The pathogenesis of cystic fibrosis (CF) airways infection is unknown. Two hypotheses, "hypotonic [low salt]/defensin" and "isotonic volume transport/mucus clearance," attempt to link defects in cystic fibrosis transmembrane conductance regulator-mediated ion transport to CF airways disease. We tested these hypotheses with planar and cylindrical culture models and found no evidence that the liquids lining airway surfaces were hypotonic or that salt concentrations differed between CF and normal cultures. In contrast, CF airway epithelia exhibited abnormally high rates of airway surface liquid absorption, which depleted the periciliary liquid layer and abolished mucus transport. The failure to clear thickened mucus from airway surfaces likely initiates CF airways infection. These data indicate that therapy for CF lung disease should not be directed at modulation of ionic composition, but rather at restoring volume (salt and water) on airway surfaces.

The amiloride-sensitive epithelial sodium channel, ENaC, is a heteromultimeric protein made up of three homologous subunits (alpha, beta and gamma) (1,2). In vitro, assembly and expression of functional active sodium channels in the Xenopus oocyte is strictly dependent on alpha-ENaC--the beta and gamma subunits by themselves are unable to induce an amiloride-sensitive sodium current in this heterologous expression system (2). In vivo, ENaC constitutes the limiting step for sodium absorption in epithelial cells that line the distal renal tubule, distal colon and the duct of several exocrine glands. The adult lung expresses alpha, beta and gamma ENaC (3,4), and an amiloride-sensitive electrogenic sodium reabsorption has been documented in upper and lower airways (3-7), but it is not established whether this sodium transport is mediated by ENaC in vivo. We inactivated the mouse alpha-ENaC gene by gene targeting. Amiloride-sensitive electrogenic Na+ transport was abolished in airway epithelia from alpha-ENaC(-/-) mice. Alpha-ENaC(-/-) neonates developed respiratory distress and died within 40 h of birth from failure to clear their lungs of liquid. This study shows that ENaC plays a critical role in the adaptation of the newborn lung to air breathing.

Cystic fibrosis (CF) transmembrane conductance regulator (CFTR)-dependent airway epithelial bicarbonate transport is hypothesized to participate in airway surface liquid pH regulation and contribute to lung defense. We measured pH and ionic composition in apical surface liquid (ASL) on polarized normal (NL) and CF primary bronchial epithelial cell cultures under basal conditions, after cAMP stimulation, and after challenge with luminal acid loads. Under basal conditions, CF epithelia acidified ASL more rapidly than NL epithelia. Two ASL pH regulatory paths that contributed to basal pH were identified in the apical membrane of airway epithelia, and their activities were measured. We detected a ouabain-sensitive (nongastric) H ؉ ,K ؉ -ATPase that acidified ASL, but its activity was not different in NL and CF cultures. We also detected the following evidence for a CFTR-dependent HCO 3 ؊ secretory pathway that was defective in CF: (i) ASL [HCO 3 ؊ ] was higher in NL than CF ASL; (ii) activating CFTR with forskolin͞3-isobutyl-1-methylxanthine alkalinized NL ASL but acidified CF ASL; and (iii) NL airway epithelia more rapidly and effectively alkalinized ASL in response to a luminal acid challenge than CF epithelia. We conclude that cultured human CF bronchial epithelial pH ASL is abnormally regulated under basal conditions because of absent CFTRdependent HCO 3 ؊ secretion and that this defect can lead to an impaired capacity to respond to airway conditions associated with acidification of ASL.

To investigate respiratory epithelial function in cystic fibrosis, we measured the transepithelial electrical potential difference across the upper and lower respiratory mucosa in patients with cystic fibrosis and control subjects. The nasal potential difference in the 24 patients with cystic fibrosis exceeded by more than 3 standard deviations the mean voltage in healthy controls, subjects with other diseases, and subjects heterozygous for cystic fibrosis. Potential differences in lower airways were measured in four patients and were significantly greater than in controls (P less than 0.05). Superfusion of the luminal surface with amiloride, an inhibitor of active sodium absorption, induced greater reductions in both nasal and airway potential difference in patients than in controls. We conclude that the increased respiratory-epithelial potential differences appear to be a specific abnormality in homozygotes for cystic fibrosis. The greater reduction in potential difference in response to amiloride suggests that absorption of excess salt and perhaps liquid from respiratory epithelial surfaces contributes to the pathogenesis of lung disease in cystic fibrosis.

The transepithelial potential difference (PD) of cystic fibrosis (CF) airway epithelium is abnormally raised and the Cl-permeability is low. We studied the contribution of active Na' absorption to the PD and attempted to increase the Cl-permeability of CF epithelia. Nasal epithelia from CF and control subjects were mounted in Ussing chambers and were short-circuited. The basal rate of Na' absorption was raised in CF polyps compared with control tissues. Whereas beta agonists induced Cl-secretion in normal and atopic epithelia, beta agonists further increased the rate of Na' absorption in CF epithelia without inducing Clsecretion. This unusual effect is not due to an abnormal CF beta receptor because similar effects were induced by forskolin, and because cAMP production was similar in normal and CF epithelia. We conclude that CF airway epithelia absorb Na' at an accelerated rate. The abnormal response to beta agonists may reflect a primary abnormality in a cAMP-modulated path, or a normal cAMP-modulated process in a Cl-impermeable epithelial cell.