Effect of diclofenac on CFTR Cl⁻ channel activity.

<p>(A) Effect of diclofenac on CFTR Cl⁻ channel activity stimulated by forskolin (20 µM), CPT-cAMP (100 µM), or genistein (20 µM) in T84 cells. In this experiment, CFTR Cl⁻ channel activity was determined using apical Cl⁻ current analysis of T84 cell monolayers. Amphotericin B (250 µg/ml) was added into basolateral solutions to permeabilize basolateral membrane of T84 cells. Diclofenac at the indicated concentrations was added to both apical and basolateral sides. Representative current tracings of 4–7 experiments were shown. (B) Effect of diclofenac on CFTR Cl⁻ channel activity in human airway epithelial (Calu-3) cells. Diclofenac at the indicated concentrations was added into both apical and basolateral solutions after stimulation of CFTR-mediated apical Cl⁻ current by CPT-cAMP (100 µM). Apical Cl⁻ current analysis was performed with basolateral membrane of Calu-3 cells being permeabilized with amphotericin B (250 µg/ml). A representative current tracing of 5 separate experiments is shown. (C) Reversibility of inhibition of CFTR Cl⁻ channel activity by diclofenac (20 µM) in T84 cells. Apical Cl⁻ current analysis was performed with basolateral membrane permeabilization by amphotericin B. After stabilization of diclofenac -inhibited apical Cl⁻ current, solutions containing forskolin and diclofenac were removed, and hemichambers were gently washed three times and filled with fresh solutions containing only forskolin. At the end of experiment, CFTR_inh-172 (5 µM) was added into apical solutions. A representative current tracing of 4 separate experiments is shown.