Breathing on Chip: Dynamic flow and stretch tune cellular composition and accelerate mucociliary maturation of airway epithelium in vitro

Abstract Human lung function is intricately linked to blood flow and breathing cycles, but it remains unknown how these dynamic cues shape human airway epithelial biology. Here we report a state-of-the-art protocol for studying effects of physiological airflow and stretch on differentiation, cellular composition, and mucociliary clearance of human primary airway epithelial cells cultured on a perfused airway chip. Perfused epithelial tissue cultures developed a large airway-like cellular composition with accelerated maturation and polarization of mucociliary clearance when compared to traditional (static) culture methods. Additional application of airflow and stretch to the airway chip resulted in a cellular composition more comparable to the small(er) airways, reduced baseline secretion of interleukin-8 and other inflammatory proteins, and reduced gene expression of matrix metalloproteinase (MMP) 9, fibronectin, and other extracellular matrix factors. These results indicate that breathing-like mechanical stimuli are important modulators of airway epithelial cell differentiation and homeostasis and that their fine-tuned application could generate models of specific epithelial regions, pathologies, and mucociliary (dys)function.