Identification of Na(+)-H+ exchange in human normal and cystic fibrotic ciliated airway epithelium.

Abstract

The pH-sensitive probe 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein was used to measure intracellular pH (pHi) and test for Na(+)-H+ exchange in single ciliated human nasal epithelial (HNE) cells from normal and cystic fibrosis (CF) subjects. In N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES)-buffered NaCl Ringer solution, average pHi for normal and CF ciliated HNE cells was 7.15 +/- 0.02 and 7.18 +/- 0.03, respectively. Utilizing the NH+4-loading technique to acid load cells, we found that pHi decreased from approximately 7.2 to approximately 6.6; subsequent alkalinization of normal and CF ciliated cells required Na+ and was independent of Cl- but was blocked by amiloride (500 microM). In the presence of extracellular Na+, initial rates (delta pHi/min) of recovery from an acid load in normal and CF cells were 0.09 +/- 0.03 and 0.12 +/- 0.05, respectively. Pretreatment of cells with phorbol 12-myristate 13-acetate (PMA, 100 nM) caused initial rates of recovery to increase over control values by approximately twofold in both cell preparations. Other studies showed that the native H+ conductance of the plasma membrane was negligible in both normal and CF cell preparations. The results provide evidence in support of an amiloride-sensitive PMA-activated Na(+)-H+ exchanger in ciliated HNE cells from normal and CF donors.