Cilia dynamics creates a dynamic barrier to penetration of the periciliary layer in human airway epithelia

Abstract

The ciliated epithelium of the human respiratory tract is covered by the airway surface liquid (ASL), a protective fluid consisting of two layers: the periciliary layer (PCL), where motile cilia reside and generate fluid flow, and an overlying mucus layer. The complex structure and stratified nature of the PCL complicate both the prediction and quantification of fluid flow at the scale of individual or small groups of cilia, making it difficult to connect microscopic flows to macroscopic clearance. To tackle this challenge, we developed a novel methodology that involves un-caging a fluorescent compound to trace the flow field within the PCL. Fluorescence is activated at micrometric spots within the cilia layer, and displacement vectors and diffusion are recorded using high-speed video. Our experiments reveal a complex fluid transport pattern, with displacement velocity along the epithelial surface varying due to a non-uniform vertical flow field. Additionally, we observed that cilia expel fluid at their tips, a mechanism likely aimed at preventing pathogen access to the epithelium. Simulations, where cilia are modeled as arrays of rigid rods with length asymmetry, support these findings and offer new insights into the dynamics of fluid transport in the respiratory tract and the critical role of cilia coordination. Significance StatementThis study introduces an experimental pipeline to investigate fluid velocity and diffusion within the PCL of the human respiratory tract. By integrating experimental data with simulations of ciliary motion, we offer a robust framework to understand how cilia, depending on their collective beating properties, propel periciliary fluid in this structurally and dynamically complex environment. Our findings significantly expand the understanding of ciliary function, revealing that when cilia are beating coherently near cilia tips fluid is actively driven away from the epithelial surface. This suggests that coordinated cilia movement not only plays a key role in maintaining respiratory health by clearing mucus, but may also provide a dynamic barrier against pathogen entry.