 5‐6 mice/treatment group. *** P

    Journal: FASEB BioAdvances

    Article Title: LCRF‐0006, a small molecule mimetic of the N‐cadherin antagonist peptide ADH‐1, synergistically increases multiple myeloma response to bortezomib, et al. LCRF‐0006, a small molecule mimetic of the N‐cadherin antagonist peptide ADH‐1, synergistically increases multiple myeloma response to bortezomib

    doi: 10.1096/fba.2019-00073

    Figure Lengend Snippet: The combination of LCRF‐0006 and bortezomib synergistically increases MM tumor response in vivo. A, Schematic representation of treatment regimen. C57BL/KaLwRij mice with established MM (day 14 post‐5TGM1 cell injection) were treated with LCRF‐0006 (100 mg/kg) and bortezomib (0.5 mg/kg) (LCRF‐0006 + bortezomib), LCRF‐0006 and DMSO vehicle (LCRF‐0006), 2‐HP‐b‐CD vehicle and bortezomib (bortezomib) or 2‐HP‐b‐CD and DMSO vehicles (vehicles) i.p., as per treatment regimen. B, Tumor burden was assessed at day 14, 21 and 28 by BLI. Graph depicts mean ± SEM. n = 5‐6 mice/treatment group. *** P

    Article Snippet: 2.2 Drugs For in vitro experiments, LCRF‐0006 (kindly provided by Crocus Laboratories) was solubilized in dimethyl sulfoxide (DMSO) (Merck).

    Techniques: In Vivo, Mouse Assay, Injection

    An mTOR inhibitor AZD8055 preferentially inhibited proliferation and phosphorylation levels in NCI N87 T-R cells (A) T-S and T-R cells were cultured for 24 h, and then treated with or without AZD8055 at the indicated concentrations (0, 25, 50, 100, 200, 400 and 800 nM) or DMSO as control, respectively. Cell viability was determined by Cell Count kit-8 assay 48 h later. Results expressed as % control represent the mean of three experiments. * p

    Journal: Oncotarget

    Article Title: Quantitative proteomics profiling reveals activation of mTOR pathway in trastuzumab resistance

    doi: 10.18632/oncotarget.17415

    Figure Lengend Snippet: An mTOR inhibitor AZD8055 preferentially inhibited proliferation and phosphorylation levels in NCI N87 T-R cells (A) T-S and T-R cells were cultured for 24 h, and then treated with or without AZD8055 at the indicated concentrations (0, 25, 50, 100, 200, 400 and 800 nM) or DMSO as control, respectively. Cell viability was determined by Cell Count kit-8 assay 48 h later. Results expressed as % control represent the mean of three experiments. * p

    Article Snippet: Chemicals used are as follows: AZD8055 (Medchem express, USA), dimethyl sulfoxide (DMSO) (Amresco, USA), Dithiothreitol (DTT) and iodoacetamide (IAA) (Sigma-Aldrich, USA), DAPI (Beyotime Biotechnology, China).

    Techniques: Cell Culture, Cell Counting

    The UV spectra of PBDE hapten, protein and conjugates: ( a ) with BSA and ( b ) with OVA; absorbance value at the characteristic peak, 292 nm: OD BSA-PBDE = 1.860, OD PBDE-hapten = 1.893, OD BSA = 0.074; 272 nm: OD OVA-PBDE = 1.767, OD PBDE - hapten = 1.893, OD OVA = 0.401; C BSA : 0.20 g/L, C OVA : 0.20 g/L, and C hapten : 0.05 g/L; protein and conjugate were dissolved in PBS buffer; hapten was dissolved in DMSO.

    Journal: Scientific Reports

    Article Title: Ultrasensitive Nano-rt-iPCR for Determination of Polybrominated Diphenyl Ethers in Natural Samples

    doi: 10.1038/s41598-017-12339-x

    Figure Lengend Snippet: The UV spectra of PBDE hapten, protein and conjugates: ( a ) with BSA and ( b ) with OVA; absorbance value at the characteristic peak, 292 nm: OD BSA-PBDE = 1.860, OD PBDE-hapten = 1.893, OD BSA = 0.074; 272 nm: OD OVA-PBDE = 1.767, OD PBDE - hapten = 1.893, OD OVA = 0.401; C BSA : 0.20 g/L, C OVA : 0.20 g/L, and C hapten : 0.05 g/L; protein and conjugate were dissolved in PBS buffer; hapten was dissolved in DMSO.

    Article Snippet: Complete Freund’s adjuvant, ovalbumin (OVA), Bovine serum albumin (BSA), dichloromethane, n-hexane, Tween-20, N-hydroxysuccinimide (NHS), 1-(3-Dimethylaminopropyl)-3-ethyl carbodiimide hydrochloride(EDC), dimethyl sulfoxide (DMSO), O-(carboxymethyl)hydroxylamine hemihydrochloride, methanol, Coomassie Brilliant Blue G250 and anhydrous sodium acetate were purchased from Sinopharm, China.

    Techniques:

    Controlling TC-83 induced ROS response in microglial cells decreases inflammatory burden. (a) Experimental workflow indicating HMC3 cells were pre-treated for 2h with 0.1% DMSO, 1μM BAY 11–7082 (BAY-82), or 100pM mitoquinone mesylate (MitoQ) prior to inoculation with TC-83 (MOI:2). Levels of the pro-inflammatory cytokines (b) IL-1α, (c) IL-1β, (d), IL-6 and (e) IL-8 produced by HMC3 cells as the direct result of TC-83 infection were measured at 1, 2, and 6hpi. The quantitative data are depicted as the means of three biologically independent experiments ± SD. * p

    Journal: Virulence

    Article Title: Direct and indirect pro-inflammatory cytokine response resulting from TC-83 infection of glial cells

    doi: 10.1080/21505594.2018.1509668

    Figure Lengend Snippet: Controlling TC-83 induced ROS response in microglial cells decreases inflammatory burden. (a) Experimental workflow indicating HMC3 cells were pre-treated for 2h with 0.1% DMSO, 1μM BAY 11–7082 (BAY-82), or 100pM mitoquinone mesylate (MitoQ) prior to inoculation with TC-83 (MOI:2). Levels of the pro-inflammatory cytokines (b) IL-1α, (c) IL-1β, (d), IL-6 and (e) IL-8 produced by HMC3 cells as the direct result of TC-83 infection were measured at 1, 2, and 6hpi. The quantitative data are depicted as the means of three biologically independent experiments ± SD. * p

    Article Snippet: Antibodies, inhibitors, and reagents The reagents and antibodies (Abs) used were as follows: dimethyl sulfoxide (DMSO) (VWR 97,063–136); the mitochondrially targeted antioxidant mitoquinone mesylate (MedKoo Biosciences 317,102); the NF-κB inhibitor BAY 11–7082 (Selleckchem S2913); the IL-1β inhibitor Pirfenidone (SelleckChem S2907); IL-1α, IL-1β, IL-6, and IL-8 recombinant protein (Aushon 101-3FF-1-AB); mouse monoclonal Ab to TOMM20 (Abcam ab56783), Alexa Fluor 488-conjugated goat anti-mouse (Invitrogen A21202), Alexa Fluor 568-conjugated donkey anti-goat (Invitrogen A11057); Anti-Venezuelan equine encephalitis antibody clone 1A3B-7 (EMD Millipore MAB8755); and polyclonal goat serum to TC-83 capsid protein (BEI Resources NR-9403).

    Techniques: Produced, Infection

    Indirect inflammatory response dependent on mitochondrially derived ROS. (a) Experimental workflow indicating that U-87 MG cells were pre-treated for 2h with 0.1% DMSO, 1µM BAY 11–7082 (BAY-82), or 100pM mitoquinone mesylate (MitoQ) prior to inoculation with TC-83 (MOI:2). Supernatants were removed from infected U-87 MG cells at 2hpi and overlaid onto naïve HMC3 microglia, with (b) IL-1α, (c) IL-1β, (d) IL-6, and (e) IL-8 cytokine levels measured at 1, 2, and 6 hours post overlay. The quantitative data are depicted as the means of three biologically independent experiments ± SD. * p

    Journal: Virulence

    Article Title: Direct and indirect pro-inflammatory cytokine response resulting from TC-83 infection of glial cells

    doi: 10.1080/21505594.2018.1509668

    Figure Lengend Snippet: Indirect inflammatory response dependent on mitochondrially derived ROS. (a) Experimental workflow indicating that U-87 MG cells were pre-treated for 2h with 0.1% DMSO, 1µM BAY 11–7082 (BAY-82), or 100pM mitoquinone mesylate (MitoQ) prior to inoculation with TC-83 (MOI:2). Supernatants were removed from infected U-87 MG cells at 2hpi and overlaid onto naïve HMC3 microglia, with (b) IL-1α, (c) IL-1β, (d) IL-6, and (e) IL-8 cytokine levels measured at 1, 2, and 6 hours post overlay. The quantitative data are depicted as the means of three biologically independent experiments ± SD. * p

    Article Snippet: Antibodies, inhibitors, and reagents The reagents and antibodies (Abs) used were as follows: dimethyl sulfoxide (DMSO) (VWR 97,063–136); the mitochondrially targeted antioxidant mitoquinone mesylate (MedKoo Biosciences 317,102); the NF-κB inhibitor BAY 11–7082 (Selleckchem S2913); the IL-1β inhibitor Pirfenidone (SelleckChem S2907); IL-1α, IL-1β, IL-6, and IL-8 recombinant protein (Aushon 101-3FF-1-AB); mouse monoclonal Ab to TOMM20 (Abcam ab56783), Alexa Fluor 488-conjugated goat anti-mouse (Invitrogen A21202), Alexa Fluor 568-conjugated donkey anti-goat (Invitrogen A11057); Anti-Venezuelan equine encephalitis antibody clone 1A3B-7 (EMD Millipore MAB8755); and polyclonal goat serum to TC-83 capsid protein (BEI Resources NR-9403).

    Techniques: Derivative Assay, Infection

    Inhibition of the Piezo1 channel in bladder ICC-LCs by GsMTx4 significantly alleviated bladder hyperactivity in CYP-8d rats. a – c Compared with the vehicle (DMSO), 2.5 μM GsMTx4 significantly decreased the MBP and prolonged the ICI in naive and CYP-8d rats, and naive rats showed lower sensitivity to GsMTx4 than CYP-8d rats (the data represent the mean ± S.D., n = 8; * P

    Journal: Experimental & Molecular Medicine

    Article Title: Increased Piezo1 channel activity in interstitial Cajal-like cells induces bladder hyperactivity by functionally interacting with NCX1 in rats with cyclophosphamide-induced cystitis

    doi: 10.1038/s12276-018-0088-z

    Figure Lengend Snippet: Inhibition of the Piezo1 channel in bladder ICC-LCs by GsMTx4 significantly alleviated bladder hyperactivity in CYP-8d rats. a – c Compared with the vehicle (DMSO), 2.5 μM GsMTx4 significantly decreased the MBP and prolonged the ICI in naive and CYP-8d rats, and naive rats showed lower sensitivity to GsMTx4 than CYP-8d rats (the data represent the mean ± S.D., n = 8; * P

    Article Snippet: Physiological saline containing Grammostola spatulata mechanotoxin 4 (GsMTx4, Piezo1 antagonist, 2.5 μM, Abcam) or dimethyl sulfoxide (DMSO) (Beyotime) at room temperature was infused into the bladder at a constant rate (10 mL/h).

    Techniques: Inhibition, Immunocytochemistry

    Infliximab prevents Zaprinast-induced glial fibrillary acidic protein (GFAP) overexpression in cultured porcine retina. Retinal explants were incubated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast as described in Methods. (A) Confocal laser scanning micrographs of retinal sections showing GFAP content. Scale bar: 50 μm. (B) Bar graphs showing the quantification of GFAP content. Values are the mean ±SEM of six different cultures. Values that are significantly different are indicated by asterisks * P

    Journal: Journal of Neuroinflammation

    Article Title: Infliximab reduces Zaprinast-induced retinal degeneration in cultures of porcine retina

    doi: 10.1186/s12974-014-0172-9

    Figure Lengend Snippet: Infliximab prevents Zaprinast-induced glial fibrillary acidic protein (GFAP) overexpression in cultured porcine retina. Retinal explants were incubated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast as described in Methods. (A) Confocal laser scanning micrographs of retinal sections showing GFAP content. Scale bar: 50 μm. (B) Bar graphs showing the quantification of GFAP content. Values are the mean ±SEM of six different cultures. Values that are significantly different are indicated by asterisks * P

    Article Snippet: Zaprinast (Sigma-Aldrich, Madrid, Spain) was diluted in dimethyl sulfoxide (DMSO) (AppliChem, Darmstadt, Germany).

    Techniques: Over Expression, Cell Culture, Incubation

    Infliximab prevents Zaprinast-induced cell death in cultured porcine retina. Retinal explants were incubated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast as described in Methods. Confocal laser scanning micrographs of retinal sections showing TUNEL-stained sections visualizing apoptotic photoreceptors (pink) (A) , cleaved caspase-3 positive cells (red) (B) and PAR accumulation (pink) (C) in SYTOX Green-counterstained retinal sections. Scale bar: 50 μm. (D) Bar graphs showing the quantification of TUNEL, cleaved caspase-3 and PAR accumulation. Values are the mean ± SEM of seven different cultures. Values that are significantly different are indicated by asterisks * P

    Journal: Journal of Neuroinflammation

    Article Title: Infliximab reduces Zaprinast-induced retinal degeneration in cultures of porcine retina

    doi: 10.1186/s12974-014-0172-9

    Figure Lengend Snippet: Infliximab prevents Zaprinast-induced cell death in cultured porcine retina. Retinal explants were incubated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast as described in Methods. Confocal laser scanning micrographs of retinal sections showing TUNEL-stained sections visualizing apoptotic photoreceptors (pink) (A) , cleaved caspase-3 positive cells (red) (B) and PAR accumulation (pink) (C) in SYTOX Green-counterstained retinal sections. Scale bar: 50 μm. (D) Bar graphs showing the quantification of TUNEL, cleaved caspase-3 and PAR accumulation. Values are the mean ± SEM of seven different cultures. Values that are significantly different are indicated by asterisks * P

    Article Snippet: Zaprinast (Sigma-Aldrich, Madrid, Spain) was diluted in dimethyl sulfoxide (DMSO) (AppliChem, Darmstadt, Germany).

    Techniques: Cell Culture, Incubation, TUNEL Assay, Staining

    Co-localization of caspase-3 , PAR and TUNEL at different nuclear layers in culture of porcine retina. Triple-imnunofluorescence labeling of retinal explants treated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast was carried out as described in Methods. Confocal laser scanning micrographs of retinal sections showing immunolocalization of TUNEL (red), cleaved caspase-3 (blue) and PAR (green)-positive cells in the nuclear layers of retina. Scale bar: 10 μm. GCL: ganglion nuclear layer; INL: inner nuclear layer; ONL: outer nuclear layer; OS: outer segments; C: control; Z100: 100 μM Zaprinast; INF: 2 μg/mL Infliximab; Z100 + INF: 100 μM Zaprinast with 2 μg/mL Infliximab. TUNEL, terminal deoxynucleotidil transferase dUTP nick and labeling.

    Journal: Journal of Neuroinflammation

    Article Title: Infliximab reduces Zaprinast-induced retinal degeneration in cultures of porcine retina

    doi: 10.1186/s12974-014-0172-9

    Figure Lengend Snippet: Co-localization of caspase-3 , PAR and TUNEL at different nuclear layers in culture of porcine retina. Triple-imnunofluorescence labeling of retinal explants treated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast was carried out as described in Methods. Confocal laser scanning micrographs of retinal sections showing immunolocalization of TUNEL (red), cleaved caspase-3 (blue) and PAR (green)-positive cells in the nuclear layers of retina. Scale bar: 10 μm. GCL: ganglion nuclear layer; INL: inner nuclear layer; ONL: outer nuclear layer; OS: outer segments; C: control; Z100: 100 μM Zaprinast; INF: 2 μg/mL Infliximab; Z100 + INF: 100 μM Zaprinast with 2 μg/mL Infliximab. TUNEL, terminal deoxynucleotidil transferase dUTP nick and labeling.

    Article Snippet: Zaprinast (Sigma-Aldrich, Madrid, Spain) was diluted in dimethyl sulfoxide (DMSO) (AppliChem, Darmstadt, Germany).

    Techniques: TUNEL Assay, Labeling

    Infliximab partially prevents Zaprinast-induced oxidative stress in cultures of porcine retina. Retinal explants were incubated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast as described in Methods. Effect of Infliximab on the total antioxidant capacity (A) , TBARS formation (B) and intracellular NOX (C) . Each sample was measured in duplicate, and the values are the mean ±SEM of eight cultures. ANOVA Newman-Keuls post-test was used for TAC analysis. Kruskal-Wallis test and Dunn’s post-test was used for TBARS and iNOX analysis. * P

    Journal: Journal of Neuroinflammation

    Article Title: Infliximab reduces Zaprinast-induced retinal degeneration in cultures of porcine retina

    doi: 10.1186/s12974-014-0172-9

    Figure Lengend Snippet: Infliximab partially prevents Zaprinast-induced oxidative stress in cultures of porcine retina. Retinal explants were incubated with dimethyl sulfoxide (DMSO), Zaprinast and Infliximab alone or combined with Zaprinast as described in Methods. Effect of Infliximab on the total antioxidant capacity (A) , TBARS formation (B) and intracellular NOX (C) . Each sample was measured in duplicate, and the values are the mean ±SEM of eight cultures. ANOVA Newman-Keuls post-test was used for TAC analysis. Kruskal-Wallis test and Dunn’s post-test was used for TBARS and iNOX analysis. * P

    Article Snippet: Zaprinast (Sigma-Aldrich, Madrid, Spain) was diluted in dimethyl sulfoxide (DMSO) (AppliChem, Darmstadt, Germany).

    Techniques: Incubation

    Inhibition of ALDH1A2 expression by GSK3 inhibitor CHIR99021 is via β-catenin signaling. (A) Effect of CHIR99021 on β-catenin nuclear translocation in WiT49 cells. WiT49 cells were treated with 5 μM CHIR99021 or 0.1% DMSO for 48 h, and then immunofluorescence was performed with β-catenin antibody and DAPI staining. Negative control represents no primary antibody control. Bar length equals 50 μm. (B) Effect of CHIR99021 on the TCF reporter in WiT49 cells. A 500-ng sample of TOPFlash or FOPFlash constructs, together with 50 ng of the pRL-TK renilla control vector, was transiently transfected into WiT49 cells. Cells were treated with 0.1% DMSO or 5 μM CHIR99021 after 24 h of transfection. Luciferase reporter activities relative to renilla (pRL-TK) are shown in percentage terms. Values are the mean ± SD of three determinations from separate experiments. ∗∗∗ P

    Journal: Frontiers in Cell and Developmental Biology

    Article Title: Inhibition of GSK3 Represses the Expression of Retinoic Acid Synthetic Enzyme ALDH1A2 via Wnt/β-Catenin Signaling in WiT49 Cells

    doi: 10.3389/fcell.2020.00094

    Figure Lengend Snippet: Inhibition of ALDH1A2 expression by GSK3 inhibitor CHIR99021 is via β-catenin signaling. (A) Effect of CHIR99021 on β-catenin nuclear translocation in WiT49 cells. WiT49 cells were treated with 5 μM CHIR99021 or 0.1% DMSO for 48 h, and then immunofluorescence was performed with β-catenin antibody and DAPI staining. Negative control represents no primary antibody control. Bar length equals 50 μm. (B) Effect of CHIR99021 on the TCF reporter in WiT49 cells. A 500-ng sample of TOPFlash or FOPFlash constructs, together with 50 ng of the pRL-TK renilla control vector, was transiently transfected into WiT49 cells. Cells were treated with 0.1% DMSO or 5 μM CHIR99021 after 24 h of transfection. Luciferase reporter activities relative to renilla (pRL-TK) are shown in percentage terms. Values are the mean ± SD of three determinations from separate experiments. ∗∗∗ P

    Article Snippet: CHIR99021 was dissolved in dimethyl sulfoxide (DMSO) (Cat.196055, MP Biomedicals).

    Techniques: Inhibition, Expressing, Translocation Assay, Immunofluorescence, Staining, Negative Control, Construct, Plasmid Preparation, Transfection, Luciferase

    GSK3 inhibitor CHIR99021 inhibits Aldh1a2 expression in WiT49 cells. (A) Effect of CHIR99021 on ALDH1A2 mRNA expression in WiT49 cells. WiT49 cells were treated with 0.1% DMSO (control vehicle) or 5 μM CHIR99021 for 48 h. ALDH1A2 expression relative to TBP was measured by real-time RT-PCR. Values are the mean ± SD of three determinations; representative results from at least three separate experiments. ∗∗∗ P

    Journal: Frontiers in Cell and Developmental Biology

    Article Title: Inhibition of GSK3 Represses the Expression of Retinoic Acid Synthetic Enzyme ALDH1A2 via Wnt/β-Catenin Signaling in WiT49 Cells

    doi: 10.3389/fcell.2020.00094

    Figure Lengend Snippet: GSK3 inhibitor CHIR99021 inhibits Aldh1a2 expression in WiT49 cells. (A) Effect of CHIR99021 on ALDH1A2 mRNA expression in WiT49 cells. WiT49 cells were treated with 0.1% DMSO (control vehicle) or 5 μM CHIR99021 for 48 h. ALDH1A2 expression relative to TBP was measured by real-time RT-PCR. Values are the mean ± SD of three determinations; representative results from at least three separate experiments. ∗∗∗ P

    Article Snippet: CHIR99021 was dissolved in dimethyl sulfoxide (DMSO) (Cat.196055, MP Biomedicals).

    Techniques: Expressing, Quantitative RT-PCR

    Increases in Mcl-1 and Bcl-xL expression were mediated by YAP1. a Relative gene expression of MCL1 and BCLXL detected by qRT-PCR in 4 ALK-rearranged lung cancer cell lines ( n = 4 independent experiments). Significance was evaluated using a one-way ANOVA followed by Sidak’s test. b Evaluation of Mcl-1 expression, Bcl-xL expression, and YAP phosphorylation (pS127). Cell lysates were blotted using an immunoblotting assay with the indicated antibody. c Initial survival rate (top) and apoptosis activity (bottom) of KTOR1 (left) and KTOR2 (right) when cells were exposed to ALC or vehicle in combination with the knockdown of MCL1 and BCLXL . Summarized results are shown in the panels. The number of independent experiments is shown in each panel. Complete results are presented in Supplementary Fig. 6c, d . Significance was evaluated using a one-way ANOVA followed by Sidak’s test. d – e Increases in Mcl-1 and Bcl-xL expression were canceled by the inhibition of YAP1. YAP1 activity was inhibited using siRNA ( d ) or verteporfin ( e ). Traced experiments using H2228 are shown in Supplementary Fig. 6f, g . f Sequence analysis of putative YAP1/TEAD binding sites (labeled as A, B) in the upstream regions of MCL1 and BCLXL . The ChIP-PCR primer pairs designed were shown as arrows. g ChIP-qPCR on KTOR1 and KTOR2, which confirmed the presence of YAP1/TEAD binding sites in the MCL1 and BCLXL upstream regions and increased binding induced by the ALC treatment. BCLXL , a known YAP target, was also used as a positive control. Results are expressed as Percent Input (Two biologically independent cells, two-independent primers, two-independent experiment in each cell lines and two-independent qRT-PCR quantification in each experiment). The titration of DNA digestion, electrophoresis of input samples, and specificity of ChIP-qRT-PCR primers were shown in Supplementary Fig. 8 . Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. h Schematic explanation of Fig. 6. The inhibition of YAP1 down-regulated Mcl-1 and Bcl-xL expression. The inhibition of both Mcl-1 and Bcl-xL induced apoptosis and inhibited initial survival. ALC alectinib, DMSO dimethyl sulfoxide, VER verteporfin, siSC si scramble control. Error bars indicate ± S.D. * P

    Journal: Nature Communications

    Article Title: YAP1 mediates survival of ALK-rearranged lung cancer cells treated with alectinib via pro-apoptotic protein regulation

    doi: 10.1038/s41467-019-13771-5

    Figure Lengend Snippet: Increases in Mcl-1 and Bcl-xL expression were mediated by YAP1. a Relative gene expression of MCL1 and BCLXL detected by qRT-PCR in 4 ALK-rearranged lung cancer cell lines ( n = 4 independent experiments). Significance was evaluated using a one-way ANOVA followed by Sidak’s test. b Evaluation of Mcl-1 expression, Bcl-xL expression, and YAP phosphorylation (pS127). Cell lysates were blotted using an immunoblotting assay with the indicated antibody. c Initial survival rate (top) and apoptosis activity (bottom) of KTOR1 (left) and KTOR2 (right) when cells were exposed to ALC or vehicle in combination with the knockdown of MCL1 and BCLXL . Summarized results are shown in the panels. The number of independent experiments is shown in each panel. Complete results are presented in Supplementary Fig. 6c, d . Significance was evaluated using a one-way ANOVA followed by Sidak’s test. d – e Increases in Mcl-1 and Bcl-xL expression were canceled by the inhibition of YAP1. YAP1 activity was inhibited using siRNA ( d ) or verteporfin ( e ). Traced experiments using H2228 are shown in Supplementary Fig. 6f, g . f Sequence analysis of putative YAP1/TEAD binding sites (labeled as A, B) in the upstream regions of MCL1 and BCLXL . The ChIP-PCR primer pairs designed were shown as arrows. g ChIP-qPCR on KTOR1 and KTOR2, which confirmed the presence of YAP1/TEAD binding sites in the MCL1 and BCLXL upstream regions and increased binding induced by the ALC treatment. BCLXL , a known YAP target, was also used as a positive control. Results are expressed as Percent Input (Two biologically independent cells, two-independent primers, two-independent experiment in each cell lines and two-independent qRT-PCR quantification in each experiment). The titration of DNA digestion, electrophoresis of input samples, and specificity of ChIP-qRT-PCR primers were shown in Supplementary Fig. 8 . Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. h Schematic explanation of Fig. 6. The inhibition of YAP1 down-regulated Mcl-1 and Bcl-xL expression. The inhibition of both Mcl-1 and Bcl-xL induced apoptosis and inhibited initial survival. ALC alectinib, DMSO dimethyl sulfoxide, VER verteporfin, siSC si scramble control. Error bars indicate ± S.D. * P

    Article Snippet: ALC, crizotinib, ceritinib, and VER were dissolved in dimethyl sulfoxide (DMSO) (Nacalai Tesque) at a concentration of 5 mmol/L.

    Techniques: Expressing, Quantitative RT-PCR, Activity Assay, Inhibition, Sequencing, Binding Assay, Labeling, Chromatin Immunoprecipitation, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Positive Control, Titration, Electrophoresis

    A subpart of ALK-rearranged tumors survived the ALC treatment. a Tumor volumes in 13 patients with ALK-rearranged lung cancer receiving ALC as first-line therapy in our institution. Representative CT image of a patient (top left). Changes in tumor volumes during the ALC treatment (top right). Significance was evaluated using a one-way ANOVA followed by Dunnett’s multiple comparison test. Tumor responses in 13 patients in the first evaluation (1–2 months from treatment initiation) or at the best response (bottom). b Schematic explanation of initial survival from ALK inhibitors. ALK inhibitors exert specific effects on ALK-rearranged tumors; however, some tumor cells survive (initial survival) this treatment. During months-years of treatment, tumors eventually relapsed with acquired resistance to ALK inhibitors. A waterfall plot of the changes in tumor diameter based on RECIST guideline are shown in Supplementary Fig. 1a . c Schematic explanation of the establishment of patient-derived cell lines. d – e Some ALK-rearranged NSCLC cells survived the ALC treatment; however, their proliferation was significantly suppressed under the ALC treatment in vitro. Proliferation tests ( d ) and initial survival rates ( e ) of KTOR1 and H2228 cells exposed to stepwise concentrations of ALC or DMSO ( n = 6 independent experiments). Significance was evaluated using a one-way ANOVA followed by Dunnett’s multiple comparison test. The survival rate was calculated using the formula: (Cell number at 96 h)/(Cell number at 0 h = baseline) × 100. Relative cell growth was calculated by (survival rate)-100. The positive columns indicate increased cell numbers during the 96-h treatment and negative columns indicate decreased cell numbers. The results for traced experiments in KTOR2 and KTOR3 were presented in Supplementary Fig. 1b, c . CR complete response, PR partial response, SD stable disease, ALK anaplastic lymphoma kinase, NSCLC non-small cell lung cancer, ALC alectinib, DMSO dimethyl sulfoxide. Error bars indicate ± S.D. * P

    Journal: Nature Communications

    Article Title: YAP1 mediates survival of ALK-rearranged lung cancer cells treated with alectinib via pro-apoptotic protein regulation

    doi: 10.1038/s41467-019-13771-5

    Figure Lengend Snippet: A subpart of ALK-rearranged tumors survived the ALC treatment. a Tumor volumes in 13 patients with ALK-rearranged lung cancer receiving ALC as first-line therapy in our institution. Representative CT image of a patient (top left). Changes in tumor volumes during the ALC treatment (top right). Significance was evaluated using a one-way ANOVA followed by Dunnett’s multiple comparison test. Tumor responses in 13 patients in the first evaluation (1–2 months from treatment initiation) or at the best response (bottom). b Schematic explanation of initial survival from ALK inhibitors. ALK inhibitors exert specific effects on ALK-rearranged tumors; however, some tumor cells survive (initial survival) this treatment. During months-years of treatment, tumors eventually relapsed with acquired resistance to ALK inhibitors. A waterfall plot of the changes in tumor diameter based on RECIST guideline are shown in Supplementary Fig. 1a . c Schematic explanation of the establishment of patient-derived cell lines. d – e Some ALK-rearranged NSCLC cells survived the ALC treatment; however, their proliferation was significantly suppressed under the ALC treatment in vitro. Proliferation tests ( d ) and initial survival rates ( e ) of KTOR1 and H2228 cells exposed to stepwise concentrations of ALC or DMSO ( n = 6 independent experiments). Significance was evaluated using a one-way ANOVA followed by Dunnett’s multiple comparison test. The survival rate was calculated using the formula: (Cell number at 96 h)/(Cell number at 0 h = baseline) × 100. Relative cell growth was calculated by (survival rate)-100. The positive columns indicate increased cell numbers during the 96-h treatment and negative columns indicate decreased cell numbers. The results for traced experiments in KTOR2 and KTOR3 were presented in Supplementary Fig. 1b, c . CR complete response, PR partial response, SD stable disease, ALK anaplastic lymphoma kinase, NSCLC non-small cell lung cancer, ALC alectinib, DMSO dimethyl sulfoxide. Error bars indicate ± S.D. * P

    Article Snippet: ALC, crizotinib, ceritinib, and VER were dissolved in dimethyl sulfoxide (DMSO) (Nacalai Tesque) at a concentration of 5 mmol/L.

    Techniques: Derivative Assay, In Vitro

    Proteome analysis to identify acute response of ALK-rearranged cells to ALC. a A schematic explanation of the experiment to identify the acute response of ALK-rearranged cells to alectinib. Four ALK-rearranged NSCLC cells with alectinib sensitivity were exposed to DMSO or the survivable inhibitory concentration (sIC) of ALC (see Table 1 ) for 96 h. Proteome and conventional biological analyses were performed to compare cells that survived ALC and proliferating control cells (DMSO-treated). b The integrated results of the proteome analysis in ALK-rearranged cell lines ( n = 3 biologically independent cell lines, KTOR3 was excluded, please see the text). Volcano plot representing all expressed proteins. Regarding every protein, fold changes in the vehicle and exposure to alectinib were plotted against the –log P -value. The integrated P -value was calculated by paired t-test. Significant differentially expressed proteins, with a fold change ≥1.5 or ≤−1.5, are depicted as red or blue, while insignificant ones are shown as black dots. Of the 3183 proteins detected after the 96-h exposure to alectinib, 48 were differentially expressed (18 up- and 30 downregulated). c Gene ontology (GO) analysis of the 48 proteins. P -values were calculated using Fisher’s test. d KEGG pathway analysis on the upregulated proteins. P -values were calculated using Fisher’s test. All annotated pathways are presented. e Average cell area on the culture plate. A plot shows the average cell area on each photographed image. Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. f Cell adhesion areas in culture plates increased when ALK-rearranged cells were treated with ALC. g A simplified diagram of the Hippo signaling pathway using KEGG pathway analysis. Two important factors (Ajuba, AFP) among the four annotated proteins are shown. All analysis results are described in Supplementary Data 2 . * P

    Journal: Nature Communications

    Article Title: YAP1 mediates survival of ALK-rearranged lung cancer cells treated with alectinib via pro-apoptotic protein regulation

    doi: 10.1038/s41467-019-13771-5

    Figure Lengend Snippet: Proteome analysis to identify acute response of ALK-rearranged cells to ALC. a A schematic explanation of the experiment to identify the acute response of ALK-rearranged cells to alectinib. Four ALK-rearranged NSCLC cells with alectinib sensitivity were exposed to DMSO or the survivable inhibitory concentration (sIC) of ALC (see Table 1 ) for 96 h. Proteome and conventional biological analyses were performed to compare cells that survived ALC and proliferating control cells (DMSO-treated). b The integrated results of the proteome analysis in ALK-rearranged cell lines ( n = 3 biologically independent cell lines, KTOR3 was excluded, please see the text). Volcano plot representing all expressed proteins. Regarding every protein, fold changes in the vehicle and exposure to alectinib were plotted against the –log P -value. The integrated P -value was calculated by paired t-test. Significant differentially expressed proteins, with a fold change ≥1.5 or ≤−1.5, are depicted as red or blue, while insignificant ones are shown as black dots. Of the 3183 proteins detected after the 96-h exposure to alectinib, 48 were differentially expressed (18 up- and 30 downregulated). c Gene ontology (GO) analysis of the 48 proteins. P -values were calculated using Fisher’s test. d KEGG pathway analysis on the upregulated proteins. P -values were calculated using Fisher’s test. All annotated pathways are presented. e Average cell area on the culture plate. A plot shows the average cell area on each photographed image. Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. f Cell adhesion areas in culture plates increased when ALK-rearranged cells were treated with ALC. g A simplified diagram of the Hippo signaling pathway using KEGG pathway analysis. Two important factors (Ajuba, AFP) among the four annotated proteins are shown. All analysis results are described in Supplementary Data 2 . * P

    Article Snippet: ALC, crizotinib, ceritinib, and VER were dissolved in dimethyl sulfoxide (DMSO) (Nacalai Tesque) at a concentration of 5 mmol/L.

    Techniques: Concentration Assay

    YAP1 was activated by ALC in vitro and in vivo. a YAP1 localized in the nucleus when ALK-rearranged lung cancer cells were exposed to ALC. The cell area was also increased by the exposure to ALC. Scale bar = 100 µm. b Representative colocalization values. The brightness of YAP1-Alexa488 and Hoechst at every dot on the image was plotted and Pearson’s R value was calculated (top). Original images of immunofluorescence-labeled YAP1 (middle) and Hoechst (bottom). Nuclear localization strongly correlated with R values. Scale bar = 100 µm. c Increased nuclear localization of YAP1 in 4 ALK-rearranged lung cancers. Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. d Dose dependency of the nuclear localization of YAP1 in KTOR1 cells. The number of images analyzed is presented in the figure. The values in the three other cell lines are shown in Supplementary Fig. 3 . Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. e ALC exposure time dependency of the nuclear localization of YAP1 in KTOR1 cells. The number of images analyzed is presented in the figure. Significance was evaluated using a one-way ANOVA followed by Dunnett’s multiple comparison test. f Experimental design of the xenograft study. ALK-rearranged xenografts on nude mice were randomized to ALC or vehicle treatment groups. Treated xenografts were fixed and immunohistochemically stained by YAP1. g Increased nuclear localization of YAP1 when ALK-rearranged xenografts were treated with ALC for 7 days. h Increased nuclear localization of YAP1 in ALK-rearranged xenografts treated with ALC ( n = 4 biologically independent xenografts). Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. i Schematic explanation of the results shown in Fig. 3. Exposure to YAP1 activation in ALK-rearranged lung cancer. YAP1 activation was evaluated by the nuclear localization of YAP1. ALC alectinib, N > C Nuclear > Cytoplasm, CRZ crizotinib, CER ceritinib, DMSO dimethyl sulfoxide, ALK anaplastic lymphoma kinase, IHC immunohistochemistry. Error bars indicate ± S.D. * P

    Journal: Nature Communications

    Article Title: YAP1 mediates survival of ALK-rearranged lung cancer cells treated with alectinib via pro-apoptotic protein regulation

    doi: 10.1038/s41467-019-13771-5

    Figure Lengend Snippet: YAP1 was activated by ALC in vitro and in vivo. a YAP1 localized in the nucleus when ALK-rearranged lung cancer cells were exposed to ALC. The cell area was also increased by the exposure to ALC. Scale bar = 100 µm. b Representative colocalization values. The brightness of YAP1-Alexa488 and Hoechst at every dot on the image was plotted and Pearson’s R value was calculated (top). Original images of immunofluorescence-labeled YAP1 (middle) and Hoechst (bottom). Nuclear localization strongly correlated with R values. Scale bar = 100 µm. c Increased nuclear localization of YAP1 in 4 ALK-rearranged lung cancers. Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. d Dose dependency of the nuclear localization of YAP1 in KTOR1 cells. The number of images analyzed is presented in the figure. The values in the three other cell lines are shown in Supplementary Fig. 3 . Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. e ALC exposure time dependency of the nuclear localization of YAP1 in KTOR1 cells. The number of images analyzed is presented in the figure. Significance was evaluated using a one-way ANOVA followed by Dunnett’s multiple comparison test. f Experimental design of the xenograft study. ALK-rearranged xenografts on nude mice were randomized to ALC or vehicle treatment groups. Treated xenografts were fixed and immunohistochemically stained by YAP1. g Increased nuclear localization of YAP1 when ALK-rearranged xenografts were treated with ALC for 7 days. h Increased nuclear localization of YAP1 in ALK-rearranged xenografts treated with ALC ( n = 4 biologically independent xenografts). Significance was evaluated using a one-way ANOVA followed by Sidak’s multiple comparison test. i Schematic explanation of the results shown in Fig. 3. Exposure to YAP1 activation in ALK-rearranged lung cancer. YAP1 activation was evaluated by the nuclear localization of YAP1. ALC alectinib, N > C Nuclear > Cytoplasm, CRZ crizotinib, CER ceritinib, DMSO dimethyl sulfoxide, ALK anaplastic lymphoma kinase, IHC immunohistochemistry. Error bars indicate ± S.D. * P

    Article Snippet: ALC, crizotinib, ceritinib, and VER were dissolved in dimethyl sulfoxide (DMSO) (Nacalai Tesque) at a concentration of 5 mmol/L.

    Techniques: In Vitro, In Vivo, Immunofluorescence, Labeling, Mouse Assay, Staining, Activation Assay, Immunohistochemistry

    ( A ) SEC elugram of AMCA-PLGA; the black line indicates the refractive index of the PLGA expressed as relative units (RIU), and the gray line indicates the ultraviolet absorption by AMCA at 350 nm expressed as relative milli-units (mAU). ( B ) Stability studies via fluorescence intensity measurements of AMCA-PLGA in DMSO for > 120 hours at 4°C (solid line, crosses), 20°C (dotted line, triangles) and 37°C (broken line, circles). Data are expressed as ratios of I/I 0 and shown as the mean ± SD of three independent experiments. Abbreviations: AMCA-PLGA, 7-amino-4-methyl-3-coumarinylacetic acid-poly( d,l -lactide-co-glycolide); DMSO, dimethyl sulfoxide; mAU, milli-absorption unit; PLGA, poly( d,l -lactide-co-glycolide); RIU, refractive index unit; SD, standard deviation; SEC, size exclusion chromatography.

    Journal: International Journal of Nanomedicine

    Article Title: A blue fluorescent labeling technique utilizing micro- and nanoparticles for tracking in LIVE/DEAD® stained pathogenic biofilms of Staphylococcus aureus and Burkholderia cepacia

    doi: 10.2147/IJN.S98401

    Figure Lengend Snippet: ( A ) SEC elugram of AMCA-PLGA; the black line indicates the refractive index of the PLGA expressed as relative units (RIU), and the gray line indicates the ultraviolet absorption by AMCA at 350 nm expressed as relative milli-units (mAU). ( B ) Stability studies via fluorescence intensity measurements of AMCA-PLGA in DMSO for > 120 hours at 4°C (solid line, crosses), 20°C (dotted line, triangles) and 37°C (broken line, circles). Data are expressed as ratios of I/I 0 and shown as the mean ± SD of three independent experiments. Abbreviations: AMCA-PLGA, 7-amino-4-methyl-3-coumarinylacetic acid-poly( d,l -lactide-co-glycolide); DMSO, dimethyl sulfoxide; mAU, milli-absorption unit; PLGA, poly( d,l -lactide-co-glycolide); RIU, refractive index unit; SD, standard deviation; SEC, size exclusion chromatography.

    Article Snippet: Stability of AMCA-PLGA A 4 mg/mL solution of AMCA-PLGA was prepared in dimethyl sulfoxide (DMSO) (ROTIPURAN® , ≥99.9%, p.a., Carl Roth GmbH), and 200 μL aliquots were transferred to 96-well plates (Greiner Bio-One, Frickenhausen, Germany).

    Techniques: Size-exclusion Chromatography, Fluorescence, Standard Deviation

    Toxicity of the four selected small molecules (SMs) on several eukaryotic models. ( A ) Cell toxicity and hemolytic activity of the SMs after 24 hrs and 1 hr treatment with 200 μM, respectively. Data were normalized with a 0.1% triton-100X control. Bar: standard deviation; n = 8 replicates per group. ( B ) Galleria mellonella larva survival rate after a single treatment with 12.5 μg of SM per larva. Larva death was monitored every 12 hrs for three days after treatment. Both untreated larvae (NC) and larvae treated with 1% dimethyl sulfoxide (DMSO) had 100% survival (blue line). RBCs: red blood cells; n = 15 replicates per group.

    Journal: Scientific Reports

    Article Title: Novel Imidazole and Methoxybenzylamine Growth Inhibitors Affecting Salmonella Cell Envelope Integrity and its Persistence in Chickens

    doi: 10.1038/s41598-018-31249-0

    Figure Lengend Snippet: Toxicity of the four selected small molecules (SMs) on several eukaryotic models. ( A ) Cell toxicity and hemolytic activity of the SMs after 24 hrs and 1 hr treatment with 200 μM, respectively. Data were normalized with a 0.1% triton-100X control. Bar: standard deviation; n = 8 replicates per group. ( B ) Galleria mellonella larva survival rate after a single treatment with 12.5 μg of SM per larva. Larva death was monitored every 12 hrs for three days after treatment. Both untreated larvae (NC) and larvae treated with 1% dimethyl sulfoxide (DMSO) had 100% survival (blue line). RBCs: red blood cells; n = 15 replicates per group.

    Article Snippet: The SMs were suspended into 100% dimethyl sulfoxide (DMSO) to 10 mM concentration and stored at −80 °C in 96-well plates sealed with Thermowell seal tape (Corning).

    Techniques: Activity Assay, Standard Deviation