Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) that impair its expression and/or chloride channel function. Here, we provide evidence that type 4 cyclic nucleotide phosphodiesterases (PDE4s) are critical regulators of the cAMP/PKA-dependent activation of CFTR in primary human bronchial epithelial cells. In non-CF cells, PDE4 inhibition increased CFTR activity under basal conditions (ΔISC 7.1 μA/cm(2)) and after isoproterenol stimulation (increased ΔISC from 13.9 to 21.0 μA/cm(2)) and slowed the return of stimulated CFTR activity to basal levels by >3-fold. In cells homozygous for ΔF508-CFTR, the most common mutation found in CF, PDE4 inhibition alone produced minimal channel activation. However, PDE4 inhibition strongly amplified the effects of CFTR correctors, drugs that increase expression and membrane localization of CFTR, and/or CFTR potentiators, drugs that increase channel gating, to reach ∼ 25% of the chloride conductance observed in non-CF cells. Biochemical studies indicate that PDE4s are anchored to CFTR and mediate a local regulation of channel function. Taken together, our results implicate PDE4 as an important determinant of CFTR activity in airway epithelia, and support the use of PDE4 inhibitors to potentiate the therapeutic benefits of CFTR correctors and potentiators.

Ϫ and mucin secretion in cultures of non-CF and CF human tracheobronchial gland mucous (HTGM and CFTGM, respectively) cells. Biochemical studies showed that HTMG cells secreted typical airway mucins, and immunohistochemical studies showed that these cells expressed MUC1, MUC4, MUC5B, MUC8, MUC13, MUC16, and MUC20. Effects of cumulative doses of methacholine (MCh), phenylephrine (Phe), isoproterenol (Iso), and ATP on mucin and Cl Ϫ secretion were studied on HTGM and CFTGM cultures. Baseline mucin secretion was not significantly altered in CFTGM cells, and the increases in mucin secretion induced by mediators were unaltered (Iso, Phe) or slightly decreased (MCh, ATP). Across mediators, there was no correlation between the maximal increases in Cl Ϫ secretion and mucin secretion. In HTGM cells, the Cl Ϫ channel blocker, diphenylamine-2-carboxylic acid, greatly inhibited Cl Ϫ secretion but did not alter mucin release. In HTGM cells, mediators (10 Ϫ5 M) increased mucin secretion in the rank order ATP Ͼ Phe ϭ Iso Ͼ MCh. They increased Cl Ϫ secretion in the sequence ATP Ͼ MCh Ϸ Iso Ͼ Phe. The responses in Cl Ϫ secretion to MCh, ATP, and Phe were unaltered by CF, but the response to Iso was greatly reduced. We conclude that mucin secretion by cultures of human tracheobronchial gland cells is independent of Cl Ϫ secretion, at baseline, and is unaltered in CF; that the ratio of Cl Ϫ secretion to mucus secretion varies markedly depending on mediator; and that secretions induced by stimulation of ␤-adrenergic receptors will be abnormally concentrated in CF.

Cystic fibrosis (CF) is caused by mutations in the gene encoding the cystic fibrosis transmembrane conductance regulator (CFTR) that impair its expression or function. PKA phosphorylation is the main mechanism to activate CFTR. However, previous reports indicated that cAMP/PKA activators are ineffective in restoring CFTR conductance in CF epithelia. Our current findings challenge this conclusion. We show that inhibition of type 4 cAMP‐phosphodiesterases (PDE4s) increased CFTR activity in non‐CF primary human bronchial epithelial cells under basal conditions and after β‐adrenergic stimulation, and slowed the return of stimulated CFTR activity to basal levels. In cells homozygous for ∆F508‐CFTR, the most common mutation found in CF patients, PDE4 inhibition alone produced minimal channel activation. However, PDE4 inhibition strongly amplified the effects of CFTR correctors, drugs that increase expression and membrane‐localization of CFTR, and CFTR potentiators, drugs that increase channel gating to reach ~25% of the chloride conductance observed in non‐CF cells. Biochemical studies indicate that PDE4s are anchored to CFTR and mediate local regulation of channel function. Taken together, our results implicate PDE4 as an important determinant of CFTR activity in airway epithelia and support the use of PDE4 inhibitors to potentiate the therapeutic benefits of CFTR correctors and potentiators. Grant Funding Source: Supported by the Cystic Fibrosis Foundation, the NIH, and Cystic Fibrosis Research Inc.

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