. In brief, positions at the base and tips of cilia at the beginning and end of an active phase of a beat cycle were recorded. Ciliary beat frequency was calculated by fast Fourier transform and video-kymography. This analysis allowed the calculation of ciliary beat pattern parameters, such as cilium length, angle of beating and beating amplitude.
. In brief, epithelial edges were identified in each sample, investigated for protrusions or abnormalities, and scored from 1 to 4, where 1 indicates a normal ciliated edge, 2 indicates a ciliated edge with minor projections, 3 indicates a ciliated edge with major projections, and 4 is for single, unattached, ciliated cells.Cells obtained from nasal-scrape brushings were seeded into collagen-coated wells containing PneumaCult-Ex Plus medium .
Before deciliation, differentiated ALI cultures were washed twice with PBS for 5 min to remove cell debris and mucus. Ice-cold PBS was then added to both compartments of the culture dish and the dish was placed on ice. After 5 min, the PBS solution was removed and 50 μl ice-cold deciliation buffer was added to the cells of each well. After incubation for 2 min without shaking, the cilia-containing solution was transferred to a microcentrifuge tube.