anisomycin  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anisomycin
    Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. <t>Anisomycin</t> was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.
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    1) Product Images from "Dioscin Exerts Protective Effects Against Crystalline Silica-induced Pulmonary Fibrosis in Mice"

    Article Title: Dioscin Exerts Protective Effects Against Crystalline Silica-induced Pulmonary Fibrosis in Mice

    Journal: Theranostics

    doi: 10.7150/thno.20270

    Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. Anisomycin was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.
    Figure Legend Snippet: Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. Anisomycin was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.

    Techniques Used: Western Blot, Enzyme-linked Immunosorbent Assay

    Dioscin regulated the TGF-β1-Samd3 signaling pathway and regulatory T cell functions. (A, B) Western blot analysis and quantification of TGF-β1 in lungs of dioscin-treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of TGF-β1 levels relative to β-Actin is shown below the band. (C) qPCR analysis of Tgf-β1 in the lung tissues (n=6-8 per group). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed in triplicate. (D, E) Western blot analysis and quantification of Smad3 signaling in the lungs of treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (F, G) Western blot analysis of p-Smad3 and Smad3 in NIH-3T3 fibroblast, 30 min after TGF-β1 (5 ng/mL) and dioscin treatment. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (H) Schematic design of transwell experiment: RAW 264.7 macrophages were primed by LPS (25 ng/mL) for 3 h, and then treated with CS (50 μg/cm 2 ) together with dioscin or anisomycin (2 ng/mL) for 24 h in the lower plate, while NIH-3T3 fibroblasts were attached in the upper chamber. After that, the upper chamber was inserted back to the lower plate, and macrophages and fibroblasts were co-cultured for another 24 h. Then, the mRNA expressions of Col1a1 (I) and α-SMA (J) in NIH-3T3 fibroblast were determined by qPCR (n=3). (K, L) Percentage of CD4+CD25+FoxP3+ cells (regulatory T cells) in hilar lymph nodes was assayed by FACS analysis (n=5). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD.
    Figure Legend Snippet: Dioscin regulated the TGF-β1-Samd3 signaling pathway and regulatory T cell functions. (A, B) Western blot analysis and quantification of TGF-β1 in lungs of dioscin-treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of TGF-β1 levels relative to β-Actin is shown below the band. (C) qPCR analysis of Tgf-β1 in the lung tissues (n=6-8 per group). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed in triplicate. (D, E) Western blot analysis and quantification of Smad3 signaling in the lungs of treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (F, G) Western blot analysis of p-Smad3 and Smad3 in NIH-3T3 fibroblast, 30 min after TGF-β1 (5 ng/mL) and dioscin treatment. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (H) Schematic design of transwell experiment: RAW 264.7 macrophages were primed by LPS (25 ng/mL) for 3 h, and then treated with CS (50 μg/cm 2 ) together with dioscin or anisomycin (2 ng/mL) for 24 h in the lower plate, while NIH-3T3 fibroblasts were attached in the upper chamber. After that, the upper chamber was inserted back to the lower plate, and macrophages and fibroblasts were co-cultured for another 24 h. Then, the mRNA expressions of Col1a1 (I) and α-SMA (J) in NIH-3T3 fibroblast were determined by qPCR (n=3). (K, L) Percentage of CD4+CD25+FoxP3+ cells (regulatory T cells) in hilar lymph nodes was assayed by FACS analysis (n=5). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD.

    Techniques Used: Western Blot, Cell Culture

    anisomycin  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anisomycin
    Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. <t>Anisomycin</t> was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.
    Anisomycin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Dioscin Exerts Protective Effects Against Crystalline Silica-induced Pulmonary Fibrosis in Mice"

    Article Title: Dioscin Exerts Protective Effects Against Crystalline Silica-induced Pulmonary Fibrosis in Mice

    Journal: Theranostics

    doi: 10.7150/thno.20270

    Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. Anisomycin was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.
    Figure Legend Snippet: Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. Anisomycin was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.

    Techniques Used: Western Blot, Enzyme-linked Immunosorbent Assay

    Dioscin regulated the TGF-β1-Samd3 signaling pathway and regulatory T cell functions. (A, B) Western blot analysis and quantification of TGF-β1 in lungs of dioscin-treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of TGF-β1 levels relative to β-Actin is shown below the band. (C) qPCR analysis of Tgf-β1 in the lung tissues (n=6-8 per group). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed in triplicate. (D, E) Western blot analysis and quantification of Smad3 signaling in the lungs of treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (F, G) Western blot analysis of p-Smad3 and Smad3 in NIH-3T3 fibroblast, 30 min after TGF-β1 (5 ng/mL) and dioscin treatment. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (H) Schematic design of transwell experiment: RAW 264.7 macrophages were primed by LPS (25 ng/mL) for 3 h, and then treated with CS (50 μg/cm 2 ) together with dioscin or anisomycin (2 ng/mL) for 24 h in the lower plate, while NIH-3T3 fibroblasts were attached in the upper chamber. After that, the upper chamber was inserted back to the lower plate, and macrophages and fibroblasts were co-cultured for another 24 h. Then, the mRNA expressions of Col1a1 (I) and α-SMA (J) in NIH-3T3 fibroblast were determined by qPCR (n=3). (K, L) Percentage of CD4+CD25+FoxP3+ cells (regulatory T cells) in hilar lymph nodes was assayed by FACS analysis (n=5). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD.
    Figure Legend Snippet: Dioscin regulated the TGF-β1-Samd3 signaling pathway and regulatory T cell functions. (A, B) Western blot analysis and quantification of TGF-β1 in lungs of dioscin-treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of TGF-β1 levels relative to β-Actin is shown below the band. (C) qPCR analysis of Tgf-β1 in the lung tissues (n=6-8 per group). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed in triplicate. (D, E) Western blot analysis and quantification of Smad3 signaling in the lungs of treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (F, G) Western blot analysis of p-Smad3 and Smad3 in NIH-3T3 fibroblast, 30 min after TGF-β1 (5 ng/mL) and dioscin treatment. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (H) Schematic design of transwell experiment: RAW 264.7 macrophages were primed by LPS (25 ng/mL) for 3 h, and then treated with CS (50 μg/cm 2 ) together with dioscin or anisomycin (2 ng/mL) for 24 h in the lower plate, while NIH-3T3 fibroblasts were attached in the upper chamber. After that, the upper chamber was inserted back to the lower plate, and macrophages and fibroblasts were co-cultured for another 24 h. Then, the mRNA expressions of Col1a1 (I) and α-SMA (J) in NIH-3T3 fibroblast were determined by qPCR (n=3). (K, L) Percentage of CD4+CD25+FoxP3+ cells (regulatory T cells) in hilar lymph nodes was assayed by FACS analysis (n=5). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD.

    Techniques Used: Western Blot, Cell Culture

    p38 agonist anisomycin  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc p38 agonist anisomycin
    The effects of the reagents on CCD-induced mechanical allodynia. (a–d) The PWMTs of CCD rats (4 days after operation) 1, 2, 4, and 8 h after RR, 4 α -PDD, SB203580, and <t>anisomycin</t> intrathecal injections ( n = 6; the data are expressed as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared ipsilaterally with the saline group; one-way ANOVA followed by Tukey's post hoc test.
    P38 Agonist Anisomycin, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Images

    1) Product Images from "Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion"

    Article Title: Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion

    Journal: BioMed Research International

    doi: 10.1155/2016/6978923

    The effects of the reagents on CCD-induced mechanical allodynia. (a–d) The PWMTs of CCD rats (4 days after operation) 1, 2, 4, and 8 h after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections ( n = 6; the data are expressed as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared ipsilaterally with the saline group; one-way ANOVA followed by Tukey's post hoc test.
    Figure Legend Snippet: The effects of the reagents on CCD-induced mechanical allodynia. (a–d) The PWMTs of CCD rats (4 days after operation) 1, 2, 4, and 8 h after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections ( n = 6; the data are expressed as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared ipsilaterally with the saline group; one-way ANOVA followed by Tukey's post hoc test.

    Techniques Used:

    Distribution changes of the p38-positive neurons in DRG tissue. (a–f) p38 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Scale bars: 100 μ m. (g) The analysis of the p38-positive neurons (shown as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.
    Figure Legend Snippet: Distribution changes of the p38-positive neurons in DRG tissue. (a–f) p38 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Scale bars: 100 μ m. (g) The analysis of the p38-positive neurons (shown as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.

    Techniques Used: Immunohistochemical staining, Staining

    The effect of reagents on the protein expression of TRPV4, p38, and P-p38. (a–d) The expression of TRPV4, p38, and P-p38 proteins after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections given to rats 4 days after CCD surgery. The bars represent means ± SEMs based on six experiments; ∗ P < 0.05 and ∗∗ P < 0.01, TRPV4 compared with controls. # P < 0.05 and ## P < 0.01, p38 compared with controls. & P < 0.05 and && P < 0.01, P-p38 compared with controls.
    Figure Legend Snippet: The effect of reagents on the protein expression of TRPV4, p38, and P-p38. (a–d) The expression of TRPV4, p38, and P-p38 proteins after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections given to rats 4 days after CCD surgery. The bars represent means ± SEMs based on six experiments; ∗ P < 0.05 and ∗∗ P < 0.01, TRPV4 compared with controls. # P < 0.05 and ## P < 0.01, p38 compared with controls. & P < 0.05 and && P < 0.01, P-p38 compared with controls.

    Techniques Used: Expressing

    Altered distribution of TRPV4-positive neurons in DRG tissue. (a–f) TRPV4 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups, respectively. Scale bars: 100 μ m. (g) The analysis of the TRPV4-positive neurons (shown as means ± SEMs); ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.
    Figure Legend Snippet: Altered distribution of TRPV4-positive neurons in DRG tissue. (a–f) TRPV4 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups, respectively. Scale bars: 100 μ m. (g) The analysis of the TRPV4-positive neurons (shown as means ± SEMs); ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.

    Techniques Used: Immunohistochemical staining, Staining

    Ectopic discharges after CCD surgery and reagent injection. (a–f) represent discharges of the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Cut from BL-420E+ biological and functional experimental system. (g) and (h) show the amplitudes and frequencies for the different groups (data are expressed as means ± SEM); ∗∗ P < 0.01, compared with the CCD group (7-8 rats in each group).
    Figure Legend Snippet: Ectopic discharges after CCD surgery and reagent injection. (a–f) represent discharges of the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Cut from BL-420E+ biological and functional experimental system. (g) and (h) show the amplitudes and frequencies for the different groups (data are expressed as means ± SEM); ∗∗ P < 0.01, compared with the CCD group (7-8 rats in each group).

    Techniques Used: Injection, Functional Assay

    gpx4  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc gpx4
    Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of <t>GPX4,</t> SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.
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    1) Product Images from "Maternal Organic Selenium Supplementation Relieves Intestinal Endoplasmic Reticulum Stress in Piglets by Enhancing the Expression of Glutathione Peroxidase 4 and Selenoprotein S"

    Article Title: Maternal Organic Selenium Supplementation Relieves Intestinal Endoplasmic Reticulum Stress in Piglets by Enhancing the Expression of Glutathione Peroxidase 4 and Selenoprotein S

    Journal: Frontiers in Nutrition

    doi: 10.3389/fnut.2022.900421

    Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of GPX4, SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.
    Figure Legend Snippet: Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of GPX4, SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.

    Techniques Used: Expressing, Activity Assay

    Roles of GPX4 and SELENOS in regulating ER stress induced by H 2 O 2 in IPEC-J2 cells. (A) IPEC-J2 cells were infected with AdshGPX4, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (B) IPEC-J2 cells were infected with AdshSELS, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (C) IPEC-J2 cells were infected with AdshGPX4 or/and AdshSELS, then were treated with H 2 O 2 . Protein levels of ER stress related markers were detected n = 4 or 3 for western blot assay. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.
    Figure Legend Snippet: Roles of GPX4 and SELENOS in regulating ER stress induced by H 2 O 2 in IPEC-J2 cells. (A) IPEC-J2 cells were infected with AdshGPX4, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (B) IPEC-J2 cells were infected with AdshSELS, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (C) IPEC-J2 cells were infected with AdshGPX4 or/and AdshSELS, then were treated with H 2 O 2 . Protein levels of ER stress related markers were detected n = 4 or 3 for western blot assay. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.

    Techniques Used: Infection, Western Blot

    General view of the effect of HMSeBA on intestinal ER stress. A model for HMSeBA supplementation regulates ER stress induced by LPS or H 2 O 2 in vivo and vitro . Only core components of the pathway are shown. LPS or H 2 O 2 administration induce ER stress through activating the GRP78-IRE1α-CHOP signaling pathway. While HMSeBA treatment improves the expression of GPX4 and SELS, thereby suppresses the ER stress signal, ameliorates the redox status. Maternal HMSeBA is involved in regulating the ER stress signal and health of intestine.
    Figure Legend Snippet: General view of the effect of HMSeBA on intestinal ER stress. A model for HMSeBA supplementation regulates ER stress induced by LPS or H 2 O 2 in vivo and vitro . Only core components of the pathway are shown. LPS or H 2 O 2 administration induce ER stress through activating the GRP78-IRE1α-CHOP signaling pathway. While HMSeBA treatment improves the expression of GPX4 and SELS, thereby suppresses the ER stress signal, ameliorates the redox status. Maternal HMSeBA is involved in regulating the ER stress signal and health of intestine.

    Techniques Used: In Vivo, Expressing

    myogenic colonic slow transmission  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anisomycin
    KEY RESOURCES TABLE
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    1) Product Images from "Labeling of heterochronic ribosomes reveals C1ORF109 and SPATA5 control a late step in human ribosome assembly"

    Article Title: Labeling of heterochronic ribosomes reveals C1ORF109 and SPATA5 control a late step in human ribosome assembly

    Journal: Cell reports

    doi: 10.1016/j.celrep.2022.110597

    KEY RESOURCES TABLE
    Figure Legend Snippet: KEY RESOURCES TABLE

    Techniques Used: Recombinant, Blocking Assay, Magnetic Beads, CRISPR, Knock-In, Software

    hrt cst sm  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anisomycin
    a Immunofluorescence staining of ERα (green) in uncultured PDEC-N and PDEC-BC samples and after 7 days in a LMx-Ag matrix. N = 5 explants examined from 3 biologically independent samples. b The heatmap shows the expression of ERα-regulated gene sets in PDEC-BCs (P182T and P184T) in different matrices in comparison to the uncultured original samples. c PCA of RNAseq data obtained from PDEC-BC and MCF7 cells cultured in LMx-Ag matrix (red) for 7 days compared to the uncultured original tumor / 2D cultured MCF7 cells (grey). Pie charts show the relative distribution of exonic (E), intronic (I) and intergenic (IGR) transcripts in the RNA sequencing. d Heatmaps from GSEA analysis show the enrichment of stress pathway in MMECs, PDEC-N and PDEC-BCs. Different comparisons and the corresponding enrichments are shown in left. e Western blot analysis shows the effect from <t>anisomycin</t> treatment on p38p/p38 and ERα expression in the DU4475 TNBC cell line. TGFβ (2 ng/ml) serves as the negative control, while MCF7 and T47D are positive controls for the ERα ( n = 3). f , Immunofluorescence staining of p38p in DU4475 cells, MMECs, PDEC-Ns, and PDEC-BCs in control and after anisomycin treatment. N = 6 explants examined from three biologically independent samples. Scale bar = 10 μm.
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    1) Product Images from "Compressive stress-mediated p38 activation required for ERα + phenotype in breast cancer"

    Article Title: Compressive stress-mediated p38 activation required for ERα + phenotype in breast cancer

    Journal: Nature Communications

    doi: 10.1038/s41467-021-27220-9

    a Immunofluorescence staining of ERα (green) in uncultured PDEC-N and PDEC-BC samples and after 7 days in a LMx-Ag matrix. N = 5 explants examined from 3 biologically independent samples. b The heatmap shows the expression of ERα-regulated gene sets in PDEC-BCs (P182T and P184T) in different matrices in comparison to the uncultured original samples. c PCA of RNAseq data obtained from PDEC-BC and MCF7 cells cultured in LMx-Ag matrix (red) for 7 days compared to the uncultured original tumor / 2D cultured MCF7 cells (grey). Pie charts show the relative distribution of exonic (E), intronic (I) and intergenic (IGR) transcripts in the RNA sequencing. d Heatmaps from GSEA analysis show the enrichment of stress pathway in MMECs, PDEC-N and PDEC-BCs. Different comparisons and the corresponding enrichments are shown in left. e Western blot analysis shows the effect from anisomycin treatment on p38p/p38 and ERα expression in the DU4475 TNBC cell line. TGFβ (2 ng/ml) serves as the negative control, while MCF7 and T47D are positive controls for the ERα ( n = 3). f , Immunofluorescence staining of p38p in DU4475 cells, MMECs, PDEC-Ns, and PDEC-BCs in control and after anisomycin treatment. N = 6 explants examined from three biologically independent samples. Scale bar = 10 μm.
    Figure Legend Snippet: a Immunofluorescence staining of ERα (green) in uncultured PDEC-N and PDEC-BC samples and after 7 days in a LMx-Ag matrix. N = 5 explants examined from 3 biologically independent samples. b The heatmap shows the expression of ERα-regulated gene sets in PDEC-BCs (P182T and P184T) in different matrices in comparison to the uncultured original samples. c PCA of RNAseq data obtained from PDEC-BC and MCF7 cells cultured in LMx-Ag matrix (red) for 7 days compared to the uncultured original tumor / 2D cultured MCF7 cells (grey). Pie charts show the relative distribution of exonic (E), intronic (I) and intergenic (IGR) transcripts in the RNA sequencing. d Heatmaps from GSEA analysis show the enrichment of stress pathway in MMECs, PDEC-N and PDEC-BCs. Different comparisons and the corresponding enrichments are shown in left. e Western blot analysis shows the effect from anisomycin treatment on p38p/p38 and ERα expression in the DU4475 TNBC cell line. TGFβ (2 ng/ml) serves as the negative control, while MCF7 and T47D are positive controls for the ERα ( n = 3). f , Immunofluorescence staining of p38p in DU4475 cells, MMECs, PDEC-Ns, and PDEC-BCs in control and after anisomycin treatment. N = 6 explants examined from three biologically independent samples. Scale bar = 10 μm.

    Techniques Used: Immunofluorescence, Staining, Expressing, Cell Culture, RNA Sequencing Assay, Western Blot, Negative Control

    a Enrichment of the gene-repressive histone trimethylation (H3K27me3) signatures in different MMECs, PDEC-N, and PDEC-BC samples. Different comparisons and the resulting enrichments are shown below of the graphs. b Immunofluorescent staining of ERα expression in control and enhancer of zeste homolog 2 (EZH2) inhibitor GSK-126 treated explants from DU4475 cells, MMECs, PDEC-N, and PDEC-BC. N = 6 explants examined from three biologically independent samples. c PDEC-BCs from four patients (PxxxT) were exposed to anisomycin or GSK-126 and the effect on ERα-regulated GREB1 and PGR genes was measured using QRT-PCR. d Western blot analysis shows ERα, CK5 and H3K27me3 expression in the DU4475 cells after 48 h treatment with anisomycin ( n = 3). e A model for the stress-mediated regulation of ERα expression. f Illustration of the magnetic cylinder-mediated compression method. g Immunofluorescence images of PDEC-Ns and PDEC-BCs in the LMx-Ag following overnight compression with the magnetic cylinders, stained as indicated. N = 4 explants examined from 3 biologically independent samples. h PDEC-BCs from three patients were exposed to magnet mediated compression for 48 h and with or without tamoxifen treatment. The effect on ERα regulated GREB1 and PGR genes was measured with QRT-PCR. Scale bar = 10 μm.
    Figure Legend Snippet: a Enrichment of the gene-repressive histone trimethylation (H3K27me3) signatures in different MMECs, PDEC-N, and PDEC-BC samples. Different comparisons and the resulting enrichments are shown below of the graphs. b Immunofluorescent staining of ERα expression in control and enhancer of zeste homolog 2 (EZH2) inhibitor GSK-126 treated explants from DU4475 cells, MMECs, PDEC-N, and PDEC-BC. N = 6 explants examined from three biologically independent samples. c PDEC-BCs from four patients (PxxxT) were exposed to anisomycin or GSK-126 and the effect on ERα-regulated GREB1 and PGR genes was measured using QRT-PCR. d Western blot analysis shows ERα, CK5 and H3K27me3 expression in the DU4475 cells after 48 h treatment with anisomycin ( n = 3). e A model for the stress-mediated regulation of ERα expression. f Illustration of the magnetic cylinder-mediated compression method. g Immunofluorescence images of PDEC-Ns and PDEC-BCs in the LMx-Ag following overnight compression with the magnetic cylinders, stained as indicated. N = 4 explants examined from 3 biologically independent samples. h PDEC-BCs from three patients were exposed to magnet mediated compression for 48 h and with or without tamoxifen treatment. The effect on ERα regulated GREB1 and PGR genes was measured with QRT-PCR. Scale bar = 10 μm.

    Techniques Used: Staining, Expressing, Quantitative RT-PCR, Western Blot, Immunofluorescence

    dyes  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc cst r ec harbored mcr genes
    Analysis of the rep-PCR results from 67 Escherichia coli ( Ec ) strains isolated from local people ( n = 20 out of 51 detected), poultry ( n = 18 out of 28 detected), chicken meat ( n = 17 out of 17 detected), Swiss residents after traveling to Laos ( n = 10 out of 10 detected), and Swiss residents before traveling ( n = 2 out of 3 detected). Strains showing an identity of ≥ 85% were considered to belong to the same clone. For each isolate, we show the phenotype, the main antimicrobial resistance genes (ARGs) detected by using microarray or PCRs, and the sequence type (ST). Isolates underlined in blue are from local people in Laos, in green from poultry, in yellow from chicken meat, in orange from Swiss residents before traveling, and in red from returning travelers. ESC-R, extended-spectrum cephalosporin-resistant; <t>CST-R,</t> colistin-resistant; Carba-R, carbapenem-resistant; NP, not performed (strains not typed with WGS); -, <t>mcr</t> and/or bla ESBL genes not detected; gr, group; subgr, subgroup.
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    1) Product Images from "Antimicrobial-Resistant Escherichia coli Strains and Their Plasmids in People, Poultry, and Chicken Meat in Laos"

    Article Title: Antimicrobial-Resistant Escherichia coli Strains and Their Plasmids in People, Poultry, and Chicken Meat in Laos

    Journal: Frontiers in Microbiology

    doi: 10.3389/fmicb.2021.708182

    Analysis of the rep-PCR results from 67 Escherichia coli ( Ec ) strains isolated from local people ( n = 20 out of 51 detected), poultry ( n = 18 out of 28 detected), chicken meat ( n = 17 out of 17 detected), Swiss residents after traveling to Laos ( n = 10 out of 10 detected), and Swiss residents before traveling ( n = 2 out of 3 detected). Strains showing an identity of ≥ 85% were considered to belong to the same clone. For each isolate, we show the phenotype, the main antimicrobial resistance genes (ARGs) detected by using microarray or PCRs, and the sequence type (ST). Isolates underlined in blue are from local people in Laos, in green from poultry, in yellow from chicken meat, in orange from Swiss residents before traveling, and in red from returning travelers. ESC-R, extended-spectrum cephalosporin-resistant; CST-R, colistin-resistant; Carba-R, carbapenem-resistant; NP, not performed (strains not typed with WGS); -, mcr and/or bla ESBL genes not detected; gr, group; subgr, subgroup.
    Figure Legend Snippet: Analysis of the rep-PCR results from 67 Escherichia coli ( Ec ) strains isolated from local people ( n = 20 out of 51 detected), poultry ( n = 18 out of 28 detected), chicken meat ( n = 17 out of 17 detected), Swiss residents after traveling to Laos ( n = 10 out of 10 detected), and Swiss residents before traveling ( n = 2 out of 3 detected). Strains showing an identity of ≥ 85% were considered to belong to the same clone. For each isolate, we show the phenotype, the main antimicrobial resistance genes (ARGs) detected by using microarray or PCRs, and the sequence type (ST). Isolates underlined in blue are from local people in Laos, in green from poultry, in yellow from chicken meat, in orange from Swiss residents before traveling, and in red from returning travelers. ESC-R, extended-spectrum cephalosporin-resistant; CST-R, colistin-resistant; Carba-R, carbapenem-resistant; NP, not performed (strains not typed with WGS); -, mcr and/or bla ESBL genes not detected; gr, group; subgr, subgroup.

    Techniques Used: Isolation, Microarray, Sequencing

    anisomycin treatment  (Cell Signaling Technology Inc)


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    Cell Signaling Technology Inc anisomycin
    Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. <t>Anisomycin</t> was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.
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    The effects of the reagents on CCD-induced mechanical allodynia. (a–d) The PWMTs of CCD rats (4 days after operation) 1, 2, 4, and 8 h after RR, 4 α -PDD, SB203580, and <t>anisomycin</t> intrathecal injections ( n = 6; the data are expressed as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared ipsilaterally with the saline group; one-way ANOVA followed by Tukey's post hoc test.
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    Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of <t>GPX4,</t> SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.
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    Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of <t>GPX4,</t> SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.
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    Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of <t>GPX4,</t> SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.
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    Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of <t>GPX4,</t> SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.
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    Cell Signaling Technology Inc cst r ec harbored mcr genes
    Analysis of the rep-PCR results from 67 Escherichia coli ( Ec ) strains isolated from local people ( n = 20 out of 51 detected), poultry ( n = 18 out of 28 detected), chicken meat ( n = 17 out of 17 detected), Swiss residents after traveling to Laos ( n = 10 out of 10 detected), and Swiss residents before traveling ( n = 2 out of 3 detected). Strains showing an identity of ≥ 85% were considered to belong to the same clone. For each isolate, we show the phenotype, the main antimicrobial resistance genes (ARGs) detected by using microarray or PCRs, and the sequence type (ST). Isolates underlined in blue are from local people in Laos, in green from poultry, in yellow from chicken meat, in orange from Swiss residents before traveling, and in red from returning travelers. ESC-R, extended-spectrum cephalosporin-resistant; <t>CST-R,</t> colistin-resistant; Carba-R, carbapenem-resistant; NP, not performed (strains not typed with WGS); -, <t>mcr</t> and/or bla ESBL genes not detected; gr, group; subgr, subgroup.
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    Cell Signaling Technology Inc anisomycin treatment
    Analysis of the rep-PCR results from 67 Escherichia coli ( Ec ) strains isolated from local people ( n = 20 out of 51 detected), poultry ( n = 18 out of 28 detected), chicken meat ( n = 17 out of 17 detected), Swiss residents after traveling to Laos ( n = 10 out of 10 detected), and Swiss residents before traveling ( n = 2 out of 3 detected). Strains showing an identity of ≥ 85% were considered to belong to the same clone. For each isolate, we show the phenotype, the main antimicrobial resistance genes (ARGs) detected by using microarray or PCRs, and the sequence type (ST). Isolates underlined in blue are from local people in Laos, in green from poultry, in yellow from chicken meat, in orange from Swiss residents before traveling, and in red from returning travelers. ESC-R, extended-spectrum cephalosporin-resistant; <t>CST-R,</t> colistin-resistant; Carba-R, carbapenem-resistant; NP, not performed (strains not typed with WGS); -, <t>mcr</t> and/or bla ESBL genes not detected; gr, group; subgr, subgroup.
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    Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. Anisomycin was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.

    Journal: Theranostics

    Article Title: Dioscin Exerts Protective Effects Against Crystalline Silica-induced Pulmonary Fibrosis in Mice

    doi: 10.7150/thno.20270

    Figure Lengend Snippet: Dioscin inhibits macrophages and fibroblasts from secreting pro-inflammatory cytokines. (A) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in LPS-primed RAW 264.7 macrophages treated with CS (50 μg/cm 2 ) together with different does of dioscin for 6 h. Anisomycin was used to reverse dioscin's effect (2 ng/mL). Quantification of each protein level relative to β-Actin is shown below each band . (B-D) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (E-H) ELISA analyses of pro-inflammation cytokines in the culture medium of different treated macrophages at 6 h. (E) MCP-1, (F) IL-1β, (G) IL-6, (H)TNF-α (n=3). Data are the mean of three independent experiments (n=3). (I) Western blot analyses of ASK1 phosphorylation and downstream MAPK proteins (p38 and JNK/SAPK) in TNF-α- (25 ng/mL) treated NIH-3T3 fibroblasts together with different does of dioscin for 30 min. Quantification of each protein level relative to β-Actin is shown below each band . (J-L) The levels of phospho-ASK1, phospho-p38 and phosphor-JNK were normalized to those of β-Actin (n=3). (M) qPCR analysis of Il-6 mRNA levels in TNF-α- (25 ng/mL) treated fibroblasts together with different concentrations of dioscin for 12 h (n=3). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed twice with similar results.

    Article Snippet: Anisomycin (2 ng/mL, Cell Signaling Technology), activator of JNK and p38 MAPKs, was used to test the biological effect of dioscin on the MAPKs.

    Techniques: Western Blot, Enzyme-linked Immunosorbent Assay

    Dioscin regulated the TGF-β1-Samd3 signaling pathway and regulatory T cell functions. (A, B) Western blot analysis and quantification of TGF-β1 in lungs of dioscin-treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of TGF-β1 levels relative to β-Actin is shown below the band. (C) qPCR analysis of Tgf-β1 in the lung tissues (n=6-8 per group). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed in triplicate. (D, E) Western blot analysis and quantification of Smad3 signaling in the lungs of treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (F, G) Western blot analysis of p-Smad3 and Smad3 in NIH-3T3 fibroblast, 30 min after TGF-β1 (5 ng/mL) and dioscin treatment. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (H) Schematic design of transwell experiment: RAW 264.7 macrophages were primed by LPS (25 ng/mL) for 3 h, and then treated with CS (50 μg/cm 2 ) together with dioscin or anisomycin (2 ng/mL) for 24 h in the lower plate, while NIH-3T3 fibroblasts were attached in the upper chamber. After that, the upper chamber was inserted back to the lower plate, and macrophages and fibroblasts were co-cultured for another 24 h. Then, the mRNA expressions of Col1a1 (I) and α-SMA (J) in NIH-3T3 fibroblast were determined by qPCR (n=3). (K, L) Percentage of CD4+CD25+FoxP3+ cells (regulatory T cells) in hilar lymph nodes was assayed by FACS analysis (n=5). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD.

    Journal: Theranostics

    Article Title: Dioscin Exerts Protective Effects Against Crystalline Silica-induced Pulmonary Fibrosis in Mice

    doi: 10.7150/thno.20270

    Figure Lengend Snippet: Dioscin regulated the TGF-β1-Samd3 signaling pathway and regulatory T cell functions. (A, B) Western blot analysis and quantification of TGF-β1 in lungs of dioscin-treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of TGF-β1 levels relative to β-Actin is shown below the band. (C) qPCR analysis of Tgf-β1 in the lung tissues (n=6-8 per group). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD. The experiment was performed in triplicate. (D, E) Western blot analysis and quantification of Smad3 signaling in the lungs of treated mice at 56 d (n=3). β-Actin was used as the loading control. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (F, G) Western blot analysis of p-Smad3 and Smad3 in NIH-3T3 fibroblast, 30 min after TGF-β1 (5 ng/mL) and dioscin treatment. Quantification of p-Smad3 levels relative to β-Actin is shown below the band. (H) Schematic design of transwell experiment: RAW 264.7 macrophages were primed by LPS (25 ng/mL) for 3 h, and then treated with CS (50 μg/cm 2 ) together with dioscin or anisomycin (2 ng/mL) for 24 h in the lower plate, while NIH-3T3 fibroblasts were attached in the upper chamber. After that, the upper chamber was inserted back to the lower plate, and macrophages and fibroblasts were co-cultured for another 24 h. Then, the mRNA expressions of Col1a1 (I) and α-SMA (J) in NIH-3T3 fibroblast were determined by qPCR (n=3). (K, L) Percentage of CD4+CD25+FoxP3+ cells (regulatory T cells) in hilar lymph nodes was assayed by FACS analysis (n=5). *, P<0.05; **, P<0.01. Error bars indicate the mean ± SD.

    Article Snippet: Anisomycin (2 ng/mL, Cell Signaling Technology), activator of JNK and p38 MAPKs, was used to test the biological effect of dioscin on the MAPKs.

    Techniques: Western Blot, Cell Culture

    The effects of the reagents on CCD-induced mechanical allodynia. (a–d) The PWMTs of CCD rats (4 days after operation) 1, 2, 4, and 8 h after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections ( n = 6; the data are expressed as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared ipsilaterally with the saline group; one-way ANOVA followed by Tukey's post hoc test.

    Journal: BioMed Research International

    Article Title: Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion

    doi: 10.1155/2016/6978923

    Figure Lengend Snippet: The effects of the reagents on CCD-induced mechanical allodynia. (a–d) The PWMTs of CCD rats (4 days after operation) 1, 2, 4, and 8 h after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections ( n = 6; the data are expressed as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared ipsilaterally with the saline group; one-way ANOVA followed by Tukey's post hoc test.

    Article Snippet: Four days after CCD surgery, the TRPV4 inhibitor Ruthenium Red (RR, Sigma, Germany), the TRPV4 agonist 4 α -PDD (CST, USA), the p38 inhibitor SB203580 (CST, USA), or the p38 agonist anisomycin (CST, USA) was given to the experimental groups at the recommended concentrations via intrathecal injection.

    Techniques:

    Distribution changes of the p38-positive neurons in DRG tissue. (a–f) p38 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Scale bars: 100 μ m. (g) The analysis of the p38-positive neurons (shown as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.

    Journal: BioMed Research International

    Article Title: Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion

    doi: 10.1155/2016/6978923

    Figure Lengend Snippet: Distribution changes of the p38-positive neurons in DRG tissue. (a–f) p38 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Scale bars: 100 μ m. (g) The analysis of the p38-positive neurons (shown as means ± SEMs); ∗ P < 0.05 and ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.

    Article Snippet: Four days after CCD surgery, the TRPV4 inhibitor Ruthenium Red (RR, Sigma, Germany), the TRPV4 agonist 4 α -PDD (CST, USA), the p38 inhibitor SB203580 (CST, USA), or the p38 agonist anisomycin (CST, USA) was given to the experimental groups at the recommended concentrations via intrathecal injection.

    Techniques: Immunohistochemical staining, Staining

    The effect of reagents on the protein expression of TRPV4, p38, and P-p38. (a–d) The expression of TRPV4, p38, and P-p38 proteins after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections given to rats 4 days after CCD surgery. The bars represent means ± SEMs based on six experiments; ∗ P < 0.05 and ∗∗ P < 0.01, TRPV4 compared with controls. # P < 0.05 and ## P < 0.01, p38 compared with controls. & P < 0.05 and && P < 0.01, P-p38 compared with controls.

    Journal: BioMed Research International

    Article Title: Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion

    doi: 10.1155/2016/6978923

    Figure Lengend Snippet: The effect of reagents on the protein expression of TRPV4, p38, and P-p38. (a–d) The expression of TRPV4, p38, and P-p38 proteins after RR, 4 α -PDD, SB203580, and anisomycin intrathecal injections given to rats 4 days after CCD surgery. The bars represent means ± SEMs based on six experiments; ∗ P < 0.05 and ∗∗ P < 0.01, TRPV4 compared with controls. # P < 0.05 and ## P < 0.01, p38 compared with controls. & P < 0.05 and && P < 0.01, P-p38 compared with controls.

    Article Snippet: Four days after CCD surgery, the TRPV4 inhibitor Ruthenium Red (RR, Sigma, Germany), the TRPV4 agonist 4 α -PDD (CST, USA), the p38 inhibitor SB203580 (CST, USA), or the p38 agonist anisomycin (CST, USA) was given to the experimental groups at the recommended concentrations via intrathecal injection.

    Techniques: Expressing

    Altered distribution of TRPV4-positive neurons in DRG tissue. (a–f) TRPV4 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups, respectively. Scale bars: 100 μ m. (g) The analysis of the TRPV4-positive neurons (shown as means ± SEMs); ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.

    Journal: BioMed Research International

    Article Title: Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion

    doi: 10.1155/2016/6978923

    Figure Lengend Snippet: Altered distribution of TRPV4-positive neurons in DRG tissue. (a–f) TRPV4 immunohistochemical staining of the DRG neurons in the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups, respectively. Scale bars: 100 μ m. (g) The analysis of the TRPV4-positive neurons (shown as means ± SEMs); ∗∗ P < 0.01 compared with controls; ## P < 0.01 compared with the CCD group.

    Article Snippet: Four days after CCD surgery, the TRPV4 inhibitor Ruthenium Red (RR, Sigma, Germany), the TRPV4 agonist 4 α -PDD (CST, USA), the p38 inhibitor SB203580 (CST, USA), or the p38 agonist anisomycin (CST, USA) was given to the experimental groups at the recommended concentrations via intrathecal injection.

    Techniques: Immunohistochemical staining, Staining

    Ectopic discharges after CCD surgery and reagent injection. (a–f) represent discharges of the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Cut from BL-420E+ biological and functional experimental system. (g) and (h) show the amplitudes and frequencies for the different groups (data are expressed as means ± SEM); ∗∗ P < 0.01, compared with the CCD group (7-8 rats in each group).

    Journal: BioMed Research International

    Article Title: Effect of TRPV4-p38 MAPK Pathway on Neuropathic Pain in Rats with Chronic Compression of the Dorsal Root Ganglion

    doi: 10.1155/2016/6978923

    Figure Lengend Snippet: Ectopic discharges after CCD surgery and reagent injection. (a–f) represent discharges of the control, CCD, CCD+RR 10 nmol/L, CCD+4 α -PDD 10 nmol/L, CCD+SB203580 20 μ mol/L, and CCD+anisomycin 25 μ g/mL groups. Cut from BL-420E+ biological and functional experimental system. (g) and (h) show the amplitudes and frequencies for the different groups (data are expressed as means ± SEM); ∗∗ P < 0.01, compared with the CCD group (7-8 rats in each group).

    Article Snippet: Four days after CCD surgery, the TRPV4 inhibitor Ruthenium Red (RR, Sigma, Germany), the TRPV4 agonist 4 α -PDD (CST, USA), the p38 inhibitor SB203580 (CST, USA), or the p38 agonist anisomycin (CST, USA) was given to the experimental groups at the recommended concentrations via intrathecal injection.

    Techniques: Injection, Functional Assay

    Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of GPX4, SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.

    Journal: Frontiers in Nutrition

    Article Title: Maternal Organic Selenium Supplementation Relieves Intestinal Endoplasmic Reticulum Stress in Piglets by Enhancing the Expression of Glutathione Peroxidase 4 and Selenoprotein S

    doi: 10.3389/fnut.2022.900421

    Figure Lengend Snippet: Effect of HMSeBA supplementation on the expression of selenoproteins and the antioxidative capacity in IPEC-J2 cells. IPEC-J2 cells were treated with 1.2 μM HMSeBA for 24 h. (A) Relative mRNA levels of selenoproteins. n = 3 for each group. (B) Relative protein abundance of GPX4, SELENOS and SELENOP. n = 4 for each group. (C) The activity of antioxidant related enzymes. n = 6 for each group. GSH-Px, glutathione peroxidase; T-SOD, total superoxide dismutase; CAT, catalase; T-AOC, total antioxidant capacity. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.

    Article Snippet: The PVDF membrane was then blocked with 1% BSA containing TBST, followed by incubation with primary antibodies at 4°C for more than 8 h. The primary antibodies were as follows: SELS (21 KDa, 15591-1-AP, Proteintech), GPX4 (22 KDa, 52455S, Cell Signaling Technology), SEPP1 (57/45 KDa, sc-376858, Santa Cruz Biotechnology), GRP78 (78 KDa, ab21685, Abcam), p-IRE1α (110 KDa, ab48187, Abcam), IRE1α (110 KDa, ab37073, Abcam), p-PERK (170 KDa, 3179S, Cell Signaling Technology), PERK (170 KDa, 3192S, Cell Signaling Technology), CHOP (27 KDa, 381679, Zen BioScience) and β-actin (45 kDa, #4967, Cell Signaling Technology).

    Techniques: Expressing, Activity Assay

    Roles of GPX4 and SELENOS in regulating ER stress induced by H 2 O 2 in IPEC-J2 cells. (A) IPEC-J2 cells were infected with AdshGPX4, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (B) IPEC-J2 cells were infected with AdshSELS, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (C) IPEC-J2 cells were infected with AdshGPX4 or/and AdshSELS, then were treated with H 2 O 2 . Protein levels of ER stress related markers were detected n = 4 or 3 for western blot assay. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.

    Journal: Frontiers in Nutrition

    Article Title: Maternal Organic Selenium Supplementation Relieves Intestinal Endoplasmic Reticulum Stress in Piglets by Enhancing the Expression of Glutathione Peroxidase 4 and Selenoprotein S

    doi: 10.3389/fnut.2022.900421

    Figure Lengend Snippet: Roles of GPX4 and SELENOS in regulating ER stress induced by H 2 O 2 in IPEC-J2 cells. (A) IPEC-J2 cells were infected with AdshGPX4, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (B) IPEC-J2 cells were infected with AdshSELS, then were treated with H 2 O 2 plus HMSeBA. Protein levels of ER stress related markers were detected. (C) IPEC-J2 cells were infected with AdshGPX4 or/and AdshSELS, then were treated with H 2 O 2 . Protein levels of ER stress related markers were detected n = 4 or 3 for western blot assay. Data are expressed as mean ± SE. * P < 0.05, ** P < 0.01, *** P < 0.001.

    Article Snippet: The PVDF membrane was then blocked with 1% BSA containing TBST, followed by incubation with primary antibodies at 4°C for more than 8 h. The primary antibodies were as follows: SELS (21 KDa, 15591-1-AP, Proteintech), GPX4 (22 KDa, 52455S, Cell Signaling Technology), SEPP1 (57/45 KDa, sc-376858, Santa Cruz Biotechnology), GRP78 (78 KDa, ab21685, Abcam), p-IRE1α (110 KDa, ab48187, Abcam), IRE1α (110 KDa, ab37073, Abcam), p-PERK (170 KDa, 3179S, Cell Signaling Technology), PERK (170 KDa, 3192S, Cell Signaling Technology), CHOP (27 KDa, 381679, Zen BioScience) and β-actin (45 kDa, #4967, Cell Signaling Technology).

    Techniques: Infection, Western Blot

    General view of the effect of HMSeBA on intestinal ER stress. A model for HMSeBA supplementation regulates ER stress induced by LPS or H 2 O 2 in vivo and vitro . Only core components of the pathway are shown. LPS or H 2 O 2 administration induce ER stress through activating the GRP78-IRE1α-CHOP signaling pathway. While HMSeBA treatment improves the expression of GPX4 and SELS, thereby suppresses the ER stress signal, ameliorates the redox status. Maternal HMSeBA is involved in regulating the ER stress signal and health of intestine.

    Journal: Frontiers in Nutrition

    Article Title: Maternal Organic Selenium Supplementation Relieves Intestinal Endoplasmic Reticulum Stress in Piglets by Enhancing the Expression of Glutathione Peroxidase 4 and Selenoprotein S

    doi: 10.3389/fnut.2022.900421

    Figure Lengend Snippet: General view of the effect of HMSeBA on intestinal ER stress. A model for HMSeBA supplementation regulates ER stress induced by LPS or H 2 O 2 in vivo and vitro . Only core components of the pathway are shown. LPS or H 2 O 2 administration induce ER stress through activating the GRP78-IRE1α-CHOP signaling pathway. While HMSeBA treatment improves the expression of GPX4 and SELS, thereby suppresses the ER stress signal, ameliorates the redox status. Maternal HMSeBA is involved in regulating the ER stress signal and health of intestine.

    Article Snippet: The PVDF membrane was then blocked with 1% BSA containing TBST, followed by incubation with primary antibodies at 4°C for more than 8 h. The primary antibodies were as follows: SELS (21 KDa, 15591-1-AP, Proteintech), GPX4 (22 KDa, 52455S, Cell Signaling Technology), SEPP1 (57/45 KDa, sc-376858, Santa Cruz Biotechnology), GRP78 (78 KDa, ab21685, Abcam), p-IRE1α (110 KDa, ab48187, Abcam), IRE1α (110 KDa, ab37073, Abcam), p-PERK (170 KDa, 3179S, Cell Signaling Technology), PERK (170 KDa, 3192S, Cell Signaling Technology), CHOP (27 KDa, 381679, Zen BioScience) and β-actin (45 kDa, #4967, Cell Signaling Technology).

    Techniques: In Vivo, Expressing

    Analysis of the rep-PCR results from 67 Escherichia coli ( Ec ) strains isolated from local people ( n = 20 out of 51 detected), poultry ( n = 18 out of 28 detected), chicken meat ( n = 17 out of 17 detected), Swiss residents after traveling to Laos ( n = 10 out of 10 detected), and Swiss residents before traveling ( n = 2 out of 3 detected). Strains showing an identity of ≥ 85% were considered to belong to the same clone. For each isolate, we show the phenotype, the main antimicrobial resistance genes (ARGs) detected by using microarray or PCRs, and the sequence type (ST). Isolates underlined in blue are from local people in Laos, in green from poultry, in yellow from chicken meat, in orange from Swiss residents before traveling, and in red from returning travelers. ESC-R, extended-spectrum cephalosporin-resistant; CST-R, colistin-resistant; Carba-R, carbapenem-resistant; NP, not performed (strains not typed with WGS); -, mcr and/or bla ESBL genes not detected; gr, group; subgr, subgroup.

    Journal: Frontiers in Microbiology

    Article Title: Antimicrobial-Resistant Escherichia coli Strains and Their Plasmids in People, Poultry, and Chicken Meat in Laos

    doi: 10.3389/fmicb.2021.708182

    Figure Lengend Snippet: Analysis of the rep-PCR results from 67 Escherichia coli ( Ec ) strains isolated from local people ( n = 20 out of 51 detected), poultry ( n = 18 out of 28 detected), chicken meat ( n = 17 out of 17 detected), Swiss residents after traveling to Laos ( n = 10 out of 10 detected), and Swiss residents before traveling ( n = 2 out of 3 detected). Strains showing an identity of ≥ 85% were considered to belong to the same clone. For each isolate, we show the phenotype, the main antimicrobial resistance genes (ARGs) detected by using microarray or PCRs, and the sequence type (ST). Isolates underlined in blue are from local people in Laos, in green from poultry, in yellow from chicken meat, in orange from Swiss residents before traveling, and in red from returning travelers. ESC-R, extended-spectrum cephalosporin-resistant; CST-R, colistin-resistant; Carba-R, carbapenem-resistant; NP, not performed (strains not typed with WGS); -, mcr and/or bla ESBL genes not detected; gr, group; subgr, subgroup.

    Article Snippet: Furthermore, 24 (82.8%) out of the 29 CST-R- Ec harbored mcr genes ( mcr-1 -like, n = 22; mcr-3 -like, n = 2).

    Techniques: Isolation, Microarray, Sequencing