Generation of Lung Epithelial-Like Tissue from hESC by AirLiquid Interface Culture

Human embryonic stem cells (hESCs) have the capacity to differentiate in vivo and in vitro into cells from all three germ lineages. The in vitro generation of lung cells and tissues from hESCs creates opportunities for fundamental research, drug development or cell-replacement therapy. In this chapter, we describe a reliable and simple protocol to differentiate hESCs into the major cell types of lung epithelial tissue. In this protocol, undifferentiated hESCs, grown on a porous membrane in hESC medium for 4 days, are switched to a differentiation medium and cultured for 4 days, followed by differentiation in air–liquid interface conditions for 20 days. The expression of several lung markers is analysed using quantitative real-time PCR and confirmed by immunohistochemistry. The functionality is determined through an enzyme immunoassay. For a more detailed discussion regarding the obtained cells and their gene and protein expression, refer to Van Haute et al. (Generation of lung epithelial-like tissue from human embryonic stem cells Respiratory Research10: 105, 2009).