RPMI 1640, penicillin/streptomycin solution, fetal bovine serum (FBS) were obtained from Invitrogen (Camarillo, CA, USA). Ketoconazole (KTZ) and sunitinib were purchased from Selleckchem (Houston, TX). U0126 and SB203580 were procured from Promega (Madison, WI, USA) and SP600125 from Sigma (Sigma-Aldrich, St. Louis, MO, USA). Unless otherwise indicated, all other drugs were purchased from Sigma (St. Louis, MO, USA)11 (link),12 (link).
Sb203580
Manufactured by Promega
Sourced in United States
SB203580 is a small molecule inhibitor that targets the p38 MAPK signaling pathway. It functions by selectively and potently inhibiting the p38α and p38β isoforms of the p38 MAPK enzyme.
Automatically generated - may contain errors
Lab products found in correlation
U0126, by Promega (7 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (5 mentions)
Ly294002, by Promega (5 mentions)
Rpmi 1640, by Thermo Fisher Scientific (3 mentions)
Sp600125, by Merck Group (3 mentions)
Penicillin streptomycin solution, by Thermo Fisher Scientific (2 mentions)
Tunicamycin, by Merck Group (2 mentions)
Rpmi 1640 medium, by Merck Group (2 mentions)
Sp600125, by Promega (2 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Pd98059, by Promega (2 mentions)
Recombinant human apn full length apn or gapn, by ProSpec (2 mentions)
Sunitinib, by Selleck Chemicals (1 mentions)
Ketoconazole, by Selleck Chemicals (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Merck Group (1 mentions)
Apigenin, by Merck Group (1 mentions)
Luteolin, by Merck Group (1 mentions)
N acetylcysteine, by Merck Group (1 mentions)
Chrysin, by Merck Group (1 mentions)
24 protocols using sb203580
1
Pharmacological Inhibition Assay
2
Oxidative Stress and Cellular Pathways
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4-Hydroxy-trans-2-nonenal (4-HNE) was purchased from Cayman Chemical (Ann Arbor, MI). Luteolin (≥98%), apigenin (≥97%), chrysin (≥97%), MTT (3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide), RPMI-1640 medium, H2O2, N-acetyl-cysteine (NAC), tunicamycin (Tm), protein kinase A (PKA) inhibitor H-89 [N-[2-((p-Bromocinnamyl)amino)ethyl]-5-isoquinolinesulfonamide] and other chemicals were purchased from Sigma-Aldrich Co. (St. Louis, MO), unless otherwise indicated. SB 203580 (4-[4'-fluorophenyl]- 2-[4'-methylsulfinylphenyl]-5-[4'-pyridyl] imidazole), a p38 MAP kinase inhibitor was purchased from Promega (Madison, WI). MEK inhibitor U0126, and c-Jun NH2-terminal kinase (JNK) inhibitor SP600125 (anthra[1,9-cd]pyrazole-6 (2H)-one) were purchased from Calbiochem (San Diego, CA).
3
Investigating Cellular Signaling Pathways
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DON was obtained from Sigma-Aldrich (St. Louis, MO, USA). Cell culture medium and supplements were purchased from Life Technologies (Grand Island, NY, USA). Anti-phospho-p38 (4511), anti-p38 (8690), anti-phospho-JNK (4668), anti-JNK (9252), anti-phospho-ERK (4370), anti-ERK (4695) and anti-β-actin (4970) antibodies were purchased from Cell Signaling Technology (Beverly, MA, USA). SB203580 was obtained from Promega (Madison, WI, USA). U0126 and SP600125 were acquired from Cayman Chemicals (Ann Arbor, MI, USA).
4
Resveratrol Modulates Egr-1 Expression
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The final concentrations of all the inhibitors were 10 μM except SP 600125 (BIOMOL International, PA), of which the concentration was 20 μM. SB 203580 (Promega, WI) inhibits p38MAPK/SAP2, whereas SP 600125 is a potent JNK inhibitor. U-0126 (Santa Cruz Biotechnology, Inc, TX) is a potent inhibitor of ERK1/2 activity and LY 294002 (Santa Cruz Biotechnology, Inc, TX) blocks PI3K/AKT pathways. Cells were treated with or without 100 μM of resveratrol for another 6 h after 1 h preincubation, and the expression of Egr-1 protein and β-actin (loading control) was examined by western blotting.
5
Anticancer Drug 5-FU and p38 MAPK Inhibitor Evaluation
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The anticancer drug 5-FU was purchased from Kyowa Hakko Bio Co., Ltd. (Tokyo, Japan). A selective p38 MAPK inhibitor, SB203580 (20 (link)), was purchased from Promega Corporation (Madison, WI, USA). The human colon cancer cell lines, HCT116, HCT15 and HT29, were obtained from the American Type Culture Collection (Manassas, VA, USA). The cells were grown in RPMI-1640 medium (Gibco-BRL, Carlsbad, CA, USA). Each culture was supplemented with 10% heat-inactivated fetal bovine serum (FBS; CSL Ltd., Melbourne, Australia) and 1% penicillin/streptomycin (1 ml) in a humidified 5% CO2 incubator at 37°C. Prior to the experiments, the cells were incubated without FBS for 24 h.
6
Culturing Keratinocytes and XPC-deficient Fibroblasts
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Human HaCaT keratinocytes (kindly provided by Prof. N. Fusenig), and XPC-deficient (XPCNull) immortalized skin fibroblasts (GM15983, also known as XP4PA-SV-EB, obtained from Coriell) were cultured in a monolayer in 95% air/5% CO2 (vol/vol) at 37°C in Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum (FBS), 100 units/ml penicillin and 100 μg/ml streptomycin (Invitrogen). HaCaT cells passaged <40 times were used. Where indicated, cells were treated with the inhibitors rapamycin (25 nM, LC Laboratories, R-5000), CX-4945 (5 μM, Selleck Chemicals, S2248), KU-60019 (1 μM, Selleck Chemicals, S1570), LY294002 (10 μM, Promega, V1201), NU7441 (1 μM, Selleck Chemicals, S2638), TCS JNK 60 (10 μM, Torcis, 3222), SB203580 (10 μM, Promega, V1161), PD98059 (20 μM, Promega, V1191), Y27632 (10 μM, Torcis, 1254) or with the vehicle (DMSO, Sigma-Aldrich). The concentrations of the kinase inhibitors were chosen based on our previously used concentrations and from literature (23 (link),32–40 (link)).
7
Cell Culture and Inhibitors Evaluation
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The ESCC cell line EC109 was cultured in 199 medium (Invitrogen, Carlsbad, USA) containing 10% (v/v) fetal bovine serum and maintained at 37°C in a humidified 5% CO2 atmosphere. TPA was purchased from Sigma-Aldrich (St. Louis, MO, USA). DMSO was purchased from Amresco Company (Solon, Ohio, USA). Empty pcDNA3 vector was purchased from Invitrogen. PKCα/β kinase inhibitors (myristoylated protein kinase C peptide inhibitor), MEK kinase inhibitors (U0126, PD98059), and p38 kinase inhibitors SB203580 were purchased from Promega (Madison, WI, USA). The c-Jun N-terminal kinase (JNK) inhibitor SP600125 was purchased from Calbiochem (La Jolla, CA, USA). Antibodies against phospho-ERK1/2 (p-ERK1/2), ERK1/2, and JNK were purchased from Santa Cruz Biotechnology (Santa Cruz, USA). β-Actin was purchased from Sigma-Aldrich (St. Louis, MO, USA). The pMD19-T Simple vector, SYBR Premix Ex-Taq™, and SYBR Primescript RT-PCR Kit were purchased from TaKaRa (Dalian, China).
8
Fisetin Treatment in Cell Culture
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Fisetin (3,3′,4′,7-tetrahydroxyflavone), dimethyl sulfoxide (DMSO), poly-L-lysine, thiazoly blue tetrazolium bromide (MTT), nonessential amino acids (NEAAs), RPMI-1640 medium, and other chemicals were purchased from Sigma–Aldrich (St. Louis, MO, USA) unless otherwise indicated. Corticosterone and SP600125 were purchased from Enzo Life Sciences (Farmingdale, New York, NY, USA). Horse serum (HS) and fetal bovine serum (FBS) were purchased from Thermo Fisher Scientific (Rockford, IL, USA). U0126, SB203580 and LY294002 were purchased from Promega (Madison, WI, USA).
9
Anisomycin and Kinase Inhibitor Assay
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TQ (Catalog no. 274 666) and anisomycin were purchased from Sigma Aldrich, St Louis, MO. U0126 and SB203580 were purchased from Promega, Madison, WI.
10
Linoleic Acid Activation of FFA1
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Linoleic acid and GW9508 were purchased from Sigma-Aldrich (St, Louis, MO, USA). EGF was obtained from ThermoFisher (Frederick, MD, USA). Phospho-ERK, phospho-p38, total ERK, total p38, and GAPDH antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). FFA1 antibody was obtained from Abcam (ab75197, USA). Inhibitors UO126 and SB203580 were obtained from Promega (Madison, WI, USA).
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