Differentiated structure and function of cultures from human tracheal epithelium.
Here we describe the conditions which allow cultured human tracheal epithelial cells to retain the ion transport properties and ultrastructure of the original tissue. The order of potency of growth supports and media additives in elevating baseline short-circuit current (Isc) and responses to mediators were vitrogen gel (VIT) greater than extracellular matrix from bovine corneal endothelial cells (ECM) greater than human placental collagen (HPC), and 2% Ultroser G serum substitute (USG) greater than 5% fetal calf serum (FCS) greater than defined growth factors (GF). For all combinations of medium and growth supports, an air interface (AIR) gave better electrical properties than immersion feeding (IMM). As opposed to our earlier conditions (HPC/FCS/IMM), the best new combination (VIT/USG/AIR) produced higher baseline Isc (58.0 +/- 10.6 vs. 5.1 +/- 1.0 microA/cm2) and increased Isc responses to isoproterenol (6.1 +/- 1.5 vs. 0.8 +/- 0.3 microA/cm2) and bradykinin (9.6 +/- 2.0 vs. 1.0 +/- 0.2 microA/cm2), while retaining high transepithelial resistance (227 +/- 5 omega.cm2). VIT/USG/AIR led to the appearance of cilia, an increase in the depth of the cell sheets (50 vs. 10 microns), longer and more frequent apical microvilli, and increased interdigitations of the basolateral membrane. Protein and DNA content were also significantly increased. Secretory granules were present which stained with antibody to goblet cells, but not to serous or mucous gland cells. CF cells grown in VIT/USG/AIR showed high baseline Isc (69 +/- 18 microA/cm2) and a proportionately larger inhibition of Isc by amiloride (70 +/- 10 vs. 34 +/- 3%). Isc did not respond to isoproterenol, and the response to bradykinin was 22% normal.
Yamaya
M
M
Cystic Fibrosis Research Center, University of California, San Francisco 94143.
Finkbeiner
W E
WE
Chun
S Y
SY
Widdicombe
J H
JH
D000584:Amiloride / Q000494:pharmacology; D001920:Bradykinin / Q000494:pharmacology; D002034:Bumetanide / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D002478:Cells, Cultured; D002923:Cilia / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D003594:Cytoplasmic Granules / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D004594:Electrophysiology / Q000379:methods; D004848:Epithelium / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D007700:Kinetics; D008564:Membrane Potentials; D008854:Microscopy, Electron / Q000379:methods; D008871:Microvilli / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D009092:Mucous Membrane / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D010042:Ouabain / Q000494:pharmacology; D014132:Trachea / Q000166:cytology / Q000502:physiology*; D062367:ortho-Aminobenzoates / Q000494:pharmacology
D062367:ortho-Aminobenzoates; D002034:Bumetanide; D010042:Ouabain; D000584:Amiloride; C001209:fenamic acid; D007545:Isoproterenol; D001920:Bradykinin