Gw9662

Human peripheral blood mononuclear cells were isolated from the blood of healthy volunteers by a density gradient centrifugation method on Lymphoprep (Abcys). Monocytes were isolated from mononuclear cells by adherence to plastic for 2 h in special macrophage serum-free medium (SFM; Life Technologies) with l-glutamine at 37°C in a humidified atmosphere containing 5% CO₂. Non-adherent cells were removed by washing with Hanks’ balanced salt solution (HBSS) (Gibco, Invitrogen), and the remaining adherent cells (>85% monocytes) were incubated in SFM medium. The h-MDMs were obtained after 24 h of culture in SFM medium. Adherent h-MDMs were pre-incubated 30 min or not before the addition of P17 (200 µg/ml) with GW9662 (1 nM; Santa Cruz Biotechnology), MAFP (20 µM; Calbiochem), MK-886 (10 µM; Calbiochem), N-acetyl-cysteine (10 mM; Sigma), or Z-Vad-FMK (ZVAD) (50 µM; Calbiochem) for 30 min.

Pioglitazone hydrochloride (Sigma Aldrich, St. Louis, MO, USA) and GW9662 (Santa Cruz, CA, USA) were dissolved in dimethyl sulfoxide (DMSO) and stored at −20°C until use. Glioma cells were treated at a number of drug concentrations and times as indicated in the results section.

HepG2 cells were obtained from ATCC (HB-8065; 41106514, H87), and grown in DMEM medium supplemented with 10% (v/v) fetal bovine serum (Invitrogen Life Technologies-GIBCO, Carlsbad, CA, USA), 100 U/mL penicillin, 100 g/mL streptomycin, and maintained under humidified atmosphere of 5% CO₂ at 37 °C. Cells were incubated with 500 μM PFD, 1 μM GW7647 (PPARγ agonist; Santa Cruz Biotechnology, Santa Cruz, CA, USA), and 100 nM GW9662 (PPARγ antagonist; Santa Cruz Biotechnology) for 24 h to evaluate PFD effect on PPARα and PPARγ proteins expression. Incubation was performed after 8 h of fetal bovine serum starving.