Figure 1

Activin A inhibited BMP-9-induced apoptosis. (A) The expression of the type I receptors ALK4 and ALK7 in eight human myeloma cell lines was determined using QRT-PCR. The delta delta Ct method using GAPDH as housekeeping gene was used to determine the relative levels of mRNA compared to the expression of ALK7 in cell line JJN-3 (Ct-value = 33) which was set to 1. (B) Phosphorylation of SMAD2 was determined using immunoblotting in INA-6 and IH-1 cells treated with activin A (10 ng/mL) or TGF-β (5 ng/mL) for 4 hours. GAPDH was used as loading control. INA-6 (C) and IH-1 cells (D) were treated for three days with activin A (10 ng/mL) and the indicated concentrations of BMP-9 before cell viability was determined by flow cytometry using annexin V/PI labeling. Cells that were negative for both annexin V and PI were considered viable. Phosphorylation of SMAD1/5/8 or SMAD2 was determined using immunoblotting in INA-6 (E) and IH-1 cells (F) treated with activin A (10 ng/mL) and/or BMP-9 (0.5 ng/mL) for one, six and 24 hours. GAPDH was used as loading control.