Fig. 3. Regulation of α-SMA, transforming growth factor (TGF)-β Receptors, and fibronectin in lung fibroblasts from control and interstitial pneumonia (IP) subjects. (A-D) Effect of TGF-β1 and BIX02189 on α-SMA (A), phospho-TGF-βRI (B), TGF-βRI (C), and TGF-βRII (D) expression. Sub-confluent fibroblasts were cultured in serum-free (SF)-DMEM for 24 h and then incubated in the presence or absence of TGF-β1 or BIX02189 for 8 h (B, C) or 24 h (A, D). Total protein was extracted, and western blotting was performed with specific antibodies against α-SMA, phospho-TGF-βRI, TGF-βRI, and TGF-βRII. The vertical axes show the relative intensity of α-SMA, phospho-TGF-βRI, TGF-βRI, and TGF-βRII vs. β-actin; the horizontal axes show the conditions. (E) Effect of BIX02189 on TGF-β1-stimulated fibronectin production by lung fibroblasts. Sub-confluent fibroblasts were cultured in serum-free (SF)-DMEM for 2 h and incubated in the presence or absence of TGF-β1 or BIX02189 for 24 h. The supernatant was harvested, and fibronectin level was quantified. The vertical axis shows fibronectin production, expressed as amount per 10⁵ cells/ml; the horizontal axis shows the conditions. The values represent mean ± SEM of at least three independent experiments. (A-E) *P<0.05, ^†P<0.01, ^§P<0.001.