Smad signaling mediated BMP4-induced squamous cell differentiation. A. Western blot analysis shows the phosphorylated Smad 1/5/8 (upper) and total Smad 1 (lower) protein expression of the airway BC in ALI culture (day 0) with or without BMP4 (10 ng/ml) stimulation for 30 min, 1, 5 and 24 hours. BMP4 solvent HCl and EGF (10 ng/ml; EGF does not activate BMP4 downstream Smad signaling [23] were used as negative controls. B. TaqMan assessment of the squamous cell markers IVL (right) and KRT14 (left) in BC with BMP4 (10 ng/ml) stimulation vs untreated control for 48 hr while silencing Smad4 by siRNA. Negative control siRNA was used as negative control, and the data were normalized to the untreated control (log2), n=4. * p<0.05, *** p<0.001.