Figure 5.

TBXA2R antagonism reduces proliferation, migration, and activation of fibroblasts from lungs of mice and patients with IPF. (A) Lung fibroblasts were isolated from lungs of mice and treated with F2-isoprostanes, ifetroban (Ifet), or both. Western blots are shown for evaluation of α-SMA, total Smad2/3 (T-Smad2/3), phospho-Smad2/3 (P-Smad2/3), Timp1 (tissue inhibitor of metalloproteinase-1), total p44/42 (T-p44/42), phospho-p44/42 (P-p44/42), total Akt (T-AKT), and phospho-Akt (P-AKT). (B–E) Lung fibroblasts were isolated from explanted lungs of six patients with IPF and treated with Ifet (300 nM) or vehicle. (B) Scratch wound closure assay measured as wound area closed at 24 hours. *P < 0.05 by Wilcoxon signed rank test. Error bars denote SEM. (C) Cell proliferation measured by BrdU incorporation. *P < 0.05 by Wilcoxon signed rank test. Error bars denote SEM. (D) Quantitative PCR for evaluation of profibrotic gene expression. Lines connect cells from the same patient with and without Ifet. Average of vehicle controls was set to 1. n = 6 per group. *P < 0.05 compared with vehicle-treated group by paired t test (each treated culture was paired to the same culture with no treatment). (E) Western blot for evaluation of T-Smad2/3, P-Smad2/3, Timp1, T-p44/42, P-p44/42, T-AKT, and P-AKT. ACTA2 = actin alpha 2, smooth muscle; α-SMA = α-smooth muscle actin; BrdU = bromodeoxyuridine; COL1A1 = collagen type I alpha 1 chain; COL1A2 = collagen type I alpha 2 chain; Hsp70 = heat shock protein 70; IPF = idiopathic pulmonary fibrosis; SERPINE1 = serpin family E member 1; TBXA2R = thromboxane–prostanoid receptor; Veh = vehicle.