Figure 7.

FXYD5 is required for the NF-κB-mediated secretion of cytokines in response to IFN-α and TNF-α by alveolar epithelial cells. (A) A549 cells were transfected with a control (siCT) or FXYD5 specific (siFXYD5) siRNA, 24 h later treated with 100 U/ml IFN-α for the indicated period of time, and cell lysates were analyzed by immunoblot using specific antibodies as indicated. Bottom panel: representative immunoblots. Top panel: densitometric quantification of pIκBα in relation to total IκBα n = 4. (B,C) A549 cells were treated as in (A), culture media was collected, and IL-6 (B) and CCL2 (C) were determined by ELISA n = 3. (D). A549 cells were transfected with siCT or siFXYD5, 24 h later treated with 50 ng/ml TNF-α for the indicated period of time, and cell lysates were analyzed by immunoblot using specific antibodies as indicated. Representative immunoblots n = 4. (E,F) A549 cells were treated as in (D), culture media was collected, and IL-6 (E) and CCL2 (F) were determined by ELISA n = 4. Values of PBS-treated controls were normalized to 1. Bars represent means ± SD. Statistical significance was analyzed by one way ANOVA and Sidak’s multiple comparison test. *p ≤ 0.05; **p ≤ 0.01; ***p ≤ 0.001; ****p ≤ 0.0001, ns, non-significant.