b isoforms activate the ERK signalling pathway through VEGF receptors. Effect of VEGF121/165b isoforms on downstream signalling pathways, using antibodies against Y1175 in VEGFR2 (KDR), S473 in AKT and T202/Y204 in ERK1/2, after treatment of HUVECs with 100 ng/mL of each cytokine. VEGF165 stimulates phosphorylation of VEGFR2 and ERK1/2. This effect is not blocked by co-administration of either VEGF121b from P. pastoris or VEGF165b of different origins (mammalian cells or P. pastoris). Furthermore, addition of VEGF121/165b isoforms alone produces signalling activation as well. KDR/ERK1/2 phosphorylation was shown to be mediated specifically through VEGF receptors, since treatment with 1 μM of the specific inhibitor GW654652 blocks phosphorylation of the receptor and the intracellular transducer.