Abstract
BACKGROUND: Although pharmacological treatment has increased the average life expectancy of patients with cystic fibrosis, the median survival of females is shorter than that of males. In vitro and in vivo studies have shown that estrogens play a relevant role in the disease progression. The aim of this study was to investigate the effects of 17β-estradiol and tamoxifen citrate (TMX) on calcium-activated chloride channel (CaCC) currents in human bronchial epithelial cells carrying the ΔPhe508-CFTR mutation both in homozygosis and in heterozygosis. METHODS: Perforated patch clamp experiments were performed on single cells of the immortalized cell lines CFBE and IB3-1. Gramicidin (10 or 20 μM) was added to the electrode solution to reach the whole cell configuration. The electrical stimulation protocol consisted of square voltages ranging from - 80 to + 80 mV, in steps of 20 mV and with a duration of 800 msec. RESULTS: The presence of 17β-estradiol significantly reduced the CaCC currents, both in basal conditions and in the presence of ATP (100 μM). The addition of TMX (10 μM) completely restored the currents abolished by 17β-estradiol, in basal conditions and after stimulation with ATP in both CFBE and IB3-1 cells. TMX had a strong, direct action on membrane current density, which significantly increased more than 4-fold in both cases. The membrane current stimulation produced by TMX was further enhanced by the addition of ATP. CFBE cells incubated for 24 h with 3 μM VX-809 (a CFTR corrector) and then acutely stimulated with VX-770 (a CFTR potentiator) in the presence of forskolin, showed an increase of chloride currents which were abolished by Inh-172. The chloride current density induced by TMX + ATP was, on average, greater than that obtained with VX-809 + VX-770 + forskolin. The currents elicited by TMX + ATP were abolished by the addition of NPPB, a CaCC inhibitor. The combined administration of TMX/ATP and VXs/FSK had an additional effect on chloride currents. CONCLUSIONS: Our results show that TMX restores CaCC currents inhibited by 17ß-estradiol and directly activates the transmembrane chloride currents potentiated by ATP, an effect which is mutation independent. The combined effect of TMX with current used treatments for cystic fibrosis could be of benefit to patients.