Figure 1

TGF-β1 suppresses VEGF-D expression in lung fibroblasts. (A, B) MRC-5 fibroblasts were incubated with 5 ng/mL TGF-β1 for the indicated time points. Total RNA was isolated and reverse-transcribed, and the resultant cDNA was subjected to quantitative real-time PCR to detect gene expressions of VEGF-A (red), VEGF-C (blue), VEGF-D (black) (A) and PAI-1 (B). Real-time PCR results were normalized to β2-macroglobulin and expressed as the fold-change relative to unstimulated control cells (Ctrl). (C–F) MRC-5 fibroblasts were stimulated with TGF-β1 (5 ng/mL) for the indicated time (C, D) or stimulated with various concentrations of TGF-β1 for 48 h (E, F). Equal amounts of protein from whole cell lysates were analyzed by Western blotting with antibodies against VEGF-D and β-actin. (G, H) Primary NLFs were stimulated with TGF-β1 (5 ng/mL) for 48 h. Equal amounts of protein from whole cell lysates were analyzed by Western blotting with antibodies against VEGF-D and β-actin. (D, F, H) Ratio of VEGF-D to β-actin density was expressed as the fold change relative to unstimulated control cells in all experiments performed. Data are expressed as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, compared with control, by one-way ANOVA (A, B, D, F) or Student t test (H).