Fig. 5.

Mechanism of action of ARN7149. (A) Representative traces (left) and summary of data (right) from short-circuit current recordings on human cultured bronchial epithelia. Ca²⁺-dependent Cl⁻ secretion mediated by TMEM16A was triggered with 0.25 µM UTP on the apical side, in the presence of vehicle or indicated compounds: ARN7149 (10 µM), ARN11391 (20 µM), or ARN4550 (20 µM). To avoid the confounding effect of other channels, recordings were done in the presence of: amiloride (10 µM) to block ENaC; paxillin (10 µM) to block large conductance Ca²⁺-dependent K⁺ channels; inh-172 (10 µM, apical) to block CFTR. **P <0.01; ***P <0.001 vs. vehicle (ANOVA with Dunnett’s post-hoc test). (B) Role of P2RY2 in mediating the effect of UTP on Ca²⁺ mobilization. Left: representative traces showing Fluo-4 fluorescence time-course in null FRT cells following extracellular addition of UTP (0.25 µM). Cells were preincubated with/without AR-C118925XX (10 µM) antagonist ± ARN7149, ARN11391, or ARN4550. Right: summary of data. The symbols report the maximal change in Fluo-4 fluorescence. ***P <0.001 vs. experiments without AR-C118925XX (ANOVA with Dunnett’s post-hoc test). (C) Effect of compounds on Ca²⁺ mobilization triggered by SLIGR-NH2 (protease-activate receptor agonist) in HEKR1 (top) and HEKR2 (bottom) cells. Left: representative traces showing Fluo-4 fluorescence time-course following extracellular addition of SLIGR-NH2 (10 µM for HEKR1 and 4 µM for HEKR2). Cells were preincubated with vehicle or indicated compounds (10 µM). Right: summary of data. The symbols report the maximal change in fluorescence. *P <0.05; **P <0.01; ***P <0.001 vs. vehicle (ANOVA with Dunnett’s post-hoc test).