Activation of chloride transport in CF airway epithelial cell lines and primary CF nasal epithelial cells by S-nitrosoglutathione

BACKGROUND: It has been suggested that low μM concentrations of S-nitrosoglutathione (GSNO), an endogenous bronchodilator, may promote maturation of the defective cystic fibrosis (CF) transmembrane conductance regulator (CFTR). Because nitric oxide (NO) and GSNO levels appear to be low in the CF airway, there is an interest in the possibility that GSNO replacement could be of therapeutic benefit in CF. METHODS: The effect of GSNO on chloride (Cl(-)) transport was investigated in primary nasal epithelial cells obtained from CF patients homozygous for the delF508 mutation, as well as in two CF cell lines (CFBE and CFSME), using both a fluorescent Cl(- )indicator and X-ray microanalysis. Maturation of delF508 CFTR was determined by immunoblotting. RESULTS: Treatment with 60 μM GSNO for 4 hours increased cAMP-induced chloride efflux in nasal epithelial cells from 18 out of 21 CF patients, but did not significantly affect Cl(- )efflux in cells from healthy controls. This Cl(- )efflux was confirmed by measurements with a fluorescent Cl(- )indicator in the CFBE and CFSME cell lines. The effect of GSNO on Cl(- )efflux in CFBE cells could be inhibited both by a specific thiazolidinone CFTR inhibitor (CFTR(inh)-172) and by 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulfonic acid (H(2)DIDS). X-ray microanalysis showed that, following 4 hours incubation with 60 μM GSNO, cAMP agonists caused a decrease in the cellular Cl(- )concentration in CFBE cells, corresponding to Cl(- )efflux. GSNO exposure resulted in an increase in the protein expression and maturation, as shown by immunoblot analysis. GSNO did not increase the cytosolic Ca(2+ )concentration in cultured airway epithelial cells. CONCLUSION: Previous studies have suggested that treatment with GSNO promotes maturation of delF508-CFTR, consistent with our results in this study. Here we show that GSNO increases chloride efflux, both in the two CF cell lines and in primary nasal epithelial cells from delF508-CF patients. This effect is at least in part mediated by CFTR. GSNO may be a candidate for pharmacological treatment of the defective chloride transport in CF epithelial cells.