Vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2), a tyrosine kinase receptor, is activated by VEGF and fluid shear stress (FSS), and its downstream signaling is important in regulation of endothelial functions, such as cell migration, endothelium-dependent relaxation, and angiogenesis. Inhibition of VEGFR2 augments cigarette smoke (CS)-induced oxidative stress and inflammatory responses leading to endothelial dysfunction. CS-derived reactive oxygen/nitrogen species interact with VEGFR2, causing posttranslational modifications that render VEGFR2 inactive for downstream signaling, resulting in endothelial dysfunction. CS-mediated oxidants/carbonyl stress decreases SIRT1 levels and causes eNOS acetylation, which has ramifications in endothelial dysfunction. CS also affects endothelial cell survival pathway by disrupting VEGF- and FSS-mediated VEGFR2/PI3-kinase signaling, leading to decreased Akt phosphorylation and eNOS acetylation. We describe here the mechanisms whereby CS alters VEGF- and FSS-mediated VEGFR2-eNOS signaling, which may have implications for understanding the pathogenesis of pulmonary and cardiovascular diseases.