Long term culture of nasal epithelial cells

Abstract

Cystic fibrosis (CF) basic research has been hampered by the lack of a suitable animal model and is therefore dependent on the availability of human tissue. To date, the basic defect remains unknown, however, over the last decade convincing data have been produced that link the defect to altered epithelial electrolyte transport. Moreover, recent results of DNA analysis could confirm that the putative CF gene product is indeed expressed in epithelial tissues. Unfortunately, epithelial cells are difficult to maintain in culture, especially transport-active epithelia tend to be short-lived. We have devised a simple method for the subculturing of nasal epithelium which has been shown to exhibit altered electrolyte transport in CF patients. Nasal epithelial cells were isolated from nasal polyps obtained at surgery by protease digestion. Subsequently, cells were plated in culture medium and irradiated mouse fibroblasts were added as a feeder layer after 48 h. After 5-15 d, epithelial cells could be subcultured without loosing their growth potential, and were continuously maintained in culture for up to 4 months. The cells were frozen in liquid nitrogen following standard methods. Growth could be reinstituted after thawing the epithelium. Subcultured and thawed cells were clearly characterized as of epithelial origin and distinguished from mesodermal cells by their reaction with antibodies against keratin, vimentin, and desmin.