Effect of vascular endothelial growth factor and epidermal growth factor on iatrogenic apoptosis in human endothelial cells

To study the effect of growth factors on iatrogenic apoptosis, we examined the influence of vascular endothelial growth factor (VEGF) and epidermal growth factor (EGF) on staurosporine-induced apoptosis in primary cultures of human umbilical vein endothelial cells (HUVEC). Apoptosis was evaluated by a cell viability test, the TUNEL-POD assay and the activation of the pro-apoptotic caspase-3. Staurosporine (10-100 nM) caused the activation of caspase-3. This effect was manifest after 2 hr of incubation and reached its maximum after 5 hr. Severe loss of viability followed within 18 hr. VEGF or EGF (10-100 ng/mL) added together with staurosporine decreased the activation of caspase-3. The loss of viability was 24 hr delayed. The action of growth factors was observed at 1% serum concentration but also at concentration optimal for HUVEC survival (10%, v/v). Furthermore, the inhibition of PI-3 kinase (PI-3K) by wortmannin or LY294002 as well as the inhibition of MEK by PD098059 or U0126 prevented the protective effect of VEGF and EGF. Western blotting analysis showed that after 3 hr of incubation with staurosporine the level of the anti-apoptotic protein Mcl-1 decreased and this effect was reverted by VEGF. It is concluded that VEGF and EGF antagonize the pro-apoptotic action of staurosporine by the combined signalling of PI-3K and ERKs pathways. (C) 2003 Elsevier Inc. All rights reserved.