Fig. 2.

(A) Effect of actinomycin D (Act D) and cycloheximide (CHX) on FnCW-induced release of CCL20 by HOECs. HOEC monolayers (mixed donors; N = 3) were pretreated with actinomycin D or cycloheximide for 5 min and then incubated with or without FnCW (10 μg/ml) for an additional 6 h. As a negative control, cells were incubated with medium or medium with vehicle control (DMSO) for actinomycin D. CCL20 levels in cell-free supernatants were measured by ELISA. (B and C) Effect of BFA on CCL20 release by HOECs. HOEC monolayers (mixed donors; N = 3) were pretreated with BFA (10 μg/ml) for 5 min, followed by incubation with or without FnCW (10 μg/ml) for an additional 6 h. As a negative control, cells were incubated with medium alone or medium with vehicle control (ethanol) for BFA. CCL20 levels in cell-free supernatants (B) and cell lysates (C) were measured by ELISA. Results are the means ± standard deviations of three independent experiments. *, P < 0.05. For comparisons of CCL20 content in supernatants and in cell pellets, the levels of CCL20 are expressed as ng/well instead of ng/ml. One ml of media per well was used in the six-well format.