Influence of apical fluid volume on the development of functional intercellular junctions in the human epithelial cell line 16HBE14o-: implications for the use of this cell line as an in vitro model for bronchial drug absorption studies.
Air-interfaced culture (AIC) versus liquid-covered culture (LCC) conditions are known to have different effects on the differentiated phenotype of several cell types, including lung epithelial cells. We report the influence of culture conditions such as apical medium volume on the development of intercellular junctions in the human epithelial cell line 16HBE14o-. Immunofluorescence staining of the tight-junctional protein, ZO-1, has revealed its presence in cells grown in both AIC and LCC. However, only LCC-grown cells exhibit protein ZO-1 localized as a zonula-occludens-like regular belt connecting neighboring cells. The presence of typical tight junctions has been confirmed by electron microscopy. Immunostaining for occludin, claudin-1, connexin43, and E-cadherin has demonstrated intercellular junction structures only in the cells in LCC. These morphological findings have been paralleled by higher transepithelial electrical resistance values and similar fluxes of the hydrophilic permeability marker, fluorescein-Na, under LCC compared with AIC conditions. We conclude that the formation of functional 16HBE14o- cell layers requires the presence of an apical fluid volume, in contrast to other culture conditions for airway epithelial cells.
Ehrhardt
Carsten
C
Department of Biopharmaceutics and Pharmaceutical Technology, Bldg. 8.1, Saarland University, 66123 Saarbrücken, Germany.
Kneuer
Carsten
C
Fiegel
Jennifer
J
Hanes
Justin
J
Schaefer
Ulrich Friedrich
UF
Kim
Kwang-Jin
KJ
Lehr
Claus-Michael
CM
D000199:Actins / Q000032:analysis; D022005:Adherens Junctions / Q000737:chemistry / Q000378:metabolism; D001980:Bronchi / Q000166:cytology*; D015820:Cadherins / Q000032:analysis; D002460:Cell Line; D016764:Cell Polarity / Q000502:physiology; D062445:Claudin-1; D018031:Connexin 43 / Q000032:analysis; D004553:Electric Conductivity; D004847:Epithelial Cells / Q000378:metabolism* / Q000648:ultrastructure; D017629:Gap Junctions / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D007223:Infant; D008565:Membrane Proteins / Q000032:analysis; D062793:Occludin; D010599:Pharmacokinetics; D020545:Respiratory Mucosa / Q000166:cytology* / Q000378:metabolism*; D019108:Tight Junctions / Q000737:chemistry / Q000378:metabolism*
D000199:Actins; C568108:CLDN1 protein, human; D015820:Cadherins; D062445:Claudin-1; D018031:Connexin 43; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin