HRG-β1 induces phosphorylation of Smad2 in SK-BR-3 (a) and MCF7 (b) cells. (a) After 16 h of serum starvation in serum-free medium, SK-BR-3 cells were treated with 25 ng/ml of HRG-β1 for the indicated times. Immunoblots were probed with anti-phospho-Smad2 and anti-Smad2 antibodies. (b) The phosphorylation of Smad2 and total Smad2 were analyzed by western blotting in MCF7 cells. In all cases, β-actin was evaluated as a loading control. Data represent the means ± SD of three independent experiments. *P < 0.05, significant difference.