Fig. 4: Omega-3 epoxides suppress the activation of lung fibroblasts.

a Relative number of lung fibroblasts stimulated by lipid extracts from cultured medium of BMMCs when treated with or without ω-3 epoxides (1 μM) for 48 h (n = 4). Data are representative of 2 independent experimental replicates. b Relative number of lung fibroblasts treated with vehicle, 17,18-EpETE (1 μM), 17,18-diHETE (1 μM), 19,20-EpDPE (1 μM), 19,20-diHDoPE (1 μM), EPA (1 μM), or DHA (1 μM) for 48 h (n = 4). Data are representative of 3 independent experimental replicates. c Relative expression levels of Col1a1, Acta2, and Il6 mRNA in lung fibroblasts treated with vehicle, 17,18-EpETE (1 μM), or 19,20-EpDPE (1 μM) for 6 h when stimulated with or without TGF-β (2.5 ng/ml) (n = 4). Expression levels were normalized to those of 18S ribosomal RNA and then to those in the unstimulated control fibroblasts. Data are representative of 3 independent experimental replicates. d Immunostaining of SM22α in lung fibroblasts treated with vehicle, 17,18-EpETE (1 μM), 19,20-EpDPE (1 μM), EPA (1 μM), or DHA (1 μM) for 24 h when stimulated with or without TGF-β (2.5 ng/ml) (left). Scale bar, 50μm. Ratio of SM22α-positive cells to total lung fibroblasts (right) (n = 4). Data are representative of 2 independent experimental replicates. e Western blotting of pSmad2, Smad2, and β-Actin in total protein extracts from lung fibroblasts when administered vehicle, 17,18-EpETE (1 μM), or 19,20-EpDPE (1 μM) with or without TGF-β stimulation (1 ng/ml and 2.5 ng/ml) for 15 min (left) and quantification by densitometry (right). Experiments were repeated three times and the data were pooled. Data are mean ± SEM. P values were determined by one-way ANOVA with Dunnett’s post hoc test (a–d) or two-way ANOVA with Tukey’s post hoc test (e).