Fig. 1

Involvement of extracellular ATP in MUC5AC release in poly(I:C)-treated NCI-H292 cells. a-c Effect of poly(I:C) on cell viability and the release of extracellular ATP and MUC5AC from NCI-H292 cells. Cells were treated with 10 μg/ml poly(I:C). At various time points after the incubation, whole cells and supernatants were harvested and assayed. a Supernatants were assessed for the concentration of extracellular ATP by fluorometric assay. b Cell viability was assessed by MTT assay. Cell viability was calculated as the percentage of viable cells among poly(I:C)-treated cells at 0 min. c Supernatants were assayed for MUC5AC release by ELISA. (D-E) Release of MUC5AC in ATP-treated cells. d Cells were treated with 10 μM ATP. At various time points after the incubation, the supernatants were harvested and assayed. e Cells were treated with various concentrations of ATP. After 24 h, the supernatants were harvested and assayed for MUC5AC release. f Panels show representative photographs of the immunoreactivity of MUC5AC in NCI-H292 cells. Treatment with 10 μM ATP augmented the immunoreactivity of MUC5AC. Original magnification: x200. g, h Effect of a pannexin channel inhibitor, carbenoxolone (CBX), on the release of extracellular ATP and MUC5AC in poly(I:C)-treated cells. g Cells were treated with 10 μM CBX or vehicle 1 h prior to the treatment with 10 μg/ml poly(I:C). At various points after incubation, the supernatants were harvested and assessed for the concentration of extracellular ATP. h Various concentrations of CBX were added 1 h before 10 μg/ml poly(I:C) treatment. After 24 h, the supernatants were harvested and assayed for MUC5AC release. The data are expressed as the means ± SEM for three to six separate experiments. *p < 0.05, **p < 0.01, ***p < 0.001 compared with the values of vehicle-treated control. ⁺⁺⁺ p < 0.001 compared with the values of poly(I:C)-treated control