(A–C) Primary IPF lung fibroblasts were transfected with Brd4 siRNA for 48 hours. (A) Cells were collected and subjected to Western blots for Nox4, Brd4, and β-actin. (B) Densitometry of Brd4 and Nox4 protein expression relative to β-actin, as in A. *P < 0.05, Brd4 siRNA compared with NT control, by 2-tailed t test. (C) RNA from NT and Brd4 siRNA–treated cells was analyzed for Nox4 mRNA by real-time PCR. *P < 0.05, compared with the NT control of the same cell line, by 2-tailed t test. (D) BET inhibitors, BET-762 (0.5 μM), JQ1 (1 μM), and OTX015 (0.5 μM) were added to primary IPF lung fibroblasts at 70% confluence for 48 hours, and RNA was collected and Nox4 mRNA expression analyzed by real-time PCR. Triangles, squares, or circles indicate 3 different IPF individuals from whom primary cells were derived. Expressed values represent mean ± SD; n = 3 experimental replicates of each cell line. *P < 0.05, treated group vs. control (vehicle) group, by 2-tailed t test.