Figure 4. ALK2 AON-induced exon skipping decreased BMP signaling in 2H11 endothelial cells.

(A) 2H11 cells were cotransfected with 100 nM of the indicated AON and the BMP reporter construct BRE-Luc for 16 hours. Subsequently, cells were starved for 8 hours, and then stimulated with 50 ng/ml BMP6 overnight. Luciferase reporter activity was measured after stimulation and normalized with β-gal activity. (B) 2H11 cells were transfected with 100 nM control AON or 100 nM ALK2 AON in proliferation medium. One day after transfection, the cells were serum starved for overnight and stimulated with 5 ng/ml BMP6 for 1 hour. Protein was isolated and western blotting was performed to check the Smad1/5/8 phosphorylation. GAPDH was used as loading control. Data are means ± SD from three independent experiments. Statistical analysis was performed using Student’s t-test, using the untransfected samples as reference. *P<0.05, **P<0.005.