Fig. 7

PKA is involved in cAMP-mediated negative regulation of IgG-IC-induced expressions of pro-inflammatory mediators and acute lung injury. RAW264.7 cells are pre-treated with 2 mM of PBS/PKA agonist/Epac agonist for 30 min. Then the cells are treated with or without 100 µg/ml of IgG-IC. Cell-free supernatants are harvested 6 h later. ELISA is used to measure expressions of TNF-α (A), MIP-1α (B), and MIP-1β (C). RAW264.7 cells are pre-treated with 0.2 mM of PBS/PKA agonist/Epac agonist for 30 min. Then the cells are treated with or without 100 µg/ml of IgG-IC. RNAs are harvested 3 h later. qPCR is used to measure expressions of TNF-α (D), MIP-1α (E), and MIP-1β (F). H-89 is dissolved in DMSO to obtain 10 mM solution. RAW264.7 cells are pre-treated with DMSO (1:500 dilution), Rolipram (10 µM) or H-89 (10 µM) + Rolipram (10 µM) for 1 h. Then the cells are treated with or without 100 µg/ml of IgG-IC. RNAs are harvested 3 h later. qPCR is used to measure expressions of TNF-α (G), MIP-1α (H), and MIP-1β (I). J. Rolipram and H-89 are dissolved in DMSO to obtain 10 mg/ml, and 25 mg/ml solutions, respectively. For DMSO group, mice are treated according to the following principle: 0.5 ml/kg. For Rolipram group, mice are treated according to the following principle: 1 mg/kg Rolipram + 0.4 ml/kg DMSO. For Rolipram + H-89 group, mice are treated according to the following principle: 1 mg/kg Rolipram + 10 mg/kg H-89. Mice are treated by intraperitoneal injection of DMSO, Rolipram or H-89 + Rolipram, which is followed by airway deposition of IgG-IC. 4 h later, lungs are harvested and histological changes are assessed. Data are expressed as mean ± SEM (N = 6 for ELISA, and N = 3 for qPCR). *, ** and *** suggest p < 0.05, 0.01 and 0.01, respectively