Ex-vivo culture of fetal lung explants Transgenic E12.5 lung explants were isolated and cultured, for 8 or 24 hours, in a 1:1 mixture of DMEM: F12 (GiBCO, 11039), supplemented with Insulin-Transferrin-Selenium (Gibco, 51300-044) and antibiotics. Explants were placed on Nuclepore Track-Etched Membranes (Whatman, 110414) and they were incubated in 4-well plates (Thermo Scientific, 144444). LDN193189 (Axon MedChem, Axon 1509) and SB431542 (Sigma-Aldrich, S4317) were used at a concentration of 10 mM. Untreated and vehicle (DMSO, Sigma-Aldrich, D2650) treated lungs were used as controls. In some experiments, E12.5 lung explants were treated with 100 ng/ml BMP4 (Peprotech, 120-05ET), or 250 ng/ml FGF10 (Peprotech, 100-26) or 500 nM Sonic Hedgehog (Shh) agonist-Purmorphamin (Calbiochem, 540220) for 8 hours.