Growth and differentiation of human distal airway epithelial cells in culture. Effects of small amounts of serum in defined medium.

We herein describe a new simple method for culturing primary epithelial cells derived from the distal airway of adult human lung. Peripheral lung tissue obtained from surgical materials was cultured as explants in Ham's F12 medium supplemented with hormones and growth factors. From days 10 to 14, outgrowth of epithelial cells started on the dish surface, and even after removal of explants at days 14 to 16, these cells continued to replicate during the 3rd and 4th week of culture, and eventually formed epithelial cell foci (10 to 20-fold increase in population). They then ceased to replicate and terminally differentiated in the 5th week. Addition of 1% serum to the culture medium enhanced the initial outgrowth of epithelial cells, whereas small amounts of serum had no effect on proliferation of cells after explant removal. On the other hand, serum modulated the differentiation phenotypes of epithelial cells. In the presence of 1% serum, numerous ciliated and secretory cells appeared, whereas the cells underwent epidermoid differentiation in the absence of serum. When replated onto 3T3 fibroblast feeder layers, the epithelial cells from earlier cultures showed a great replicative capability and formed colonies at higher frequencies (colony-forming efficiency, 8 to 27% at days 14 to 21), but the replicative capability was significantly reduced at the confluent stage (colony-forming efficiency, 0.44 to 2.0% at day 28). Morphologic examinations of explants strongly suggested that the primary cells were derived from the bronchiolar epithelium. We conclude that this new culture system provides an excellent model for studying the growth, differentiation, and function of human bronchiolar epithelial cells in vitro.