Fig. 5From: Recombinant RGD-disintegrin DisBa-01 blocks integrin αvβ3 and impairs VEGF signaling in endothelial cellsDisBa-01 inhibits ERK1/2 and PI3K phosphorylation. HUVECs (5 × 105 cells/well) were seeded in 6-well plates and left to adhere at 37 °C, 5% CO2, overnight, followed by a period of 24 h of starvation at serum-free medium. Cells were treated with 1 ml of DMEM supplemented with 10% FBS and either DisBa-01 (1000 nM), VEGF (10 ng/mL) or a co-treatment and incubated for 1 and 24 h at 37 °C, 5% CO2, followed by cell lysis. Twenty micrograms of protein from the cell lysate were separated on SDS-PAGE. Blots were probed with antibodies to a P-TY187 ERK1 + ERK2 and anti-ERK1 + ERK2; to b P-TY607 PI3K and anti-PI3K and GAPDH, this last used to normalize loading. Bands corresponding to all proteins were quantified by densitometry using the ImageJ FIJI program. Bar graph shows the mean ± SE of phosphorylated ERK1 + ERK2/ERK1 + ERK2/GAPDH and PI3K/PI3K/GAPDH expression from three independent experiments. Values of *p < 0.05 were significantly different when compared to untreated (a), DisBa-01 (b) and VEGF (c) groupsBack to article page