In vitro culture of microdissected rat nasal airway tissues

The surface epithelium lining the nasal airways is a potential target For i nhaled contaminants such as ozone, endotoxin, formaldehyde, tobacco smoke a nd organic dusts. The epithelial response to injury may depend on the toxic ant, the type of epithelium, the severity of the injury, and the presence o f inflammatory cells and their secreted products. To study mechanisms of to xicant-induced epithelial injury and repair, in the absence of cellular inf lammation or other systemic effects, we have developed a culture system to maintain morphologically distinct nasal airway epithelium in vitro. Microdi ssected maxilloturbinates and proximal nasal septa of male F344/N rats were cultured at an air-liquid interface for up to 14 d in supplemented serum-f ree medium. Maxilloturbinates are lined by nonciliated cuboidal nasal trans itional epithelium (NTE) with few or no mucous cells. The proximal nasal se ptum is lined by a mucociliary respiratory epithelium (RE) that normally co ntains numerous mucous cells. Preservation of the normal RE and NTE phenoty pe in culture was assessed by light and electron microscopy, and analysis o f an airway mucin gene (rMuc-5AC) messenger RNA (mRNA), Both RE and NTE ret ained normal cell morphology for 14 d in culture (DIC), After 14 DIC there were 20% fewer RE cells in the septa (equal loss of ciliated and mucous cel ls) and 25% more NTE cells in the maxilloturbinates (increased number of ba sal cells). Compared with the RE, the NTE expressed consistently low levels of rMuc-5AC mRNA and had little to no histochemically detectable intraepit helial mucosubstances (IM) after 0, 3, 7, or 14 DIG. The amount of stored I M and the steady-state levels of rMuc-5AC mRNA in the RE decreased with tim e in culture. In summary, this culture system can maintain fully differenti ated secretory and nonsecretory rat airway epithelia in vitro for up to 14 d. This study was an essential first step in developing a system to study t he pathogenesis of toxicant-induced airway epithelial injury and mechanisms of cellular repair and adaptation in the absence of cellular inflammation and other systemic influences.