Published ahead of print on September 22, 2005, doi:10.1165/rcmb.2005-0161OC Am. J. Respir. Cell Mol. Biol., Volume 34, Number 1, January 2006, 39-48 A more recent version of this article appeared on January 1, 2006
Submitted on May 2, 2005 Long-term Cultures of Polarized Airway Epithelial Cells from CF PatientsLudovic Wiszniewski1,1 Department of Pediatrics, Laboratory of Clinical Investigation III, Geneva University Hospitals, Geneva, Switzerland, 2 Division of Pulmonary Medicine, Geneva University Hospitals, Geneva, Switzerland, 3 Department of Pathology, Geneva Medical School, Geneva, Switzerland, 4 Clinic of Oto-Rhino-Laryngology, Geneva University Hospitals, Geneva, Switzerland * To whom correspondence should be addressed. E-mail: Marc.Chanson{at}hcuge.ch.
The poor ability of respiratory epithelial cells to proliferate and differentiate in vitro into a pseudostratified mucociliated epithelium limits the general use of primary airway epithelial cell (AEC) cultures generated from patients with rare diseases such as cystic fibrosis (CF). Here, we described a procedure to amplify AEC isolated from nasal polyps and generate long-term cultures of the respiratory epithelium. AEC were seeded onto microporous permeable supports that carried on their undersurface a preformed feeder layer of primary human airway fibroblasts. As compared to previously described methods, the use of fibroblast feeder layers strongly stimulated the proliferation of epithelial cells, allowing the expansion of the cell pool with successive passages. AEC at increasing passage were seeded onto supports undercoated with airway fibroblasts and exposed to air. Either freshly isolated or amplified AEC could differentiate into a pseudostratified mucociliated epithelium for at least 10 months. Thus, CF epithelia cultures showed elevated Na+ transport, drastic hyper-absorption of surface liquid and absence of cAMP-induced Cl- secretion as compared to non-CF cultures. They were also characterized by thick apical secretion that hampered the movement of cell surface debris by cilia. However, CF respiratory epithelia did not show increased production of mucins or IL-8. The method described here is now routinely used in our laboratory to establish long-term cultures of well-differentiated respiratory epithelia from human airway biopsies.
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