CHARACTERIZATION OF NA-H+ ANTIPORTER ACTIVITY ASSOCIATED WITH HUMAN CHEEK EPITHELIAL-CELLS()

Na+ transport activity was characterized in human cheek epithelial cel ls obtained from normotensive adult subjects. The cells were isolated using a mouth-wash procedure and assayed for Na+ uptake using a radioa ctive (Na-22(+)) rapid filtration assay. Cheek cells displayed proton- dependent Na+ uptake activity that was dependent on the magnitude of t he externally directed proton gradient measured using the fluorescent probe 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein to determine int racellular pH. Amiloride, ethylisopropylamiloride (EIPA), 5-(N,N-dimet hyl)-amiloride, 5-(N-methyl-N-isobutyl)-amiloride (MIA), and 5-(N,N-he xamethylene)-amiloride (NNHA) all inhibited proton-dependent Na+ uptak e, with MIA, EIPA, and NNHA being the most potent. The Michaelis const ant (K-m) for extracellular Na+ was 5.7 mM, while the maximum velocity for Na+-H+ antiporter activity was 4.3 nmol Na+.mg protein(-)1.30 s(- 1). The K-m for intracellular H+ was 0.17 mu M, with a Hill coefficien t of 0.7. Stimulation by ouabain and inhibition by bumetanide of cheek cell proton-dependent Na+ uptake indicated only relatively low activi ties of Na+-K+-ATPase and Na+-K+-2Cl(-) cotransport, respectively. The se results are consistent with the presence of Na+-H+ antiporter activ ity in cheek cells. Cheek cells therefore provide a convenient, relati vely noninvasive source of tissue for examining Na+-H+ antiporter acti vity in human subjects.