BMP4 suppression of normal airway BC differentiation. To mimic the effects of the up-regulated BMP4 observed in the airway epithelium in asymptomatic smokers and COPD smokers, BMP4 (10 ng/ml) was added to the basolateral side when established the ALI cultures, and expression of airway epithelial differentiation-related genes and protein were assessed. A. TaqMan assessment of fold-change (log2) in the mRNA expression of ciliated cell differentiation-related genes (FOXJ1, DNAI1, MCIDAS and RFX2) and secretory cell differentiation-related genes (SCGB1A1, SPDEF, MUC5B and TFF3) on the airway epithelium after 14 days ALI culture with BMP4 treatment vs untreated control. * p<0.05, ** p<0.01, *** p<0.001, n=3 or 4. B. Immunofluorescence top staining of ciliated cell marker DNAI1 (red) and secretory cell marker SCGB1A1 (green) on the airway epithelium after 28 days ALI culture with BMP4 (10 ng/ml) stimulation from the basolateral side (right) vs untreated control (left). Top – DNAI1 (red), middle – SCGB1A1 (green), bottom – overlapping of DNAI1 and SCGB1A1. Nuclei are stained with DAPI (blue). Scale bar – 20 μm.