Human umbilical vein ECs (HUVECs) were isolated as described previously [10 (link), 26 (link)]. HUVECs from 3 to 4 cords were pooled for experiments, and passage 2–3 HUVECs were used for all experiments. HUVEC culture medium included M199 media (Invitrogen, Carlsbad, CA) supplemented with 10 % fetal bovine serum (FBS) (Invitrogen), 30 µg/ml endothelial cell growth factor supplement (ECGF) (BD Biosciences, San Jose, CA), 100 µg/ml heparin sodium (Sigma, St Louis, MO), 1 % sodium bicarbonate (Invitrogen), 1 % Glutamax™-1 (Invitrogen), 1 % penicillin/streptomycin (Invitrogen), 0.25 % fungizone (Invitrogen), and 0.25 % gentamicin (Invitrogen). Cells were cultured at 37 °C with 5 % CO2 and humidity. In some experiments, cells were treated with VEGF-A 165 (R&D Systems, Minneapolis, MN) or chemically defined lipid mixture 1 (Sigma). For induction of apoptosis, HUVECs were cultured in medium containing 5 % FBS and no ECGF, or treated with TNFα (40 ng/ml) for 24 h. PPARδ agonist GW0742 and PPARδ inhibitor GSK0660 were purchased from Cayman Chemicals (Ann Arbor, MI) and R&D Systems, respectively.
Gw0742
Manufactured by Cayman Chemical
Sourced in United States
GW0742 is a synthetic organic compound that functions as a selective agonist for the peroxisome proliferator-activated receptor delta (PPAR-delta). It is used in scientific research to study the effects of PPAR-delta activation.
Automatically generated - may contain errors
Lab products found in correlation
Gw501516, by Cayman Chemical (2 mentions)
Chemically defined lipid mixture 1, by Merck Group (1 mentions)
Sodium bicarbonate, by Thermo Fisher Scientific (1 mentions)
Gentamicin, by Thermo Fisher Scientific (1 mentions)
Heparin sodium, by Merck Group (1 mentions)
Fungizone, by Thermo Fisher Scientific (1 mentions)
Glutamax 1, by Thermo Fisher Scientific (1 mentions)
M199 media, by Thermo Fisher Scientific (1 mentions)
Vegf a165, by R&D Systems (1 mentions)
Gsk0660, by Cayman Chemical (1 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (1 mentions)
Gw1929, by Merck Group (1 mentions)
Pht 427, by Selleck Chemicals (1 mentions)
Gsk3787, by Santa Cruz Biotechnology (1 mentions)
Pd98059, by Cell Signaling Technology (1 mentions)
Fh535, by Bio-Techne (1 mentions)
Ly294002, by Merck Group (1 mentions)
Quinine, by Cayman Chemical (1 mentions)
Catalase, by Merck Group (1 mentions)
Anti rabbit igg, by Cell Signaling Technology (1 mentions)
5 protocols using gw0742
1
Isolation and Culture of Human Umbilical Vein Endothelial Cells
2
Macrophage Inflammasome Activation by Palmitic Acid
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Recombinant mouse Il-4 (rIl-4, 10 ng/ml, Peprotech, Rocky Hill, NJ, USA), GW501516, GW0742 (0.1 μM for both, Cayman Chemical, Ann Arbor, MI, USA), GW1929 (1 μM, Sigma, St. Louis, MO, USA), or vehicle (DMSO for Ppar agonists) were incubated with macrophages overnight in 10% FBS, DMEM. PA (Sigma) was prepared as previously described [26] (link) and added to cells at final concentrations of 300 μM in 0.45% BSA, 2% double stripped FBS, DMEM for 16 h. For PA-induced inflammasome activation, lipopolysaccharide (LPS, 10 ng/ml, Sigma) was pre-incubated with macrophages for 3 h in 10% FBS, DMEM before the addition of PA.
3
Neonatal and Adult Cardiomyocyte Isolation and Culture
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The investigation conforms with the Guide for the Care and Use of Laboratory Animals published by the US National Institutes of Health (NIH Publication No. 85-23, revised 1996). The local Committee approved animal experiments for Care and Use of Laboratory Animals (Regierungspräsidium Darmstadt, Gen. Nr. B 2/Anz. 75). Ventricular cardiomyocytes from 3-day-old (P3), 8-day-old (P8) and 12-week-old (adult) Sprague-Dawley rats were isolated and cultured as described55 (link),56 (link). Cells were initially cultured for 48-72 h in the presence of 20 μM cytosine-𝒟-arabinofuranoside and 5% horse serum before stimulation to prevent non-myocyte proliferation. Cardiomyocytes were subsequently treated in the presence of serum (0.2% FCS: neonatal, 5% FCS: adult) with DMSO as a negative control, 1 μM Carbacyclin if not stated otherwise (Enzo Life Science) or 100 nM GW0742 (Cayman Chemical). Postnatal cardiomyocytes were stimulated once; adult cardiomyocytes daily while the medium was exchanged every 3 days. As positive control 50 μg/ml FGF1 + 5 μM SB203580 (p38i) were used. Signaling pathway inhibitors were added 1 h before stimulation: 20 μM PHT-427 (Selleckchem), 100 nM GSK3787 (Santa Cruz Biotechnology), 15 μM FH535 (Tocris Bioscience), 10 μM LY294002 (Sigma), 20 μM PD98059 (Cell Signaling).
4
Topical Formulation of PY-60 Compound
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Quinine, GW0742, and ATRA were from Cayman Chemical. Topical PY-60 was formulated (1.5 or 0.15 weight percent) as a gel (propylene glycol: 26%, Labrasol ALF: 17.5%, PEG 400: 46.5%, Povidone K-90: 8.5%) at Aragen Life Sciences. The methods to synthesize the batch of PY-60 used for the gel formulation used in this work is provided in the SI Appendix .
5
PPARδ Antagonist-Mediated Oxidative Stress
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PT-S58 (PPARδ antagonist), catalase, thapsigargin and β-actin antibodies were from Sigma-Aldrich (St Louis, MO, USA). DCFH was from Life-Tech (Carlsbad, CA, USA) while 7-aminoactinomycin D was from Biolegend (San Diego, CA, USA). GW0742 and GW501516 (PPARδ agonists) were from Cayman Chemical (Ann Arbor, MI, USA). Dexamethasone (Omega, Montreal, QC, Canada), insulin (Eli Lilly, Toronto, ON, Canada) and interferon-α2b (Schering Canada Inc., Pointe-Claire, QC, Canada) were purchased from the hospital pharmacy. AKT inhibitor IV was from Calbiochem (San Diego, CA, USA). DG172 (PPARδ antagonist) has been previously described.28 (link) NXT1511 (PPARδ antagonist) was provided by Peppi Prasit (Inception, San Diego, CA, USA).
Antibodies to PERK, PDPK1, p-AKT(T308), AKT, p-SAPK/JNK (T183/Y185), CHOP, anti-Rabbit IgG and anti-Mouse IgG were from Cell Signaling Technology (Danvers, MA, USA). The PPARδ antibody (101720) was from Cayman.
Antibodies to PERK, PDPK1, p-AKT(T308), AKT, p-SAPK/JNK (T183/Y185), CHOP, anti-Rabbit IgG and anti-Mouse IgG were from Cell Signaling Technology (Danvers, MA, USA). The PPARδ antibody (101720) was from Cayman.
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