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  • 99
    Millipore sp600125
    Kinetic inhibition of MAPKs. ELISA quantitation of IL-8 concentrations in cells pretreated with ERK1/2 PD980559, JNK <t>SP600125,</t> and SB203580 inhibitors. Control is HEp-2 cells infected with live HPIV-1. (a) ERK1/2 inhibitor assay is displayed in red cells treated with the inhibitor. (b) JNK inhibitor assay, green, occurs in cells treated with the inhibitor. (c) p38 inhibitor assay, purple, occurs in cells treated with inhibitor. Data are the means ± SD for triplicate measurements from the three separate experiments.
    Sp600125, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 5486 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Tocris sp600125
    MAP kinases activation is not involved in 4-HNE-elicited lipolytic response. A, Fully-differentiated 3T3-L1 cells were treated with exogenous 4-HNE (20 microM) for indicated time points. The activations of the MAP kinases, including ERK1/2, p38, and JNK, were detected by immunoblotting. B, Fully-differentiated 3T3-L1 cells were pretreated with specific kinase inhibitors for ERK1/2 (U0126, 10 microM), JNK <t>(SP600125,</t> 10 microM), and p38 (SB203580, 10 microM) for 1 hours, followed by exogenous 4-HNE (20 microM) exposure. Glycerol releases were measured 6 hours later. All values are denoted as Means ± SD from three or more independent batches of cells. Bars with different characters differ significantly (p
    Sp600125, supplied by Tocris, used in various techniques. Bioz Stars score: 99/100, based on 579 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/sp600125/product/Tocris
    Average 99 stars, based on 579 article reviews
    Price from $9.99 to $1999.99
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    99
    Selleck Chemicals sp 600125 jnki
    MAP kinases activation is not involved in 4-HNE-elicited lipolytic response. A, Fully-differentiated 3T3-L1 cells were treated with exogenous 4-HNE (20 microM) for indicated time points. The activations of the MAP kinases, including ERK1/2, p38, and JNK, were detected by immunoblotting. B, Fully-differentiated 3T3-L1 cells were pretreated with specific kinase inhibitors for ERK1/2 (U0126, 10 microM), JNK <t>(SP600125,</t> 10 microM), and p38 (SB203580, 10 microM) for 1 hours, followed by exogenous 4-HNE (20 microM) exposure. Glycerol releases were measured 6 hours later. All values are denoted as Means ± SD from three or more independent batches of cells. Bars with different characters differ significantly (p
    Sp 600125 Jnki, supplied by Selleck Chemicals, used in various techniques. Bioz Stars score: 99/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Avantor 1 9 pyrazoloanthrone
    MAP kinases activation is not involved in 4-HNE-elicited lipolytic response. A, Fully-differentiated 3T3-L1 cells were treated with exogenous 4-HNE (20 microM) for indicated time points. The activations of the MAP kinases, including ERK1/2, p38, and JNK, were detected by immunoblotting. B, Fully-differentiated 3T3-L1 cells were pretreated with specific kinase inhibitors for ERK1/2 (U0126, 10 microM), JNK <t>(SP600125,</t> 10 microM), and p38 (SB203580, 10 microM) for 1 hours, followed by exogenous 4-HNE (20 microM) exposure. Glycerol releases were measured 6 hours later. All values are denoted as Means ± SD from three or more independent batches of cells. Bars with different characters differ significantly (p
    1 9 Pyrazoloanthrone, supplied by Avantor, used in various techniques. Bioz Stars score: 88/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Price from $9.99 to $1999.99
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    Image Search Results


    Kinetic inhibition of MAPKs. ELISA quantitation of IL-8 concentrations in cells pretreated with ERK1/2 PD980559, JNK SP600125, and SB203580 inhibitors. Control is HEp-2 cells infected with live HPIV-1. (a) ERK1/2 inhibitor assay is displayed in red cells treated with the inhibitor. (b) JNK inhibitor assay, green, occurs in cells treated with the inhibitor. (c) p38 inhibitor assay, purple, occurs in cells treated with inhibitor. Data are the means ± SD for triplicate measurements from the three separate experiments.

    Journal: Journal of Immunology Research

    Article Title: Parainfluenza Virus Type 1 Induces Epithelial IL-8 Production via p38-MAPK Signalling

    doi: 10.1155/2014/515984

    Figure Lengend Snippet: Kinetic inhibition of MAPKs. ELISA quantitation of IL-8 concentrations in cells pretreated with ERK1/2 PD980559, JNK SP600125, and SB203580 inhibitors. Control is HEp-2 cells infected with live HPIV-1. (a) ERK1/2 inhibitor assay is displayed in red cells treated with the inhibitor. (b) JNK inhibitor assay, green, occurs in cells treated with the inhibitor. (c) p38 inhibitor assay, purple, occurs in cells treated with inhibitor. Data are the means ± SD for triplicate measurements from the three separate experiments.

    Article Snippet: IL-8 Quantitation in the Presence of Kinase Inhibitors For the best reproduction of larynx conditions, assays were conducted using HEp-2 cells that were pretreated with the kinase inhibitors PD980559 for ERK1/2, SB203580* for p38 MAPK, and SP600125* for JNK from Calbiochem Millipore* (San Diego, CA).

    Techniques: Inhibition, Enzyme-linked Immunosorbent Assay, Quantitation Assay, Infection

    Inhibitors of ERK1/2 PD980559, JNK SP600125, and p38 SB203580 inhibited the expression of each of the MAPKs in the HEp-2 cell line treated cell 1 hour before infection and maintained for 120 minutes after the test. The experiments were performed by adding inhibitor at concentrations from 5 μ M to 20 μ M (1.3 to 5.2 μ L, resp.). Protein levels were detected by Western blot. Immunoblotting was done with monoclonal antibody specific for protein phosphorylated technique that was described in “Materials and Methods”. The extracted proteins (−) represent HEp-2 cells without infecting and the proteins (+) refer to cells infected with the HPIV-1. The tests were performed in triplicate.

    Journal: Journal of Immunology Research

    Article Title: Parainfluenza Virus Type 1 Induces Epithelial IL-8 Production via p38-MAPK Signalling

    doi: 10.1155/2014/515984

    Figure Lengend Snippet: Inhibitors of ERK1/2 PD980559, JNK SP600125, and p38 SB203580 inhibited the expression of each of the MAPKs in the HEp-2 cell line treated cell 1 hour before infection and maintained for 120 minutes after the test. The experiments were performed by adding inhibitor at concentrations from 5 μ M to 20 μ M (1.3 to 5.2 μ L, resp.). Protein levels were detected by Western blot. Immunoblotting was done with monoclonal antibody specific for protein phosphorylated technique that was described in “Materials and Methods”. The extracted proteins (−) represent HEp-2 cells without infecting and the proteins (+) refer to cells infected with the HPIV-1. The tests were performed in triplicate.

    Article Snippet: IL-8 Quantitation in the Presence of Kinase Inhibitors For the best reproduction of larynx conditions, assays were conducted using HEp-2 cells that were pretreated with the kinase inhibitors PD980559 for ERK1/2, SB203580* for p38 MAPK, and SP600125* for JNK from Calbiochem Millipore* (San Diego, CA).

    Techniques: Expressing, Infection, Western Blot

    Effect of inhibition of JNK on HLMVEC network formation in Matrigel. A : representative bright field images of HLMVECs plated on Matrigel as described in MATERIALS AND METHODS in the presence of vehicle (DMSO), SP-600125 (5 μM), or the negative

    Journal:

    Article Title: Role of JNK in network formation of human lung microvascular endothelial cells

    doi: 10.1152/ajplung.00496.2007

    Figure Lengend Snippet: Effect of inhibition of JNK on HLMVEC network formation in Matrigel. A : representative bright field images of HLMVECs plated on Matrigel as described in MATERIALS AND METHODS in the presence of vehicle (DMSO), SP-600125 (5 μM), or the negative

    Article Snippet: The ERK1/2 inhibitors PD 98059, U0126, SB-203580, and SP-600125 and negative JNK control (N1 -Methyl-1,9-pyrazoloanthrone, cat. no. 420123) were purchased from Calbiochem, La Jolla, CA.

    Techniques: Inhibition

    WISP1 inhibits DOX-induced MAPK activation and cardiomyocyte death A , DOX induces p38 MAPK activation. Cardiomyocytes were treated with DOX (1 μM) for the indicated time periods. p38 MAPK activation was analyzed by immunoblotting using activation-specific antibodies (p-p38 MAPK Thr180/Tyr182) and cleared whole cell homogenates (n=3). B , DOX-mediated p38 MAPK activation is inhibited by SB 203580. Cardiomyocytes were treated with SB 203580 (SB; 1 μM in DMSO for 30 min) prior to DOX (1 μM for 30 min) addition, and then analyzed as in A (n=3). C , DOX induces JNK activation. Cardiomyocytes treated as in A were analyzed for JNK activation by immunoblotting using activation-specific antibodies and cleared whole cell homogenates (n=3). D , SP 600125 inhibits DOX-mediated JNK activation (pJNK Thr183/Tyr185). Cardiomyocytes were treated with SP 600125 (SP; 20 μM in DMSO for 30 min) prior to DOX (1 μM for 30 min) treatment, and then analyzed as in C (n=3). E , SB and SP attenuate DOX-induced Bax translocation to mitochondria. Cardiomyocytes were treated with SB and SP prior to DOX addition (1 μM for 8 h). Bax levels in mitochondria were analyzed by immunoblotting. V-β served as a loading and purity control. F , SB and SP attenuate DOX-induced cardiomyocyte death. Cardiomyocytes were treated with SB and SP prior to DOX addition (1 μM for 24 h). Cell death was analyzed as in 1 A . * P

    Journal: Cellular signalling

    Article Title: WNT1-Inducible Signaling Pathway Protein-1 Activates Diverse Cell Survival Pathways and Blocks Doxorubicin-induced Cardiomyocyte Death

    doi: 10.1016/j.cellsig.2010.01.005

    Figure Lengend Snippet: WISP1 inhibits DOX-induced MAPK activation and cardiomyocyte death A , DOX induces p38 MAPK activation. Cardiomyocytes were treated with DOX (1 μM) for the indicated time periods. p38 MAPK activation was analyzed by immunoblotting using activation-specific antibodies (p-p38 MAPK Thr180/Tyr182) and cleared whole cell homogenates (n=3). B , DOX-mediated p38 MAPK activation is inhibited by SB 203580. Cardiomyocytes were treated with SB 203580 (SB; 1 μM in DMSO for 30 min) prior to DOX (1 μM for 30 min) addition, and then analyzed as in A (n=3). C , DOX induces JNK activation. Cardiomyocytes treated as in A were analyzed for JNK activation by immunoblotting using activation-specific antibodies and cleared whole cell homogenates (n=3). D , SP 600125 inhibits DOX-mediated JNK activation (pJNK Thr183/Tyr185). Cardiomyocytes were treated with SP 600125 (SP; 20 μM in DMSO for 30 min) prior to DOX (1 μM for 30 min) treatment, and then analyzed as in C (n=3). E , SB and SP attenuate DOX-induced Bax translocation to mitochondria. Cardiomyocytes were treated with SB and SP prior to DOX addition (1 μM for 8 h). Bax levels in mitochondria were analyzed by immunoblotting. V-β served as a loading and purity control. F , SB and SP attenuate DOX-induced cardiomyocyte death. Cardiomyocytes were treated with SB and SP prior to DOX addition (1 μM for 24 h). Cell death was analyzed as in 1 A . * P

    Article Snippet: PI3K inhibitors (25 μM in DMSO for 1 h) and wortmannin (250 nM in DMSO for 1 h), Akt inhibitor X (Akti-X, # 124040, 2.5 μM in water for 1 h), p38 MAPK inhibitor SB 203580 (1 μM in DMSO for 30 min), JNK inhibitor SP 600125 (20 μM in DMSO for 30 min), proteasomal inhibitor lactacystin (5 μM in DMSO for 30 min), and DMSO were purchased from EMD Biosciences (Gibbstown, NJ 08027).

    Techniques: Activation Assay, Translocation Assay

    MAP kinases activation is not involved in 4-HNE-elicited lipolytic response. A, Fully-differentiated 3T3-L1 cells were treated with exogenous 4-HNE (20 microM) for indicated time points. The activations of the MAP kinases, including ERK1/2, p38, and JNK, were detected by immunoblotting. B, Fully-differentiated 3T3-L1 cells were pretreated with specific kinase inhibitors for ERK1/2 (U0126, 10 microM), JNK (SP600125, 10 microM), and p38 (SB203580, 10 microM) for 1 hours, followed by exogenous 4-HNE (20 microM) exposure. Glycerol releases were measured 6 hours later. All values are denoted as Means ± SD from three or more independent batches of cells. Bars with different characters differ significantly (p

    Journal: PLoS ONE

    Article Title: Increased 4-Hydroxynonenal Formation Contributes to Obesity-Related Lipolytic Activation in Adipocytes

    doi: 10.1371/journal.pone.0070663

    Figure Lengend Snippet: MAP kinases activation is not involved in 4-HNE-elicited lipolytic response. A, Fully-differentiated 3T3-L1 cells were treated with exogenous 4-HNE (20 microM) for indicated time points. The activations of the MAP kinases, including ERK1/2, p38, and JNK, were detected by immunoblotting. B, Fully-differentiated 3T3-L1 cells were pretreated with specific kinase inhibitors for ERK1/2 (U0126, 10 microM), JNK (SP600125, 10 microM), and p38 (SB203580, 10 microM) for 1 hours, followed by exogenous 4-HNE (20 microM) exposure. Glycerol releases were measured 6 hours later. All values are denoted as Means ± SD from three or more independent batches of cells. Bars with different characters differ significantly (p

    Article Snippet: SP600125 and SQ22536 were purchased from Tocris (Bristol, UK).

    Techniques: Activation Assay

    Effect of MEK and ERK on ERRγ protein levels A, Inhibition of ERK, but not p38 or JNK, reduces exogenous ERRγ expression. MCF7 cells were transiently transfected with the pSG5 empty vector or HA-ERRγ, then treated with DMSO vehicle, 5 μM U0126 (MEK inhibitor), 25 μM SB203580 (p38 inhibitor), or 10 μM SP600125 (JNK inhibitor) for 24 hours prior to lysis and Western blot analysis. Left panels show ERRγ (HA) levels, phosphorylated ERK (pERK), and total ERK from a representative experiment repeated at least twice. Right panels show total and phosphorylated p38 and JNK (p-p38 and pJNK, respectively) from the same experiment. β–actin = loading control. B, Constitutively active, mutant MEK enhances ERRγ protein levels. MCF7 cells were transiently co-transfected with HA-ERRγ and either MEKDD or additional pSG5 empty vector. *denotes the transfected MEKDD construct. β–actin = loading control. C, Exogenous, wild type ERK2 enhances ERRγ protein levels. MCF7 cells were transiently co-transfected with HA-ERRγ and either MEKDD, wild type HA-tagged ERK2, or additional pSG5 empty vector. *denotes the transfected MEKDD construct. The arrowhead and ^ denote transfected HA-ERRγ and HA-ERK2, respectively. β–actin = loading control. D, EGF-mediated enhancement of ERRγ protein levels is reversed by concomitant ERK inhibition. MCF7 cells were transiently transfected with HA-ERRγ, then cultured in low-serum conditions for 20 hours before treatment with DMSO vehicle, 25 ng/ml EGF, or 25 ng/ml EGF plus 5 μM U0126 for 2 hours. β–actin = loading control. E, Inhibition of ERK reduces exogenous ERRγ expression in a second ER+ breast cancer cell line. SUM44 cells were transiently transfected with the pSG5 empty vector or HA-ERRγ, then treated with DMSO vehicle or 5 μM U0126 for 22 hours. β–actin = loading control.

    Journal: The FEBS journal

    Article Title: ERK/MAPK regulates ERRγ expression, transcriptional activity, and receptor-mediated Tamoxifen resistance in ER+ breast cancer

    doi: 10.1111/febs.12797

    Figure Lengend Snippet: Effect of MEK and ERK on ERRγ protein levels A, Inhibition of ERK, but not p38 or JNK, reduces exogenous ERRγ expression. MCF7 cells were transiently transfected with the pSG5 empty vector or HA-ERRγ, then treated with DMSO vehicle, 5 μM U0126 (MEK inhibitor), 25 μM SB203580 (p38 inhibitor), or 10 μM SP600125 (JNK inhibitor) for 24 hours prior to lysis and Western blot analysis. Left panels show ERRγ (HA) levels, phosphorylated ERK (pERK), and total ERK from a representative experiment repeated at least twice. Right panels show total and phosphorylated p38 and JNK (p-p38 and pJNK, respectively) from the same experiment. β–actin = loading control. B, Constitutively active, mutant MEK enhances ERRγ protein levels. MCF7 cells were transiently co-transfected with HA-ERRγ and either MEKDD or additional pSG5 empty vector. *denotes the transfected MEKDD construct. β–actin = loading control. C, Exogenous, wild type ERK2 enhances ERRγ protein levels. MCF7 cells were transiently co-transfected with HA-ERRγ and either MEKDD, wild type HA-tagged ERK2, or additional pSG5 empty vector. *denotes the transfected MEKDD construct. The arrowhead and ^ denote transfected HA-ERRγ and HA-ERK2, respectively. β–actin = loading control. D, EGF-mediated enhancement of ERRγ protein levels is reversed by concomitant ERK inhibition. MCF7 cells were transiently transfected with HA-ERRγ, then cultured in low-serum conditions for 20 hours before treatment with DMSO vehicle, 25 ng/ml EGF, or 25 ng/ml EGF plus 5 μM U0126 for 2 hours. β–actin = loading control. E, Inhibition of ERK reduces exogenous ERRγ expression in a second ER+ breast cancer cell line. SUM44 cells were transiently transfected with the pSG5 empty vector or HA-ERRγ, then treated with DMSO vehicle or 5 μM U0126 for 22 hours. β–actin = loading control.

    Article Snippet: The MEK inhibitor U0126, JNK inhibitor SP600125 and p38 inhibitor SB203580 (Tocris Bioscience, Ellisville, MO) were dissolved in dimethyl sulfoxide (DMSO), stored as 10 and 50mM stocks (respectively) at −20°C, and used at the concentrations indicated.

    Techniques: Inhibition, Expressing, Transfection, Plasmid Preparation, Lysis, Western Blot, Mutagenesis, Construct, Cell Culture

    Influence of kinase inhibitors on ppNOC and NOP mRNA expression. ppNOC and NOP expression in blood leukocytes cultured with PMA or without (control) and with or without pre-treatment with the specific kinase inhibitors PD98059 (PD, ERK inhibitor) 30 μM, SB203580 (SB, p38 inhibitor) 10 μM, SP600125 (SP, JNK inhibitor) 10 μM, Bay 11-7821 (Bay, NFκB inhibitor) 3 μM, or the combination of PD and SB. Box-and-whisker plots, medians with interquartile ranges, and 5 to 95 percentiles, n =10 for the PD+SB+PMA group, n =17 for all other groups. * P

    Journal: Molecular Pain

    Article Title: ERK and p38 contribute to the regulation of nociceptin and the nociceptin receptor in human peripheral blood leukocytes

    doi: 10.1177/1744806919828921

    Figure Lengend Snippet: Influence of kinase inhibitors on ppNOC and NOP mRNA expression. ppNOC and NOP expression in blood leukocytes cultured with PMA or without (control) and with or without pre-treatment with the specific kinase inhibitors PD98059 (PD, ERK inhibitor) 30 μM, SB203580 (SB, p38 inhibitor) 10 μM, SP600125 (SP, JNK inhibitor) 10 μM, Bay 11-7821 (Bay, NFκB inhibitor) 3 μM, or the combination of PD and SB. Box-and-whisker plots, medians with interquartile ranges, and 5 to 95 percentiles, n =10 for the PD+SB+PMA group, n =17 for all other groups. * P

    Article Snippet: Blood was pre-treated with PD98059 (PD) 30 μM, SP600125 (SP) 10 μM, SB203580 (SB) 10 μM, Bay 11-7871 (Bay) 3 μM, or the combination of PD and SB (all from Tocris Bioscience, Bristol, UK) for 1 h prior to culturing with or without PMA 10 ng/ml for 6 and 24 h. The concentrations of the inhibitors were based on doses used in previous studies., , A culture without any stimulus and one cultured only with PMA 10 ng/ml served as control group and reference group, respectively.

    Techniques: Expressing, Cell Culture, Whisker Assay

    JNK maintains high levels of GRP78 in human CCA cells. (A) After treated with SP600125 (SP, 20 µM) for indicated time periods, GRP78 was analyzed using western blot in QBC939, RBE and HCCC-9810 cells. (B) After transfected with siJNK for 60 h, GRP78 was analyzed using western blot in QBC939, RBE and HCCC-9810 cells. (C) After treated with salubrinal (Sal, 25 µM) for 30 h with or without SP600125 (SP, 20 µM) preincubation for 1 h, GRP78 was analyzed using western blot in HepG2 cells. (D) After treated with tunicamycin (Tun, 2.0 µg/ml) for 24 h with or without SP600125 (SP, 20 µM) preincubation for 1 h, GRP78 was analyzed using western blot in HepG2 cells.

    Journal: PLoS ONE

    Article Title: JNK Contributes to the Tumorigenic Potential of Human Cholangiocarcinoma Cells through the mTOR Pathway Regulated GRP78 Induction

    doi: 10.1371/journal.pone.0090388

    Figure Lengend Snippet: JNK maintains high levels of GRP78 in human CCA cells. (A) After treated with SP600125 (SP, 20 µM) for indicated time periods, GRP78 was analyzed using western blot in QBC939, RBE and HCCC-9810 cells. (B) After transfected with siJNK for 60 h, GRP78 was analyzed using western blot in QBC939, RBE and HCCC-9810 cells. (C) After treated with salubrinal (Sal, 25 µM) for 30 h with or without SP600125 (SP, 20 µM) preincubation for 1 h, GRP78 was analyzed using western blot in HepG2 cells. (D) After treated with tunicamycin (Tun, 2.0 µg/ml) for 24 h with or without SP600125 (SP, 20 µM) preincubation for 1 h, GRP78 was analyzed using western blot in HepG2 cells.

    Article Snippet: Chemicals and antibodies JNK inhibitor SP600125 (SP), eIF2α phosphatase enzymes inhibitor salubrinal (Sal) and mTOR inhibitor rapamycin (Rap) were purchased from Tocris Bioscience (Bristol, UK). p70S6K inhibitor PF-4708671 (PF) was purchased from Selleck Chemicals (Houston, TX, USA).

    Techniques: Western Blot, Transfection

    JNK promotes human CCA cells proliferation and invasion. (A) Western blot analysis of phosphorylated JNK and phosphorylated c-Jun in human CCA cells. (B) SP600125 (SP) inhibits JNK activity in human CCA cells. After treated with various doses of SP600125 (SP) for 12 h, phosphorylated c-Jun in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. (C) JNK blocking inhibits human CCA cells proliferation. QBC939, RBE and HCCC-9810 cells were treated with various doses of SP600125 (SP) for indicated time periods. Cell viability was determined by CCK8 assay. (D and E) JNK blocking suppresses human CCA cells migration and invasion. Human CCA cells were treated with SP600125 (SP, 20 µM) for 24 h before transferring to 24-well transwell chambers. The migration (D) and invasion (E) of QBC939, RBE and HCCC-9810 cells with or without SP600125 (SP, 20 µM) treatment were analyzed using transwell assay. Values are means±S.D. Columns, mean of three individual experiments; bars, SD. *Significantly different from control value.

    Journal: PLoS ONE

    Article Title: JNK Contributes to the Tumorigenic Potential of Human Cholangiocarcinoma Cells through the mTOR Pathway Regulated GRP78 Induction

    doi: 10.1371/journal.pone.0090388

    Figure Lengend Snippet: JNK promotes human CCA cells proliferation and invasion. (A) Western blot analysis of phosphorylated JNK and phosphorylated c-Jun in human CCA cells. (B) SP600125 (SP) inhibits JNK activity in human CCA cells. After treated with various doses of SP600125 (SP) for 12 h, phosphorylated c-Jun in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. (C) JNK blocking inhibits human CCA cells proliferation. QBC939, RBE and HCCC-9810 cells were treated with various doses of SP600125 (SP) for indicated time periods. Cell viability was determined by CCK8 assay. (D and E) JNK blocking suppresses human CCA cells migration and invasion. Human CCA cells were treated with SP600125 (SP, 20 µM) for 24 h before transferring to 24-well transwell chambers. The migration (D) and invasion (E) of QBC939, RBE and HCCC-9810 cells with or without SP600125 (SP, 20 µM) treatment were analyzed using transwell assay. Values are means±S.D. Columns, mean of three individual experiments; bars, SD. *Significantly different from control value.

    Article Snippet: Chemicals and antibodies JNK inhibitor SP600125 (SP), eIF2α phosphatase enzymes inhibitor salubrinal (Sal) and mTOR inhibitor rapamycin (Rap) were purchased from Tocris Bioscience (Bristol, UK). p70S6K inhibitor PF-4708671 (PF) was purchased from Selleck Chemicals (Houston, TX, USA).

    Techniques: Western Blot, Activity Assay, Blocking Assay, CCK-8 Assay, Migration, Transferring, Transwell Assay

    JNK promotes the activity of mTOR in human CCA cells. (A) After treated with SP600125 (SP, 20 µM) for indicated time periods, phosphorylated p70S6K in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. Rapamycin (Rap, 20 nM)-treated cholangiocarcinoma cells were used as positive control. (B) After transfected with siJNK for 60 h, phosphorylated p70S6K in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. (C) After treated with SP600125 (SP, 20 µM) for indicated time periods, phosphorylated mTOR in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. (D) After treated with SP600125 (SP, 20 µM) for indicated time periods, phosphorylated Raptor and phosphorylated 4E-BP1 in QBC939, RBE and HCCC-9810 cells were analyzed using western blot.

    Journal: PLoS ONE

    Article Title: JNK Contributes to the Tumorigenic Potential of Human Cholangiocarcinoma Cells through the mTOR Pathway Regulated GRP78 Induction

    doi: 10.1371/journal.pone.0090388

    Figure Lengend Snippet: JNK promotes the activity of mTOR in human CCA cells. (A) After treated with SP600125 (SP, 20 µM) for indicated time periods, phosphorylated p70S6K in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. Rapamycin (Rap, 20 nM)-treated cholangiocarcinoma cells were used as positive control. (B) After transfected with siJNK for 60 h, phosphorylated p70S6K in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. (C) After treated with SP600125 (SP, 20 µM) for indicated time periods, phosphorylated mTOR in QBC939, RBE and HCCC-9810 cells was analyzed using western blot. (D) After treated with SP600125 (SP, 20 µM) for indicated time periods, phosphorylated Raptor and phosphorylated 4E-BP1 in QBC939, RBE and HCCC-9810 cells were analyzed using western blot.

    Article Snippet: Chemicals and antibodies JNK inhibitor SP600125 (SP), eIF2α phosphatase enzymes inhibitor salubrinal (Sal) and mTOR inhibitor rapamycin (Rap) were purchased from Tocris Bioscience (Bristol, UK). p70S6K inhibitor PF-4708671 (PF) was purchased from Selleck Chemicals (Houston, TX, USA).

    Techniques: Activity Assay, Western Blot, Positive Control, Transfection

    JNK/mTOR regulates GRP78 induction through ATF4 in human CCA cells. (A) After treated with SP600125 (SP, 20 µM) for 48 h, ATF4 and phosphorylated eIF2α in QBC939, RBE and HCCC-9810 cells were analyzed using western blot. (B) After treated with rapamycin (Rap, 20 nM) for 48 h, phosphorylated eIF2α and phosphorylated p70S6K in QBC939, RBE and HCCC-9810 cells were analyzed using western blot. (C) After treated with salubrinal (Sal, 25 µM) for 30 h with or without SP600125 (SP, 20 µM) and rapamycin (Rap, 20 nM) preincubation for 1 h, ATF4 and phosphorylated eIF2α were analyzed using western blot in HepG2 cells. (D) After treated with PF-4708671 (PF, 10 µM) and 4EGI-1 (50 µM) for 48 h, GRP78 and ATF4 in QBC939, RBE and HCCC-9810 cells were analyzed using western blot. (E) QBC939, RBE and HCCC-9810 cells were treated with rapamycin (Rap, 20 nM) for 12 h, and ATF4 and GRP78 mRNA levels were analyzed using real-time RT-PCR. Values are means±S.D. Columns, mean of three individual experiments; bars, SD. *Significantly different from control value.

    Journal: PLoS ONE

    Article Title: JNK Contributes to the Tumorigenic Potential of Human Cholangiocarcinoma Cells through the mTOR Pathway Regulated GRP78 Induction

    doi: 10.1371/journal.pone.0090388

    Figure Lengend Snippet: JNK/mTOR regulates GRP78 induction through ATF4 in human CCA cells. (A) After treated with SP600125 (SP, 20 µM) for 48 h, ATF4 and phosphorylated eIF2α in QBC939, RBE and HCCC-9810 cells were analyzed using western blot. (B) After treated with rapamycin (Rap, 20 nM) for 48 h, phosphorylated eIF2α and phosphorylated p70S6K in QBC939, RBE and HCCC-9810 cells were analyzed using western blot. (C) After treated with salubrinal (Sal, 25 µM) for 30 h with or without SP600125 (SP, 20 µM) and rapamycin (Rap, 20 nM) preincubation for 1 h, ATF4 and phosphorylated eIF2α were analyzed using western blot in HepG2 cells. (D) After treated with PF-4708671 (PF, 10 µM) and 4EGI-1 (50 µM) for 48 h, GRP78 and ATF4 in QBC939, RBE and HCCC-9810 cells were analyzed using western blot. (E) QBC939, RBE and HCCC-9810 cells were treated with rapamycin (Rap, 20 nM) for 12 h, and ATF4 and GRP78 mRNA levels were analyzed using real-time RT-PCR. Values are means±S.D. Columns, mean of three individual experiments; bars, SD. *Significantly different from control value.

    Article Snippet: Chemicals and antibodies JNK inhibitor SP600125 (SP), eIF2α phosphatase enzymes inhibitor salubrinal (Sal) and mTOR inhibitor rapamycin (Rap) were purchased from Tocris Bioscience (Bristol, UK). p70S6K inhibitor PF-4708671 (PF) was purchased from Selleck Chemicals (Houston, TX, USA).

    Techniques: Western Blot, Quantitative RT-PCR