Expansion of Airway Basal Cells and Generation of Polarized Epithelium.

Airway basal stem cells are the progenitor cells within the airway that exhibit the capacity to self-renew and give rise to multiple types of differentiated airway epithelial cells. This stem cell-derived epithelium displays organized architecture with functional attributes of the airway mucosa. A protocol has been developed to culture and expand human airway basal stem cells while preserving their stem cell properties and capacity for subsequent mucociliary differentiation. This achievement presents a previously unrealized opportunity to maintain a durable supply of progenitor cells derived from healthy donors to differentiate into human primary airway epithelium for cellular and molecular-based studies. Further, basal stem cells can be harvested from patients with a specific airway disease, such as cystic fibrosis, enabling investigation of potentially altered behavior of disease-specific cells in the appropriate context of the airway mucosa. Here we describe, in detail, a protocol for the serial expansion of airway basal stem cells to enable the generation of nearly unlimited airway basal cells that can be stored and readily available for subsequent culturing and differentiation. In addition, we describe culturing and differentiation of airway basal stem cells on permeable transwell filters at air-liquid interface to create functional mucociliary pseudostratified polarized airway epithelial mucosa.