An improved method to obtain highly differentiated monolayers of human bronchial epithelial cells

Luis J V Galietta, Sabina Lantero, Andrea Gazzolo, Oliviero Sacco, Luca Romand, Giovanni A. Rossi, Olga Zegarra-Moran

Research output: Contribution to journalArticlepeer-review

Abstract

Electrophysiological studies of human bronchial epithelial cells in vitro are limited by the scarcity of biological material available for primary culture. To overcome this problem, we set up a protocol in which the cell number is first enlarged in LHC9/RPMI 1640 serum-free medium for up to six passages, each passage giving a four- to eightfold amplification. The cells are then plated at high density on permeable supports. Cell differentiation, monitored by measuring transepithelial potential difference (PD) and electrical resistance (R), is induced with a medium containing serum and a cocktail of different supplements and hormones. Maximal values of PD and R, obtained after 4-7 d of culture on permeable supports, are around -50 mV and 3000-4000 Ω/cm2, respectively. Using chamber experiments show that basal short-circuit current (I(sc)) is partially inhibited by the epithelial Na+ channel blocker amiloride. Stimulation with a cAMP-elevating agent induces a I(sc) increase that is inhibited by the cystic fibrosis transmembrane conductance regulator (CFTR) blocker glibenclamide. Our culture protocol provides a large number of differentiated bronchial epithelial cell monolayers starting from a law amount of material. This characteristic is useful for in vitro studies of ion transport in airway epithelium.

Original languageEnglish
Pages (from-to)478-481
Number of pages4
JournalIn Vitro Cellular & Developmental Biology - Animal
Volume34
Issue number6
Publication statusPublished - 1998

Keywords

  • Bronchial epithelium
  • Cystic fibrosis
  • Ion transport
  • Short-circuit current
  • Transepithelial resistance

ASJC Scopus subject areas

  • Developmental Biology
  • Clinical Biochemistry
  • Cell Biology

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