β-actin Search Results


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  • 90
    Thermo Fisher β actin mn00607939 s1
    β Actin Mn00607939 S1, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 90/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β actin mn00607939 s1/product/Thermo Fisher
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    β actin mn00607939 s1 - by Bioz Stars, 2020-05
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    99
    Cell Signaling Technology Inc anti βactin
    ( A,C ) Changes in the expression levels of matrix metalloproteinases (MMPs) following treatment with 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B,D ) The band intensities of TIMP-1,MMP-9 and MMP-2 relative to untreated control cells were quantified upon normalizing to <t>β-actin</t> expression, and are expressed as the mean ± standard deviation of three independent experiments.
    Anti βactin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 99/100, based on 7757 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti βactin/product/Cell Signaling Technology Inc
    Average 99 stars, based on 7757 article reviews
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    92
    Proteintech β‑actin
    ( A,C ) Changes in the expression levels of matrix metalloproteinases (MMPs) following treatment with 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B,D ) The band intensities of TIMP-1,MMP-9 and MMP-2 relative to untreated control cells were quantified upon normalizing to <t>β-actin</t> expression, and are expressed as the mean ± standard deviation of three independent experiments.
    β‑Actin, supplied by Proteintech, used in various techniques. Bioz Stars score: 92/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/β‑actin/product/Proteintech
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    β‑actin - by Bioz Stars, 2020-05
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    90
    Cell Signaling Technology Inc β actin protein
    HBxΔ127 has a greater capacity to stimulate SREBP-1c relative to wild type HBx. (A, B) HepG2-X (or H7402-X) cells and HepG2-XΔ127 (or H7402-XΔ127) cells were transfected with pSilencer 3.0-X plasmid for 48 h. (A) The mRNA levels of SREBP-1c were detected by RT-PCR. (B) Immunoblot analysis to detect SREBP-1c (top panel) and <t>β-actin</t> protein levels (bottom panel). (C, D) HepG2-X (or H7402-X) cells and HepG2-XΔ127 (or H7402-XΔ127) cells were co-transfected with or without pSilencer 3.0-X plasmid and SREBP-1c-571-Luc-WT. Data are representative of 3 independent experiments. Values represent means±SD. n =3. b P
    β Actin Protein, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 90/100, based on 72 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Average 90 stars, based on 72 article reviews
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    Image Search Results


    ( A,C ) Changes in the expression levels of matrix metalloproteinases (MMPs) following treatment with 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B,D ) The band intensities of TIMP-1,MMP-9 and MMP-2 relative to untreated control cells were quantified upon normalizing to β-actin expression, and are expressed as the mean ± standard deviation of three independent experiments.

    Journal: Molecules

    Article Title: 28-Hydroxy-3-oxoolean-12-en-29-oic Acid, a Triterpene Acid from Celastrus Orbiculatus Extract, Inhibits the Migration and Invasion of Human Gastric Cancer Cells In Vitro

    doi: 10.3390/molecules24193513

    Figure Lengend Snippet: ( A,C ) Changes in the expression levels of matrix metalloproteinases (MMPs) following treatment with 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B,D ) The band intensities of TIMP-1,MMP-9 and MMP-2 relative to untreated control cells were quantified upon normalizing to β-actin expression, and are expressed as the mean ± standard deviation of three independent experiments.

    Article Snippet: Reagents The reagents used included the following: RPMI 1640 cell culture medium, trypsin (HyClone, USA); fetal bovine serum (FBS) (Gibco, Waltham, MA, USA); artificially reconstituted basement membrane, Transwell chambers (Corning, Corning, NY, USA); thiazole blue (MTT) powder (Sigma, St. Louis, MO, USA); antibodies specific for E-cadherin, N-cadherin, vimentin, Snail, MMP-2, MMP-9, Akt, p-AKt, PI3K, p-PI3K, and β-actin (β-actin) (Cell Signaling, Danvers, MA, USA); and HRP-labeled goat anti-rabbit immunoglobulin (Ig) G (Hangzhou Huaan Biotechnology Co., Ltd., Hangzhou, China).

    Techniques: Expressing, Western Blot, Standard Deviation

    ( A , C ) Changes in PI3K/AKt/Snail biomarker expression levels following treatment with 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B , D ) The band intensities of AKt, (p)-Akt, PI3K, (p)-PI3K and Snail relative to untreated control cells were quantified upon normalizing to β-actin expression, and are expressed as the mean ± standard deviation of three independent experiments.

    Journal: Molecules

    Article Title: 28-Hydroxy-3-oxoolean-12-en-29-oic Acid, a Triterpene Acid from Celastrus Orbiculatus Extract, Inhibits the Migration and Invasion of Human Gastric Cancer Cells In Vitro

    doi: 10.3390/molecules24193513

    Figure Lengend Snippet: ( A , C ) Changes in PI3K/AKt/Snail biomarker expression levels following treatment with 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B , D ) The band intensities of AKt, (p)-Akt, PI3K, (p)-PI3K and Snail relative to untreated control cells were quantified upon normalizing to β-actin expression, and are expressed as the mean ± standard deviation of three independent experiments.

    Article Snippet: Reagents The reagents used included the following: RPMI 1640 cell culture medium, trypsin (HyClone, USA); fetal bovine serum (FBS) (Gibco, Waltham, MA, USA); artificially reconstituted basement membrane, Transwell chambers (Corning, Corning, NY, USA); thiazole blue (MTT) powder (Sigma, St. Louis, MO, USA); antibodies specific for E-cadherin, N-cadherin, vimentin, Snail, MMP-2, MMP-9, Akt, p-AKt, PI3K, p-PI3K, and β-actin (β-actin) (Cell Signaling, Danvers, MA, USA); and HRP-labeled goat anti-rabbit immunoglobulin (Ig) G (Hangzhou Huaan Biotechnology Co., Ltd., Hangzhou, China).

    Techniques: Biomarker Assay, Expressing, Western Blot, Standard Deviation

    ( A , C ) Changes in epithelial–mesenchymal transition (EMT) biomarker expression levels following 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B , D ) The band intensities of E-cadherin, N-cadherin and vimentin relative to untreated control cells were quantified upon normalizing to β-actin expression, and are expressed as the mean ± standard deviation of three independent experiments.

    Journal: Molecules

    Article Title: 28-Hydroxy-3-oxoolean-12-en-29-oic Acid, a Triterpene Acid from Celastrus Orbiculatus Extract, Inhibits the Migration and Invasion of Human Gastric Cancer Cells In Vitro

    doi: 10.3390/molecules24193513

    Figure Lengend Snippet: ( A , C ) Changes in epithelial–mesenchymal transition (EMT) biomarker expression levels following 28-hydroxy-3-oxoolean-12-en-29-oic acid for 24 h were assessed by Western blotting. ( B , D ) The band intensities of E-cadherin, N-cadherin and vimentin relative to untreated control cells were quantified upon normalizing to β-actin expression, and are expressed as the mean ± standard deviation of three independent experiments.

    Article Snippet: Reagents The reagents used included the following: RPMI 1640 cell culture medium, trypsin (HyClone, USA); fetal bovine serum (FBS) (Gibco, Waltham, MA, USA); artificially reconstituted basement membrane, Transwell chambers (Corning, Corning, NY, USA); thiazole blue (MTT) powder (Sigma, St. Louis, MO, USA); antibodies specific for E-cadherin, N-cadherin, vimentin, Snail, MMP-2, MMP-9, Akt, p-AKt, PI3K, p-PI3K, and β-actin (β-actin) (Cell Signaling, Danvers, MA, USA); and HRP-labeled goat anti-rabbit immunoglobulin (Ig) G (Hangzhou Huaan Biotechnology Co., Ltd., Hangzhou, China).

    Techniques: Biomarker Assay, Expressing, Western Blot, Standard Deviation

    4-PG induces HO-1 mRNA and protein expression in macrophages and lung epithelial cells. (a) RAW 264.7 cells were treated with 4-PG at various concentrations (0, 5, 10, 20, and 40 μ M) for 8 h, and cell viability was determined by MTT assay. To evaluate the beneficial effect of 4-PG on HO-1 induction, cells were treated with 4-PG (0, 1, 5, and 10 μ M) at the indicated concentrations for 8 h. The mRNA and protein levels of HO-1 were measured by RT-PCR (b) and Western blotting (c). RAW 264.7 cells were treated with 4-PG (10 μ M) at the indicated time points (0, 2, 4, and 8 h). The mRNA and protein levels of HO-1 were determined by RT-PCR (d) and Western blotting (e). (f and g) PBMC and U937 cells were treated with 4-PG at the indicated concentrations (0, 1, 5, and 10 μ M) for 8 h. The mRNA and protein levels of HO-1 were determined by RT-PCR (top) and Western blotting (bottom). GAPDH and β -actin were used as internal controls. (h) RAW 264.7 cells were cotransduced with a pCignal Lenti-ARE reporter and pCignal Lenti-TK-Renilla. After treatment with 4-PG, luciferase activity was analyzed. The expression levels obtained from pCignal Lenti-ARE reporter-transduced cells without 4-PG treatment were normalized to 1. (i) RAW 264.7 cells were treated with 4-PG (10 μ M) for 8 h. Several antioxidative genes including TRX1, GCLC, and NQO1 were measured by RT-qPCR. (j and k) PBMC and U937 cells were pretreated with 4-PG (10 μ M) for 6 h followed by the stimulation of LPS (100 ng/ml) for another 4 h. (l) A549 cells pretreated with 4-PG (10 μ M) for 4 h and then stimulated with LPS (10 μ g/ml) for 6 h. The mRNA levels of HO-1, TNF- α , and IL-6 were determined by RT-PCR. Data were expressed as mean ± SD ( n = 5 determined in five independent experiments). One-way ANOVA with Turkey post hoc tests were performed; ∗ p

    Journal: Oxidative Medicine and Cellular Longevity

    Article Title: Pterostilbene 4′-β-Glucoside Attenuates LPS-Induced Acute Lung Injury via Induction of Heme Oxygenase-1

    doi: 10.1155/2018/2747018

    Figure Lengend Snippet: 4-PG induces HO-1 mRNA and protein expression in macrophages and lung epithelial cells. (a) RAW 264.7 cells were treated with 4-PG at various concentrations (0, 5, 10, 20, and 40 μ M) for 8 h, and cell viability was determined by MTT assay. To evaluate the beneficial effect of 4-PG on HO-1 induction, cells were treated with 4-PG (0, 1, 5, and 10 μ M) at the indicated concentrations for 8 h. The mRNA and protein levels of HO-1 were measured by RT-PCR (b) and Western blotting (c). RAW 264.7 cells were treated with 4-PG (10 μ M) at the indicated time points (0, 2, 4, and 8 h). The mRNA and protein levels of HO-1 were determined by RT-PCR (d) and Western blotting (e). (f and g) PBMC and U937 cells were treated with 4-PG at the indicated concentrations (0, 1, 5, and 10 μ M) for 8 h. The mRNA and protein levels of HO-1 were determined by RT-PCR (top) and Western blotting (bottom). GAPDH and β -actin were used as internal controls. (h) RAW 264.7 cells were cotransduced with a pCignal Lenti-ARE reporter and pCignal Lenti-TK-Renilla. After treatment with 4-PG, luciferase activity was analyzed. The expression levels obtained from pCignal Lenti-ARE reporter-transduced cells without 4-PG treatment were normalized to 1. (i) RAW 264.7 cells were treated with 4-PG (10 μ M) for 8 h. Several antioxidative genes including TRX1, GCLC, and NQO1 were measured by RT-qPCR. (j and k) PBMC and U937 cells were pretreated with 4-PG (10 μ M) for 6 h followed by the stimulation of LPS (100 ng/ml) for another 4 h. (l) A549 cells pretreated with 4-PG (10 μ M) for 4 h and then stimulated with LPS (10 μ g/ml) for 6 h. The mRNA levels of HO-1, TNF- α , and IL-6 were determined by RT-PCR. Data were expressed as mean ± SD ( n = 5 determined in five independent experiments). One-way ANOVA with Turkey post hoc tests were performed; ∗ p

    Article Snippet: The membrane was blocked with 5% nonfat milk in phosphate-buffered saline Tween 20 (PBS-T) and then incubated with a primary antibody against HO-1 (1 : 2000 v /v in PBS-T, Enzo Life Sciences, USA) or β -actin (1 : 2500 v /v in PBS-T, Cell Signaling Technology, MA) followed by incubation with a secondary antibody.

    Techniques: Expressing, MTT Assay, Reverse Transcription Polymerase Chain Reaction, Western Blot, Luciferase, Activity Assay, Quantitative RT-PCR

    4-PG prevents LPS-induced acute lung injury and upregulates HO-1 expression. (a) The scheme depicts the experimental protocol used to assess the protective effect of pterostilbene 4′-glucoside (4-PG) and pterostilbene (PTER) on LPS-induced ALI. 10-week-old mice were injected with 4-PG (10 mg/kg, i.p.) and PTER (10 mg/kg, i.p.) for 4 days prior to intranasal administration of LPS (2.5 mg/kg) for 24 h. (b) Chemical structures of 4-PG and PTER. (c) Lung sections were stained with hematoxylin and eosin (H E) for morphological evaluation, and the representative lung sections of each group are shown. Scale bar = 100 μ m. (left). Quantitative analysis of histologic lung section by lung injury score for six experimental groups. The score generates the average of two independent investigators (right). (d, e) The mRNA expression of proinflammatory cytokines and chemokines (TNF- α , IL-6, IL-1 β , CXCL1, and CXCL2) in lung tissues was detected by RT-PCR. Furthermore, mRNA and protein levels of HO-1 were assessed by RT-PCR (f, left: HO-1mRNA levels, right: quantification of the relative band density) and Western blotting (g, left: HO-1 protein levels, right: quantification of the relative band density) from lung tissues, respectively. 18S and β -actin were used as internal controls. Data were expressed as mean ± SD ( n = 5 per group); ∗∗ p

    Journal: Oxidative Medicine and Cellular Longevity

    Article Title: Pterostilbene 4′-β-Glucoside Attenuates LPS-Induced Acute Lung Injury via Induction of Heme Oxygenase-1

    doi: 10.1155/2018/2747018

    Figure Lengend Snippet: 4-PG prevents LPS-induced acute lung injury and upregulates HO-1 expression. (a) The scheme depicts the experimental protocol used to assess the protective effect of pterostilbene 4′-glucoside (4-PG) and pterostilbene (PTER) on LPS-induced ALI. 10-week-old mice were injected with 4-PG (10 mg/kg, i.p.) and PTER (10 mg/kg, i.p.) for 4 days prior to intranasal administration of LPS (2.5 mg/kg) for 24 h. (b) Chemical structures of 4-PG and PTER. (c) Lung sections were stained with hematoxylin and eosin (H E) for morphological evaluation, and the representative lung sections of each group are shown. Scale bar = 100 μ m. (left). Quantitative analysis of histologic lung section by lung injury score for six experimental groups. The score generates the average of two independent investigators (right). (d, e) The mRNA expression of proinflammatory cytokines and chemokines (TNF- α , IL-6, IL-1 β , CXCL1, and CXCL2) in lung tissues was detected by RT-PCR. Furthermore, mRNA and protein levels of HO-1 were assessed by RT-PCR (f, left: HO-1mRNA levels, right: quantification of the relative band density) and Western blotting (g, left: HO-1 protein levels, right: quantification of the relative band density) from lung tissues, respectively. 18S and β -actin were used as internal controls. Data were expressed as mean ± SD ( n = 5 per group); ∗∗ p

    Article Snippet: The membrane was blocked with 5% nonfat milk in phosphate-buffered saline Tween 20 (PBS-T) and then incubated with a primary antibody against HO-1 (1 : 2000 v /v in PBS-T, Enzo Life Sciences, USA) or β -actin (1 : 2500 v /v in PBS-T, Cell Signaling Technology, MA) followed by incubation with a secondary antibody.

    Techniques: Expressing, Mouse Assay, Injection, Staining, Reverse Transcription Polymerase Chain Reaction, Western Blot

    IL-5 and CD40L stimulation expands a population of B cells enriched for FasL expression. ( A ) Diagram of B cell co-culture experiments with CD40L-expressing fibroblasts for study of the effects of IL-5 on B cell growth and function. ( B ) CD19 + B cells were cultured as illustrated in (A) and harvested after five days. Viable B cells recovered from culture were quantified from replicate samples using a hemocytometer and trypan blue exclusion. The dotted line represents the concentration of B cells at the beginning of the experiment. ( C ) Proliferation of B cells cultured with CD40L-expressing fibroblasts in the presence or absence of IL-5 was assessed by the incorporation of 3 H-thymidine. ( D ) B cells stimulated with CD40L in the presence or absence of IL-5 were stained for FasL, CD5, and CD1d and compared with freshly-isolated CD19 + B cells. ( E ) Median fluorescence intensity of surface markers stained as in (D) on freshly-isolated CD19 + B cells, B cells stimulated with CD40L, and B cells stimulated with CD40L and IL-5. ( F ) The frequency of FasL + B cells among freshly-isolated CD19 + B cells and B cells stimulated with CD40L and IL-5 was measured by flow cytometry as in (D). ( G ) Cell lysates from equal numbers of freshly-isolated CD19 + B cells and IL-5-stimulated B cells were probed by immunoblot for FasL and β-Actin. Data are representative of more than four independent experiments. *** p

    Journal: PLoS ONE

    Article Title: Interleukin-5 Supports the Expansion of Fas Ligand-Expressing Killer B Cells that Induce Antigen-Specific Apoptosis of CD4+ T Cells and Secrete Interleukin-10

    doi: 10.1371/journal.pone.0070131

    Figure Lengend Snippet: IL-5 and CD40L stimulation expands a population of B cells enriched for FasL expression. ( A ) Diagram of B cell co-culture experiments with CD40L-expressing fibroblasts for study of the effects of IL-5 on B cell growth and function. ( B ) CD19 + B cells were cultured as illustrated in (A) and harvested after five days. Viable B cells recovered from culture were quantified from replicate samples using a hemocytometer and trypan blue exclusion. The dotted line represents the concentration of B cells at the beginning of the experiment. ( C ) Proliferation of B cells cultured with CD40L-expressing fibroblasts in the presence or absence of IL-5 was assessed by the incorporation of 3 H-thymidine. ( D ) B cells stimulated with CD40L in the presence or absence of IL-5 were stained for FasL, CD5, and CD1d and compared with freshly-isolated CD19 + B cells. ( E ) Median fluorescence intensity of surface markers stained as in (D) on freshly-isolated CD19 + B cells, B cells stimulated with CD40L, and B cells stimulated with CD40L and IL-5. ( F ) The frequency of FasL + B cells among freshly-isolated CD19 + B cells and B cells stimulated with CD40L and IL-5 was measured by flow cytometry as in (D). ( G ) Cell lysates from equal numbers of freshly-isolated CD19 + B cells and IL-5-stimulated B cells were probed by immunoblot for FasL and β-Actin. Data are representative of more than four independent experiments. *** p

    Article Snippet: Membranes were blocked and incubated with polyclonal rabbit anti-FasL IgG (Millipore) or anti-β-Actin (Cell Signaling).

    Techniques: Expressing, Co-Culture Assay, Cell Culture, Concentration Assay, Staining, Isolation, Fluorescence, Flow Cytometry, Cytometry

    IL-5-mediated induction of killer B cell function, but not FasL expression, is inhibited by IL-4. CD19 + B cells from naïve mice were cultured for five days with CD40L Fb and IL-4, IL-5, or both cytokines concurrently. ( A ) The ability of B cells stimulated under these conditions to induce apoptosis in activated CD4 + T cells was then assessed as in Figure 6 . Apoptotic cells were identified and enumerated (mean ± SEM) on the basis of positive staining for Annexin V by flow cytometry. ( B ) B cells were cultured with CD40L-expressing fibroblasts in the presence or absence of IL-4 and IL-5, or in the presence of both cytokines. Cells were harvested after 5 days in culture and cell lysates were probed for FasL and β-Actin proteins as in Figure 5G . *** p

    Journal: PLoS ONE

    Article Title: Interleukin-5 Supports the Expansion of Fas Ligand-Expressing Killer B Cells that Induce Antigen-Specific Apoptosis of CD4+ T Cells and Secrete Interleukin-10

    doi: 10.1371/journal.pone.0070131

    Figure Lengend Snippet: IL-5-mediated induction of killer B cell function, but not FasL expression, is inhibited by IL-4. CD19 + B cells from naïve mice were cultured for five days with CD40L Fb and IL-4, IL-5, or both cytokines concurrently. ( A ) The ability of B cells stimulated under these conditions to induce apoptosis in activated CD4 + T cells was then assessed as in Figure 6 . Apoptotic cells were identified and enumerated (mean ± SEM) on the basis of positive staining for Annexin V by flow cytometry. ( B ) B cells were cultured with CD40L-expressing fibroblasts in the presence or absence of IL-4 and IL-5, or in the presence of both cytokines. Cells were harvested after 5 days in culture and cell lysates were probed for FasL and β-Actin proteins as in Figure 5G . *** p

    Article Snippet: Membranes were blocked and incubated with polyclonal rabbit anti-FasL IgG (Millipore) or anti-β-Actin (Cell Signaling).

    Techniques: Cell Function Assay, Expressing, Mouse Assay, Cell Culture, Staining, Flow Cytometry, Cytometry

    Fluorescence activated cell sorting (FACS) analysis of stained cells and validation of cell cycle sorted populations in HDFa, NCI-H295R and HeLa cells. Panels a - c : FACS analysis shows cell cycle distribution of stained cells before (first image) and after sorting to G1, S and G2 phases (second, third and fourth images, respectively; Panel a : HDFa, Panel b : NCI-H295R, Panel c : HeLa. Intervals of fluorescence intensity (shown as vertical bands) were defined to gate G1, S and G2 phases, respectively. Panels d-e : Western blot analysis confirms the high efficacy of cell cycle sort (Panel d : NCI-H295R, Panel e : HeLa). Density values of phospho(Tyr15)-CDC-2 were first normalized to β-actin loading control and were further normalized to G1 phase (displayed above each band)

    Journal: BMC Genomics

    Article Title: Fluorescence activated cell sorting followed by small RNA sequencing reveals stable microRNA expression during cell cycle progression

    doi: 10.1186/s12864-016-2747-6

    Figure Lengend Snippet: Fluorescence activated cell sorting (FACS) analysis of stained cells and validation of cell cycle sorted populations in HDFa, NCI-H295R and HeLa cells. Panels a - c : FACS analysis shows cell cycle distribution of stained cells before (first image) and after sorting to G1, S and G2 phases (second, third and fourth images, respectively; Panel a : HDFa, Panel b : NCI-H295R, Panel c : HeLa. Intervals of fluorescence intensity (shown as vertical bands) were defined to gate G1, S and G2 phases, respectively. Panels d-e : Western blot analysis confirms the high efficacy of cell cycle sort (Panel d : NCI-H295R, Panel e : HeLa). Density values of phospho(Tyr15)-CDC-2 were first normalized to β-actin loading control and were further normalized to G1 phase (displayed above each band)

    Article Snippet: Thereafter membranes were stripped with mild stripping buffer (0.2 M glycine, 0.1 % sodium dodecyl sulfate, 0.1 % Tween-20, pH = 2.2) by gentle agitation for 45 min at room temperature, and were blocked again for subsequent detection of loading control β-actin (Cell Signaling Technology, cat. No.: 4967, dilution: 1:2000).

    Techniques: Fluorescence, FACS, Staining, Western Blot

    DENV infection activates cGAS-dependent antiviral responses. ( A ) In vitro assay for functional cGAS expression in A549 and THP-1 cell lysates as indicated by the production of cGAMP in the presence of 1 μg/ml poly(dA:dT). Results demonstrate that cGAS is functional in A549 as it is in THP-1, as shown previously 3 . ( B ) Western blot analysis of cGAS expression in A549 and THP-1 cells (control). THP-1 cells used as positive control and not as point of comparison for cGAS expression levels between A549 and THP-1 cell lines. ( C ) cGAMP production in A549 cells that are uninfected or infected with DENV2-PDK53 at MOI 1 for 24 hrs. Absorbance values normalized by cell count of each group. Results shown are representative of at least three independent experiments. ( D , E ) DENV RNA levels in infected A549 cells that were pre-treated ( D ) or post-treated ( E ) with 1 μg/ml cGAMP or 250 units/ml IFNα. Results are normalized by GAPDH expression and represent mean ± SD of at least four independent experiments. ( F ) DENV RNA levels in cGAS knockdown (si-cGAS) A549 cells compared to siRNA scrambled control (si-control) when infected with DENV2-PDK53 at MOI 1 for 24 hrs, assessed by RT-qPCR and normalized by GAPDH expression. Western blot confirmation of decreased cGAS expression in si-cGAS A549 cells, normalized with β-actin. Data is represented as mean ± SD of at least three independent experiments. In this figure, *depicts P

    Journal: Scientific Reports

    Article Title: Dengue virus activates cGAS through the release of mitochondrial DNA

    doi: 10.1038/s41598-017-03932-1

    Figure Lengend Snippet: DENV infection activates cGAS-dependent antiviral responses. ( A ) In vitro assay for functional cGAS expression in A549 and THP-1 cell lysates as indicated by the production of cGAMP in the presence of 1 μg/ml poly(dA:dT). Results demonstrate that cGAS is functional in A549 as it is in THP-1, as shown previously 3 . ( B ) Western blot analysis of cGAS expression in A549 and THP-1 cells (control). THP-1 cells used as positive control and not as point of comparison for cGAS expression levels between A549 and THP-1 cell lines. ( C ) cGAMP production in A549 cells that are uninfected or infected with DENV2-PDK53 at MOI 1 for 24 hrs. Absorbance values normalized by cell count of each group. Results shown are representative of at least three independent experiments. ( D , E ) DENV RNA levels in infected A549 cells that were pre-treated ( D ) or post-treated ( E ) with 1 μg/ml cGAMP or 250 units/ml IFNα. Results are normalized by GAPDH expression and represent mean ± SD of at least four independent experiments. ( F ) DENV RNA levels in cGAS knockdown (si-cGAS) A549 cells compared to siRNA scrambled control (si-control) when infected with DENV2-PDK53 at MOI 1 for 24 hrs, assessed by RT-qPCR and normalized by GAPDH expression. Western blot confirmation of decreased cGAS expression in si-cGAS A549 cells, normalized with β-actin. Data is represented as mean ± SD of at least three independent experiments. In this figure, *depicts P

    Article Snippet: Primary antibodies included human cGAS (Sigma-Aldrich HPA031700, 1:500) and human β-actin (Cell Signaling 8H10D10, 1:1000).

    Techniques: Infection, In Vitro, Functional Assay, Expressing, Western Blot, Positive Control, Cell Counting, Quantitative RT-PCR

    GSK2606414 inhibits GANT-61 induced cell autophagy in MYCN amplified NB cells ( A ) Assessment of LC3 conversion by LC3 immunoblotting. Membranes were reprobed with β-actin antibody. Four cell treatments CON (non-treatment), GANT-61 (10 μM GANT-61 treatment 48 h), GSK2606414(0.5 μM GSK2606414 treatment 3 h), GSK2606414+GANT-61 (0.5 μM GSK2606414 pretreatment 3 h with 10 μM GANT-61 treatment 48 h) were tested in NBL-W-S and SK-N-AS cells ( B ) The LC3-II/β-ACTIN ratio was plotted as histogram (mean ± SD), * P

    Journal: Oncotarget

    Article Title: The protective autophagy activated by GANT-61 in MYCN amplified neuroblastoma cells is mediated by PERK

    doi: 10.18632/oncotarget.24214

    Figure Lengend Snippet: GSK2606414 inhibits GANT-61 induced cell autophagy in MYCN amplified NB cells ( A ) Assessment of LC3 conversion by LC3 immunoblotting. Membranes were reprobed with β-actin antibody. Four cell treatments CON (non-treatment), GANT-61 (10 μM GANT-61 treatment 48 h), GSK2606414(0.5 μM GSK2606414 treatment 3 h), GSK2606414+GANT-61 (0.5 μM GSK2606414 pretreatment 3 h with 10 μM GANT-61 treatment 48 h) were tested in NBL-W-S and SK-N-AS cells ( B ) The LC3-II/β-ACTIN ratio was plotted as histogram (mean ± SD), * P

    Article Snippet: Antibodies and reagents β-ACTIN, CLEAVED-CASPASE3, PERK, phospho-eIF2α, ATF4 rabbit antibodies were purchased from Cell Signaling Technology, (USA).

    Techniques: Amplification

    Assessment of GSK2606414 cytotoxicity and inhibitory on NB cell lines ( A ) NBL-W-S and SK-N-AS NB cells were treated with 10μM GANT-61 during 48 h. Western blots evaluated protein expression of PERK, P-eIF2α, ATF4. The PERK/β-ACTIN, P-eIF2α/β-ACTIN, and the ATF4/β-ACTIN ratio. The results are presented as mean ± SD of three independent experiments. * P

    Journal: Oncotarget

    Article Title: The protective autophagy activated by GANT-61 in MYCN amplified neuroblastoma cells is mediated by PERK

    doi: 10.18632/oncotarget.24214

    Figure Lengend Snippet: Assessment of GSK2606414 cytotoxicity and inhibitory on NB cell lines ( A ) NBL-W-S and SK-N-AS NB cells were treated with 10μM GANT-61 during 48 h. Western blots evaluated protein expression of PERK, P-eIF2α, ATF4. The PERK/β-ACTIN, P-eIF2α/β-ACTIN, and the ATF4/β-ACTIN ratio. The results are presented as mean ± SD of three independent experiments. * P

    Article Snippet: Antibodies and reagents β-ACTIN, CLEAVED-CASPASE3, PERK, phospho-eIF2α, ATF4 rabbit antibodies were purchased from Cell Signaling Technology, (USA).

    Techniques: Western Blot, Expressing

    Activation of mitochondria-dependent apoptosis pathway and inhibition of EGFR-mediated PI3K/AKT and Ras/Raf/MEK/ERK pathways by GE11-Ori-Se NPs. (A) Effects of GE11-Ori-Se NPs, and the same dosage of oridonin or Chi-Se NPs on the production of ROS in KYSE-150 cells, n = 3. (B) Western blot analysis for the expression of Bcl-2 and Bax in KYSE-150 cells after GE11-Ori-Se NPs treatment. β-Actin was used as loading control. (C) Effects of GE11-Ori-Se NPs on the caspase-3 and caspase-9 activity in KYSE-150 cells, n = 3,** p

    Journal: Drug Delivery

    Article Title: GE11 peptide conjugated selenium nanoparticles for EGFR targeted oridonin delivery to achieve enhanced anticancer efficacy by inhibiting EGFR-mediated PI3K/AKT and Ras/Raf/MEK/ERK pathways

    doi: 10.1080/10717544.2017.1386729

    Figure Lengend Snippet: Activation of mitochondria-dependent apoptosis pathway and inhibition of EGFR-mediated PI3K/AKT and Ras/Raf/MEK/ERK pathways by GE11-Ori-Se NPs. (A) Effects of GE11-Ori-Se NPs, and the same dosage of oridonin or Chi-Se NPs on the production of ROS in KYSE-150 cells, n = 3. (B) Western blot analysis for the expression of Bcl-2 and Bax in KYSE-150 cells after GE11-Ori-Se NPs treatment. β-Actin was used as loading control. (C) Effects of GE11-Ori-Se NPs on the caspase-3 and caspase-9 activity in KYSE-150 cells, n = 3,** p

    Article Snippet: RIPA lysis buffer, Bcl-2 antibody, Bax antibody, EGFR antibody, p-EGFR antibody, Ras antibody, Raf antibody, MEK antibody, p-MEK antibody, ERK antibody, p-ERK antibody, PI3K antibody, p-PI3K antibody, AKT antibody, p-AKT antibody, β-actin antibody, anti-mouse IgG and anti-rabbit IgG were from Cell Signaling Technology (Danvers, MA).

    Techniques: Activation Assay, Inhibition, Western Blot, Expressing, Activity Assay

    Triple-drug (LY294002, PLX4720 and lexatumumab) and dual-drug combinations (LY294002 and lexatumumab) triggered the intrinsic and extrinsic apoptotic pathways in 8505c and SW1736 cells. 8505c and SW1736 cells were treated for 8 h with drug combinations as indicated in the figure and analyzed for pro- and anti-apoptotic proteins by western blot. ( a ) Treatment with triple-drug combination in 8505c and the dual combination in SW1736 resulted in cleavage of caspase 8/9/3 and PARP. ( b ) Triple-drug combination resulted in the increase in pro-apoptotic proteins Bad, Bim, Bax and Bak in 8505c cells. Dual inhibitor combination, on the other hand, resulted in an increase in BimL  S forms in SW1736 cells. ( c ) Triple-drug combination significantly decreased the anti-apoptotic proteins Bcl-xL and Mcl-1 in 8505c cells at 8 h of treatment. ( d ) cFLIP decreased in both the dual and triple-drug combinational treatments of 8505c cells. ( e ) Treatment of 8505c cells at different time points up to 48 h with the triple-drug combination also showed a significant decrease in anti- apoptotic proteins, Bcl-xL, Mcl-1, cFlip and an increase in pro-apoptotic proteins, Bax and Bim isoforms.  β -Actin was used as loading control

    Journal: Cell Death & Disease

    Article Title: Blocks to thyroid cancer cell apoptosis can be overcome by inhibition of the MAPK and PI3K/AKT pathways

    doi: 10.1038/cddis.2014.78

    Figure Lengend Snippet: Triple-drug (LY294002, PLX4720 and lexatumumab) and dual-drug combinations (LY294002 and lexatumumab) triggered the intrinsic and extrinsic apoptotic pathways in 8505c and SW1736 cells. 8505c and SW1736 cells were treated for 8 h with drug combinations as indicated in the figure and analyzed for pro- and anti-apoptotic proteins by western blot. ( a ) Treatment with triple-drug combination in 8505c and the dual combination in SW1736 resulted in cleavage of caspase 8/9/3 and PARP. ( b ) Triple-drug combination resulted in the increase in pro-apoptotic proteins Bad, Bim, Bax and Bak in 8505c cells. Dual inhibitor combination, on the other hand, resulted in an increase in BimL S forms in SW1736 cells. ( c ) Triple-drug combination significantly decreased the anti-apoptotic proteins Bcl-xL and Mcl-1 in 8505c cells at 8 h of treatment. ( d ) cFLIP decreased in both the dual and triple-drug combinational treatments of 8505c cells. ( e ) Treatment of 8505c cells at different time points up to 48 h with the triple-drug combination also showed a significant decrease in anti- apoptotic proteins, Bcl-xL, Mcl-1, cFlip and an increase in pro-apoptotic proteins, Bax and Bim isoforms. β -Actin was used as loading control

    Article Snippet: Antibodies – β -actin, phospho-p44/42 MAPK (ERK1/2) (Thr202/Tyr204) (phospho-ERK), p44/42 MAPK (ERK1/2) (Total ERK), total Akt, phospho-Akt (Ser473 ), caspases 3,8,9, cleaved caspase-3, PARP, c-FLIP, TRAIL-R2, Bcl2, Bcl-xL, Mcl-1, Bim, Bid, Bax, Bad, Bak were purchased from Cell Signaling Technology.

    Techniques: Western Blot

    Knockdown of Bcl-xL sensitized 8505c cells to lexatumumab-induced apoptosis. Cells were transfected with 50-nM Bcl-xL or control siRNA for 48 h and then treated with 1000 ng/ml of lexatumumab for another 24 h. ( a ) Bcl-xL protein level significantly decreased after 48 h of transfection with Bcl-xL siRNA.  β -Actin served as a loading control. ( b ) Treatment of Bcl-xL knockdown cells with lexatumumab showed a significant increase in cell death (52.3%,  P

    Journal: Cell Death & Disease

    Article Title: Blocks to thyroid cancer cell apoptosis can be overcome by inhibition of the MAPK and PI3K/AKT pathways

    doi: 10.1038/cddis.2014.78

    Figure Lengend Snippet: Knockdown of Bcl-xL sensitized 8505c cells to lexatumumab-induced apoptosis. Cells were transfected with 50-nM Bcl-xL or control siRNA for 48 h and then treated with 1000 ng/ml of lexatumumab for another 24 h. ( a ) Bcl-xL protein level significantly decreased after 48 h of transfection with Bcl-xL siRNA. β -Actin served as a loading control. ( b ) Treatment of Bcl-xL knockdown cells with lexatumumab showed a significant increase in cell death (52.3%, P

    Article Snippet: Antibodies – β -actin, phospho-p44/42 MAPK (ERK1/2) (Thr202/Tyr204) (phospho-ERK), p44/42 MAPK (ERK1/2) (Total ERK), total Akt, phospho-Akt (Ser473 ), caspases 3,8,9, cleaved caspase-3, PARP, c-FLIP, TRAIL-R2, Bcl2, Bcl-xL, Mcl-1, Bim, Bid, Bax, Bad, Bak were purchased from Cell Signaling Technology.

    Techniques: Transfection

    TAM tyrosine kinase receptors are novel ubiquitylation targets of Cbl-b. a, Out of 9000 human proteins tested, Tyro3 had the highest Cbl-b mediated ubiquitylation signal. Signal intensities are shown for the corresponding Tyro3 spots in the protein arrays incubated with the E2 enzyme without Cbl-b, the Cbl-b C373A mutant, and wild-type Cbl-b proteins (in duplicates). Data are shown as mean values. RFU: relative fluorescent units. b, In vitro ubiquitylation of recombinant Flag-tagged Tyro3, Axl, and Mer in the presence (left panel) and absence (right panel) of Cbl-b. Blots were probed with anti-Flag Abs. c, Immunoprecipitation showing time-dependent recruitment of Cbl-b to TAM tyrosine kinase receptors in HeLa cells upon stimulation with His-tagged Gas6 (upper panel). Input levels of Cbl-b and β-actin are shown as controls. d , Gas6-induced Axl ubiquitylation depends on Cbl-b expression. HeLa cells were transfected with scrambled siControl or siCbl-b and then stimulated with 450ng/ml Gas6 for 15 minutes or left untreated. Lysates were immunoprecipiated with anti-Axl antibodies. Blots were then probed using anti-Ubiquitin and anti-Axl Abs. The location of the mature form of Axl (~140kDa) is shown with an arrow. Axl, Cbl-b, and tubulin protein levels are shown to control for the input (lower panels). e, Representative histograms showing equal expression of Tyro3, Axl, and Mer receptors at the cell surface of freshly isolated NK1.1 + CD3ε - splenic Cbl-b +/+ Cbl-b -/- and C373A KI/KI NK cells. Background staining with control isotype Abs is shown for each blot in grey. f, In vitro proliferation of Cbl-b +/- and Cbl-b -/- NK cells stimulated with anti-NKG2D Abs (30μg/ml) in the presence of different concentrations of Gas6 (mean values ± s.e.m., n=4). *P

    Journal: Nature

    Article Title: The E3 Ligase Cbl-b and TAM receptors regulate cancer metastasis via natural killer cells

    doi: 10.1038/nature12998

    Figure Lengend Snippet: TAM tyrosine kinase receptors are novel ubiquitylation targets of Cbl-b. a, Out of 9000 human proteins tested, Tyro3 had the highest Cbl-b mediated ubiquitylation signal. Signal intensities are shown for the corresponding Tyro3 spots in the protein arrays incubated with the E2 enzyme without Cbl-b, the Cbl-b C373A mutant, and wild-type Cbl-b proteins (in duplicates). Data are shown as mean values. RFU: relative fluorescent units. b, In vitro ubiquitylation of recombinant Flag-tagged Tyro3, Axl, and Mer in the presence (left panel) and absence (right panel) of Cbl-b. Blots were probed with anti-Flag Abs. c, Immunoprecipitation showing time-dependent recruitment of Cbl-b to TAM tyrosine kinase receptors in HeLa cells upon stimulation with His-tagged Gas6 (upper panel). Input levels of Cbl-b and β-actin are shown as controls. d , Gas6-induced Axl ubiquitylation depends on Cbl-b expression. HeLa cells were transfected with scrambled siControl or siCbl-b and then stimulated with 450ng/ml Gas6 for 15 minutes or left untreated. Lysates were immunoprecipiated with anti-Axl antibodies. Blots were then probed using anti-Ubiquitin and anti-Axl Abs. The location of the mature form of Axl (~140kDa) is shown with an arrow. Axl, Cbl-b, and tubulin protein levels are shown to control for the input (lower panels). e, Representative histograms showing equal expression of Tyro3, Axl, and Mer receptors at the cell surface of freshly isolated NK1.1 + CD3ε - splenic Cbl-b +/+ Cbl-b -/- and C373A KI/KI NK cells. Background staining with control isotype Abs is shown for each blot in grey. f, In vitro proliferation of Cbl-b +/- and Cbl-b -/- NK cells stimulated with anti-NKG2D Abs (30μg/ml) in the presence of different concentrations of Gas6 (mean values ± s.e.m., n=4). *P

    Article Snippet: L. Prieto) antibodies and as loading control anti-β-actin, anti-GAPDH, or anti-β tubulin (Cell Signaling Technology) according to standard procedures.

    Techniques: Incubation, Mutagenesis, In Vitro, Recombinant, Immunoprecipitation, Expressing, Transfection, Isolation, Staining

    HBxΔ127 has a greater capacity to stimulate SREBP-1c relative to wild type HBx. (A, B) HepG2-X (or H7402-X) cells and HepG2-XΔ127 (or H7402-XΔ127) cells were transfected with pSilencer 3.0-X plasmid for 48 h. (A) The mRNA levels of SREBP-1c were detected by RT-PCR. (B) Immunoblot analysis to detect SREBP-1c (top panel) and β-actin protein levels (bottom panel). (C, D) HepG2-X (or H7402-X) cells and HepG2-XΔ127 (or H7402-XΔ127) cells were co-transfected with or without pSilencer 3.0-X plasmid and SREBP-1c-571-Luc-WT. Data are representative of 3 independent experiments. Values represent means±SD. n =3. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: A mutant of HBx (HBxΔ127) promotes hepatoma cell growth via sterol regulatory element binding protein 1c involving 5-lipoxygenase

    doi: 10.1038/aps.2010.5

    Figure Lengend Snippet: HBxΔ127 has a greater capacity to stimulate SREBP-1c relative to wild type HBx. (A, B) HepG2-X (or H7402-X) cells and HepG2-XΔ127 (or H7402-XΔ127) cells were transfected with pSilencer 3.0-X plasmid for 48 h. (A) The mRNA levels of SREBP-1c were detected by RT-PCR. (B) Immunoblot analysis to detect SREBP-1c (top panel) and β-actin protein levels (bottom panel). (C, D) HepG2-X (or H7402-X) cells and HepG2-XΔ127 (or H7402-XΔ127) cells were co-transfected with or without pSilencer 3.0-X plasmid and SREBP-1c-571-Luc-WT. Data are representative of 3 independent experiments. Values represent means±SD. n =3. b P

    Article Snippet: For protein loading controls, the amount of β-actin protein was also determined using a β-actin-specific antibody (Cell Signaling Technology, Danvers, MA, USA).

    Techniques: Transfection, Plasmid Preparation, Reverse Transcription Polymerase Chain Reaction