Cilia Dynamics and Forces

We have measured the motion of 2.8 µm magnetic beads bound to the tips of the human bronchial epithelial cilia, experiencing an external load provide by the 3-Dimensional Force Microscope. Human bronchial epithe-lial (HBE) cells are cultured until fully ciliated (~ 6 weeks). The cultures are washed thoroughly to remove the mucus layer. The cilia are then coated with biotinylated wheat germ agglutinin, which facilitates the binding of then 1 and 2.8 µm diameter streptavidin coated magnetic beads to the cilia. Forces ranging from 10’s of pN to > 1 nN are then applied to the beads over time scales of 2-5 seconds. To ensure binding to single cilium, we have developed a new binding strategy we call “spot labeling” which provides a ~100nm active area for bind-ing. Our data shows that over time scales on the order of seconds, the amplitude of a cilia-bound bead de-creases while maintaining a constant frequency. The application for a simple model of cilia dynamics allows us to assign a generated effective force of 65pN at the cilia tip. Most interesting, the applied force does not speed up the cilium motion in the direction of the applied force. The cilium slows down whether beating in the direc-tion of the force or opposed to it. These results are being tested against a computational model of the axoneme coded by Mitran that includes the 9+2 architecture of the axoneme and the appropriate distribution of force ge-nerators.