Tirofiban counteracts endothelial cell apoptosis through the VEGF/VEGFR2/pAkt axis

Abstract

Tirofiban is used in the treatment of patients with acute coronary syndrome submitted to percutaneous coronary intervention (PCI). We have, previously, shown that tirofiban stimulates VEGF expression and promotes proliferation of endothelial cells. VEGF is a well known inhibitor of endothelial cell apoptosis. TNF-α is a pro-apoptotic cytokine released in the site of a vascular injury, including balloon angioplasty. We thought to investigate whether tirofiban was able to protect endothelial cells from cell death induced by TNF-α. For this study, we used human umbilical vein endothelial cells (HUVEC). Analysis of apoptosis was performed by propidium iodide incorporation, annexin V staining and measure of active caspase 3 levels. Western blot served for a semiquantitative measure of Akt activation, VEGF, and the pro-apoptotic Bim and Bak. Our results show that TNF-α was unable to activate caspase 3 and produce cell death in the presence of tirofiban. Activation of apoptosis was preceded by upregulation of Bim and Bak that resulted decreased after addition of tirofiban. The anti-apoptosis effect of tirofiban was reproduced by VEGF and counteracted by VEGFR2 blockade and the cation chelating agent ethylene glycol tetraacetic acid (EGTA). The use of p-Akt inhibitor, BEZ235,and Akt knockdown, suggested that pAkt mediated the prosurvival effect of tirofiban. In conclusion, tirofiban protects endothelial cells from apoptosis stimulated by TNF-α, due to its ability to stimulate VEGF production.