AN IMPROVED METHOD TO OBTAIN HIGHLY DIFFERENTIATED MONOLAYERS OF HUMAN BRONCHIAL EPITHELIAL-CELLS

Citation
Ljv. Galietta et al., AN IMPROVED METHOD TO OBTAIN HIGHLY DIFFERENTIATED MONOLAYERS OF HUMAN BRONCHIAL EPITHELIAL-CELLS, In vitro cellular & developmental biology. Animal, 34(6), 1998, pp. 478-481
Citations number
15
Categorie Soggetti
Developmental Biology","Cell Biology
ISSN journal
10712690
Volume
34
Issue
6
Year of publication
1998
Pages
478 - 481
Database
ISI
SICI code
1071-2690(1998)34:6<478:AIMTOH>2.0.ZU;2-O
Abstract
Electrophysiological studies of human bronchial epithelial cells in vi tro are limited by the scarcity of biological material available for p rimary culture. To overcome this problem, vie set up a protocol in whi ch the cell number is first enlarged in LHC9/RPMI 1640 serum-free medi um for up to six passages, each passage giving a four- to eightfold am plification. The cells are then plated at high density on permeable su pports. Cell differentiation, monitored by measuring transepithelial p otential difference (PD) and electrical resistance (R), is induced wit h a medium containing serum and a cocktail of different supplements an d hormones. Maximal values of PD and R, obtained after 4-7 d of cultur e on permeable supports, are around - 50 mV and 3000-4000 Ohm/cm(2), r espectively. Ussing chamber experiments show that basal short-circuit current (I-sc) is partially inhibited by the epithelial Na+ channel bl ocker amiloride. Stimulation with a cAMP-elevating agent induces a I-s c increase that is inhibited by the cystic fibrosis transmembrane cond uctance regulator (CFTR) blocker glibenclamide. Our culture protocol p rovides a large number of differentiated bronchial epithelial cell mon olayers starling from a low amount of material. This characteristic is useful for in vitro studies of ion transport in airway epithelium.