5 aza 2 deoxycytidine

Defined compounds used for validation experiments were ordered de novo from various suppliers: SB-431542, PD-173074, U0126, Salinomycin, Nigericin, PD-161570, PD-166285, PD-166866, Idoxuridine and 5-Azacytidine (all from Sigma); Y-27632 2HCl, GSK429286A, Nintedanib, Pazopanib, Sorafenib, Vatalanib, Axitinib, Sunitinib, Erlotinib, Lapatinib, Gefitinib, Dasatinib, Crizotinib, Vandetanib, AEE788, Cediranib, AZD4547, CP673451, TSU-68 and KX2-391 (all from Selleck Chemicals); PP1, PP2, Rho kinase inhibitor V, SR-3677, GSK269962, SB-772077B (all from Tocris); Fasudil from Enamine; CAY10622 and PD-166326 from Cayman; BGJ398 from Axon Medchem; 5-Aza-2′-deoxycytidine from Chem-Impex. The reordered compounds were tested in extended dilution series ranging from 25 μM to 0.04 nM on TGFβ-treated NMuMG cells and from 25 μM to 1 nM on TGFβ-treated Py2T cells.

Materials were purchased from the following sources: fibroblast growth factor (FGF) 2 and activin A (PeproTech, Rocky Hill, NJ, USA); iMatrix-511 (Nippi, Tokyo, Japan); CHIR99021 and Y-27632 (Focus Biomolecules, Plymouth Meeting, PA, USA); KSR, DMEM, DMEM/F-12, Advanced DMEM/F-12, N2 supplement and B27 serum-free supplement (Thermo Fisher Scientific, Waltham, MA, USA); FBS (Nichirei Biosciences, Tokyo, Japan); R-spondin 1, Noggin and epidermal growth factor (EGF) (GenScript, Piscataway, NJ, USA); N-acetylcysteine (Sigma-Aldrich, St Louis, MO, USA); 8-Br-cAMP (Enzo Life Sciences, New York, NY, USA); PD98059 and A-83-01 (AdooQ BIOSCIENCE, Irvine, CA, USA); 5-aza-2′-deoxycytidine (Chem-impex International, Wood Dale, IL, USA); L-glutamine, minimum essential medium nonessential amino acid solution (NEAA), penicillin-streptomycin solution, valproic acid and nicotinamide (Wako Pure Chemical Industries, Osaka, Japan); and total RNA from human small intestine samples (5 donors; BioChain Institute, Newark, CA, USA). All other reagents were of the highest quality available, including SB 202190 (ChemScene, Monmouth Junction, NJ, USA) and WNT3a (ATGen Co. Ltd., Gyeonggi-do, South Korea).