“…Human PC346C cells (van Weerden et al, 1996;Marques et al, 2006) were cultured in DMEM-F12 medium (Cambrex BioWhittaker, Verviers, Belgium) supplemented with 2% FBS (PAN Biotech, Aidenbach, Germany), 1% insulintransferrin-selenium (GIBCO-BRL/Invitrogen), 0.01% bovine serum albumin (Boehringer Mannheim, Mannheim, Germany), epidermal growth factor (10 ng/ml; SigmaAldrich), penicillin-streptomycin antibiotics (penicillin, 100 U/ml; streptomycin, 100 mg/ml; Cambrex BioWhittaker) plus the following additions: fibronectin (100 ng/ml; Harbor Bio-Products, Tebu-bio, Boechout, Belgium), fetuin (20 ng/ ml; ICN Biomedicals, Zoetermeer, The Netherlands), cholera toxin (50 ng/ml), 0.1 mM phosphoethanolamine, triiodothyronine (0.6 ng/ml), dexamethasone (500 ng/ml) (all from Sigma-Aldrich), and 0.1 nM 17-methyltrienolone (R1881; New England Nuclear/PerkinElmer Life Sciences, Boston MA). Cells were grown in T25 Primaria tissue culture flasks (BD Biosciences, Benelux, The Netherlands) at 37°C under a 5% CO 2 humidified atmosphere.…”