In Vitro Cellular & Developmental Biology

, Volume 24, Issue 5, pp 420–428 | Cite as

A biphasic chamber system for maintaining polarity of differentiation of culture respiratory tract epithelial cells

  • Michael J. Whitcutt
  • Kenneth B. Adler
  • Reen Wu
Regular Papers


A simple, disposable, biphasic cultivation chamber has been developed for respiratory tract epithelial cells. This chamber, the Whicutt chamber, contains a movable, transparent, permeable gelatin membrane that can be employed either submerged in the culture medium, thereby feeding the cells by the traditional immersion method, or raised to the surface of the culture medium, to bring the apical surfaces of the cells into contact with air and provide nutrients only from below (basal feeding). The effects of biphasic cultivation on the growth and differentiation of respiratory tract epithelial cells from different sources have been studied in Whitcutt chambers. Primary hamster tracheal epithelial (HTE) cells grown to confluence with basal feeding developed a ciliated columnar morphology, with differentiated features (cilia and mucous granules) located in the apical region of the epithelial layer. These cells secreted mucinlike molecules from the apical surface (i.e. the surface in contact with air). Although the apical localization of differentiation features was greater, mucous cell differentiation achieved by basal feeding was quantitatively not greater than that achieved by continuous immersion feeding. Similarly, basal feeding did not alter the degree of epithelial cell differentiation in cultures derived from rat, rabbit, and monkey tracheas or from human bronchial and nasal tissues. In contrast, the differentiation of guinea pig tracheal epithelial cells in culture was significantly influenced by the feeding method employed. When fed basally, guinea pig tracheal epithelial cell cultures expressed various mucociliary functions with resemblance to mucociliary layers in vivo, whereas constantly immersed cultures seemed stratified and squamous. These results suggest that, at least for guinea pigs, the combination of feeding methods provided by the Whitcutt chamber can be used to achieve differentiated cultures of tracheal epithelial cells with a polarity of differentiation that is similar to that observed in intact airways in vivo.

Key words

airways differentiation biphasic cultivation polarity mucous cells epithelial cells 


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  1. 1.
    Chambard, M.; Garbrion, J.; Mauchamp, J. Influence of collagen gel on the orientation of epithelial cell polarity: follicle formation from isolated thyroid cells and from performed monolayers. J. Cell. Biol. 91: 157–166; 1981.PubMedCrossRefGoogle Scholar
  2. 2.
    Chlapowksi, F. J.; Haynes, L. Growth and differentiation of transitional epithelium in vitro. J. Cell. Biol. 83: 605–614; 1979.CrossRefGoogle Scholar
  3. 3.
    Emerman, J. T.; Burwen, S. J.; Pitelka, D. R. Substrate properties influencing ultrastructural differentiation of mammary epithelial cells in culture. Tissue Cell 11: 109–119; 1979.PubMedCrossRefGoogle Scholar
  4. 4.
    Emerman, J. T.; Pitelka, D. R. Maintenance and induction of morphological differentiation in dissociated mammary epithelium on floating collagen membranes. In Vitro 13: 316–328; 1977.PubMedCrossRefGoogle Scholar
  5. 5.
    Kleinman, H. K.; Klebe, R. J.; Martin, G. R. Role of collagenous matrices in the adhesion and growth of cells. J. Cell. Biol. 88: 473–485; 1981.PubMedCrossRefGoogle Scholar
  6. 6.
    Laemmli, U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680–685; 1970.PubMedCrossRefGoogle Scholar
  7. 7.
    Lee, E. Y. P.; Lee, W. H.; Kaetzel, C. S., et al. Interaction of mouse mammary epithelial cells with collagen substrata regulation of casein gene expression and secretion. Proc. Natl. Acad. Sci. USA 82: 1419–1423; 1985.PubMedCrossRefGoogle Scholar
  8. 8.
    Lillie, J. H.; MacCallum, D. K.; Jepsen, A. Fine structure of subcultivated stratified squamous epithelium growth on collagen rafts. Exp. Cell. Res. 125: 153–165; 1980.PubMedCrossRefGoogle Scholar
  9. 9.
    Maciag, T. S.; Cerudola, S.; Ilsley, P., et al. An endothelial cell growth factor from bovine hypothalamus: identification and partial characterization. Proc. Natl. Acad. Sci. USA 76: 5674–5678; 1979.PubMedCrossRefGoogle Scholar
  10. 10.
    Michalopoulos, G.; Pitot, H. C. Primary cultures of parenchymal liver cells on collagen membranes. Exp. Cell. Res. 94: 70–78; 1975.PubMedCrossRefGoogle Scholar
  11. 11.
    Wu, R. In Vitro differentiation of airway epithelial cells. In: Schiff, L. J., ed. In Vitro models of respiratory epithelium. Boca Raton, FL: CRC Press, Inc.; 1986: 1–26.Google Scholar
  12. 12.
    Wu, R.; Cheng, E.; Yankaskas, J., et al. Growth and differentiation of human nasal epithelial cells in culture: serumfree hormone-supplemented medium and proteoglycan synthesis. Am. Rev. Respir. Dis. 132: 311–320; 1985.PubMedGoogle Scholar
  13. 13.
    Wu, R.; Nolan, E.; Turner, C. Expression of tracheal differentiated functions in serum-free hormone-supplemented medium. J. Cell. Physiol 125: 167–181; 1985.PubMedCrossRefGoogle Scholar
  14. 14.
    Wu, R.; Smith, D. Continuous multiplication of rabbit tracheal epithelial cells in a defined hormone-supplemented medium. In Vitro 18: 800–812; 1982.PubMedGoogle Scholar
  15. 15.
    Wu, R.; Wu, M. M. J. Effects of retinoids on human bronchial epithelial cells: differential regulation of hyaluronate synthesis and keratin protein synthesis. J. Cell. Physiol. 127: 73–82; 1986.PubMedCrossRefGoogle Scholar
  16. 16.
    Yang, J.; Nandi, S. Grwoth of cultured cells using collagen as substrate. Int. Rev. Cytol. 81: 249–286; 1983.PubMedCrossRefGoogle Scholar
  17. 17.
    Jeffery, P. K.; Reid, L. M. Ultrastructure of airway epithelium and submucosal gland during development. In: Hodson, W. A., ed. Development of the lung. New York: Marcel Dekker; 1977: 87–134.Google Scholar

Copyright information

© Tissue Culture Association, Inc. 1988

Authors and Affiliations

  • Michael J. Whitcutt
    • 1
  • Kenneth B. Adler
    • 2
  • Reen Wu
    • 3
  1. 1.W. Alton Jones Cell Science CenterLake Placid
  2. 2.Department of pathology, College of MedicineUniversity of VermontBurlington
  3. 3.Department of Anatomy and California Primate Research Center, School of Veterinary MedicineUniversity of CaliforniaDavis

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