Figure 4.
TGF-β treated with DTT functions as competitive inhibitors. (A) Comparison of the luciferase activity released by TMLC cells that were incubated with recombinant human TGF-β (°) or DTT-treated TGF-β (•). Recombinant TGF-β is activate below 25 pM, while the DTT-treated TGF-β only showed activity at 5 nM. (B) TMLC cells (1.0 × 104 TMLC cells/well) were incubated with recombinant human TGF-β at 50 pM and increasing concentrations of DTT-treated TGF-β. Competitive agonist effect of DTT-treated TGF-β was apparent at ~100 fold higher concentration. The data shown are representative of three independent studies done in quadruplicate. (C) Mouse epithelial Akr cells (1.0 × 106 cells/condition) containing TGF-β receptors, were treated with 5 ng/ml of rhTGF-β and phosphorylated SMAD2/3 was measured by western blotting. The blots were quantitated by densitometry (bar graph) and 50 ng/ml monomeric TGF-β lowers the impact of the dimeric TGF-β (p < 0.01 vs non stimulated, p = 0.127 vs dimer treated). The data are representative of eleven independent studies. Error bars represent the standard error of the mean.