Fig. 1

Sunitinib impairs VEGF-induced VE-cadherin tyrosine phosphorylation and cleavage: primary ECs were grown 3–4 days to reach confluency (3 × 10⁶ cells). After 3 h of serum starvation, ECs were stimulated by VEGF alone (50 ng/mL) or in the presence of SUT (20 μM) or TEM (50 ng/mL) during indicated times. Lysates were analysed by SDS-PAGE on 10% polyacrylamide gels and western blotting (5 μg of total protein lysate/lane). a Analysis of phosphotyrosine-containing protein pattern using the anti-phosphotyrosine antibody 4G10. b, c Analysis of phospho-Y⁶⁸⁵VE-cadherin d. Conditioned media from untreated ECs (CTL), or VEGF-stimulated-ECs or VEGF-stimulated-ECs treated with SUT or TEM for 20 min (same amount of cells in each condition) were collected, centrifuged to discard floating cells, and concentrated on Centriprep Centrifugal Filter Units with an Ultracel YM-30 membrane. Volume of 5 μL of concentrated medium of each condition was analysed by SDS-PAGE and western blotting using the monoclonal anti-VE-cadherin antibody (BV9 clone). e Same experiment as in d with VEGF-treated ECs in the presence of either INFα2 A (7.5 ng/mL), Irinotecan (IRN, 10 μM) and SN-38 (10 μM). Results are representative of three independent experiments