Kinematics and Kinetics of the Ciliary Waveform in Human Tracheal Epithelial Cells

Abstract

Cilia function is critical to human health. We developed quantitative methods to describe waveform, propulsive force, and net work of cilia on multi-ciliated human tracheal epithelial cells (HTEC).Lung tracheobronchial epithelial cells were isolated from healthy donor subjects for culture and differentiated using growth factor-enriched medium and air-liquid interface conditions to yield multi-ciliated cells. Detached cells in 20 µL of culture medium at room temperature were sandwiched between a glass slide and coverslip and lateral views of beating cilia were acquired (Zeiss Axiophot, 100X, oil immersion, DIC) for 2s at 100 fps (320x240 pixels, 15.07 pixels/µm). Waveforms were analyzed in N=21 cilia from 3 subjects. In each video frame, 30 points evenly distributed from the base to the tip of the cilium were fitted to a 4th-order polynomial curve. The resulting parameter sets were Fourier-transformed and filtered temporally to obtain a space-time surface representing a typical beat. Local velocity vectors and curvature were computed by differentiation of this surface, and local fluid forces on the cilium were estimated using resistive force theory: fN = CNvN and fT = CTvT (CN≈1.5x10−3 pN-s/μm; CT ≈0.7x10−3 pN-s/μm). Power was calculated from the product f·v. Beat frequency, curvature, and shear amplitude were also computed.Waveforms of active cilia were asymmetric with distinct power and recovery strokes. The average propulsive force and work for active HTEC cilia were: Fprop=3.73±4.42pN, W=1.63±1.84aJ. We also identified apparently passive cilia with nearly symmetric waveforms and little propulsive effectiveness (Fprop= −0.02 ±1.52 pN, W=0.33±0.24aJ). Active cilia were longer (6.48 ±0.66 vs 5.09±0.73µm) and beat more rapidly (3.54 ±0.77Hz vs 2.64±1.16Hz) than passive cilia. It is not clear whether passive cilia are immature or simply represent normal heterogeneity. These results provide baseline data for future studies of motility defects.