The ability of the flavonoids genistein, apigenin, kaempferol, and quercetin to activate cystic fibrosis transmembrane conductance regulator-mediated Cl currents in human airway epithelium was investigated. We used the patch-clamp technique on single Calu-3 cells, transepithelial measurements in Calu-3 monolayers, and in vivo measurements of nasal potential difference. All flavonoids stimulated Cl currents in transepithelial experiments dose dependently. Half-maximal stimulatory concentrations were kaempferol (5.5 ± 1.7 μM) ≤ apigenin (11.2 ± 2.1 μM) ≤ genistein (13.6 ± 3.5 μM) ≤ quercetin (22.1 ± 4.5 μM). Stimulation of monolayers with forskolin significantly increased their sensitivity to flavonoids: kaempferol (2.5 ± 0.7 μM) ≤ apigenin (3.4 ± 0.9 μM) ≤ quercetin (4.1 ± 0.7 μM) ≤ genistein (6.9 ± 2.2 μM). Forskolin pretreatment significantly reduced the Hill coefficient ( n H) for all flavonoids. Control monolayers showed n H = 2.00 ± 0.21 (all flavonoids combined), and forskolin-stimulated monolayers showed n H = 1.07 ± 0.07, which was not different among the flavonoids. These data imply that the activation kinetics and the binding site(s) for flavonoids were significantly altered by forskolin stimulation. In whole cell patch-clamp experiments, maximal flavonoid-stimulated currents (percentage of forskolin-stimulated currents) were apigenin (429 ± 86%) ≥ kaempferol (318 ± 45%) ≥ genistein (258 ± 20%) = quercetin (256 ± 26%). Stimulation of the currents was caused by an increase in channel open probability. No other Cl conductances contributed significantly to the flavonoid-activated Cl currents in Calu-3 cells. In vivo, flavonoids significantly stimulated nasal potential difference by, on average, 27.8% of isoproterenol responses.
Somatic cell hemoglobin modulates nitrogen oxide metabolism in the human airway epithelium
