Correction of cAMP-stimulated fluid secretion in cystic fibrosis airway epithelia: efficiency of adenovirus-mediated gene transfer in vitro.
Adenovirus vectors are a promising vehicle to deliver cystic fibrosis transmembrane conductance regulator (CFTR) cDNA to airway epithelia. However, the value of adenovirus vectors will depend on the efficiency with which the vector can correct the defective fluid transport that is though to underlie the pathogenesis of the disease. To address the efficiency of gene transfer, we applied adenovirus vectors expressing CFTR (Ad2/CFTR-1) or beta-galactosidase to the mucosal surface of primary cultures of airway epithelial cells grown as polarized epithelial monolayers on permeable filter supports. These conditions provide a model that reproduces the physiology of the airways in vivo. We found that after adding 1 moi Ad2/CFTR-1 to the mucosal surface, cAMP agonists stimulated fluid secretion that was within the range observed in epithelia from normal subjects. When we measured electrolyte transport, we found that as little as 0.1 moi partially restored cAMP-stimulated Cl- secretion, and at 10 moi Cl- secretion was in the normal range. A related vector encoding beta-galactosidase generated activity in approximately 20% of cells at an moi of 1 and 90% of cells at an moi of 10. These data suggest that Ad2/CFTR-1 is very efficient at restoring normal fluid and electrolyte transport to CF airway epithelia. Thus, they suggest that relatively low input doses could be used for gene transfer to CF airway epithelia.
Zabner
J
J
Howard Hughes Medical Institute, University of Iowa College of Medicine, Iowa City 52242.
Couture
L A
LA
Smith
A E
AE
Welsh
M J
MJ
D000256:Adenoviridae / Q000235:genetics*; D002712:Chlorides / Q000378:metabolism*; D000242:Cyclic AMP / Q000502:physiology*; D003550:Cystic Fibrosis / Q000235:genetics / Q000378:metabolism* / Q000628:therapy; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D004848:Epithelium / Q000378:metabolism; D018014:Gene Transfer Techniques; D005822:Genetic Vectors; D006801:Humans; D008168:Lung / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D001616:beta-Galactosidase / Q000378:metabolism
C505032:CFTR protein, human; D002712:Chlorides; D008565:Membrane Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000242:Cyclic AMP; D001616:beta-Galactosidase