Figure 3

Activity of purified TGF-β NAC is dependent on Redox state. Purified active TGF-β (R&D Systems) was treated with either NAC or DTT as described, was then incubated in the absence or presence of 10 mM diamide for two hours at 37°C. Samples were dialyzed for two hours then either (A) incubated with TMLC cells to measured luciferase production or (B) separated by polyacrylamide gel electrophoresis and subjected to western blot analysis TGF-β. Data represents percent of control ± standard error where control is TGF-β sample untreated (p < 0.05 for NAC 2 mM and 10 mM versus TGF-β alone). Diamide restored activity levels to that of TGF-β alone (p > 0.05 of TGF-β alone versus TGF-β + NAC (20 mM) or TGF-β + DTT (10 mM) + diamide). Recombinant human TGF-β migrated as a ~25 kD protein that was reduced to a ~12.5 kD protein by the addition of NAC or DTT. Data shown in (B) is a representative of six independent experiments.