Ciliogenesis in submersion and suspension cultures of human nasal epithelial cells

Human nasal respiratory cells lose cilia in submerged cultures. This study compares the effect of extracellular matrix (ECM) molecules of the basal lamina on ciliogenesis in submerged cell cultures to ECM-free suspension cultures. Respiratory mucosa of nasal turbinates was the routine source for the cultures of nasal epithelial cells. For the submersion cultures, enzymatically isolated cells were seeded either on a layer of lethally irradiated ((60)Co, 60 Gy) murine 3T3-feeder fibroblasts or on an ECM-coated culture flask. For suspension cultures, the flasks were rotated for 3 days after cell seeding. In ECM-coated flasks, epithelial cell attachment and confluence was promoted and always much better than in cultures on a feeder layer. Respiratory cells lost cilia during the first 5 weeks in submerged cultures. Genesis of new, actively beating cilia was seen after 5-6 weeks when plastic culture dishes were coated with ECM molecules. Cells grown on uncoated plastic dishes together with 3T3-fibroblasts showed no ciliogenesis. Spheroids of epithelial cells in suspension cultures lost cilia during the 1st week and developed new cilia after 1-2 weeks in vitro. Our results suggest that ECM molecules are not the only signal for ciliary differentiation of respiratory cells in vitro, because suspension cultures are ECM free. However, the presence of ECM molecules in submerged cell cultures promotes the attachment and early confluence of seeded epithelial cells with a high density of cuboidal epithelial cells. The specific cellular shape and intense intercellular contact of these cuboidal cells may be among the most important signals inducing terminal differentiation and ciliogenesis.