Sphingosine 1-phosphate (Sph-1-P) is now regarded as a lysophospholipid mediator which is present in plasma and other body fluids, and exerts potent and pleiotropic biological effects [1,2]. Such extracellular mediator activities of Sph-1-P are mainly mediated by subfamilies of G proteincoupled receptors, of which the most completely characterized are those encoded by the endothelial differentiation genes (EDGs) [2,3]. Previously, we showed that in platelets, Sph-1-P is rapidly formed from sphingosine (Sph) by the action of Sph kinase, abundantly stored intracellularly, and released into the extracellular environment upon stimulation [1,4]. Furthermore, vascular endothelial cells express the EDG family Sph-1-P receptors, i.e., EDG-1 and EDG-3 [5]. Consistent with the expression of the high-affinity Sph-1-P receptors in these cells, nanomolar Sph-1-P reportedly induces a variety of vascular endothelial cell responses [5–9]. Sph-1-P stimulates endothelial cell survival or proliferation through a Gi-coupled receptor, probably EDG-1 [5,7]. Furthermore, Sph-1-P induces adherens junction assembly, migration, capillary tube formation, and promotion of angiogenesis [5,8,9]. In this case, the signals mediated via EDG-1 (coupled with Gi) and EDG-3 (coupled with G13 and Gq) are both necessary, and the small GTPase Rhoand Rac-mediated signalings are also involved [5,8]. Very recently, Sph-1-P has been shown to activate endothelial nitric oxide synthase through a novel mechanism involving Akt activation and produce nitric oxide, which plays an important role in endothelial survival and maintenance of the endothelial function [10]. Accordingly, Sph-1-P should be added to the list of platelet-derived bioactive mediators and may play an important role in platelet–endothelial cell interactions, which constitute a central part of vascular biology. To obtain insight into the physiological and pathophysiological role(s) of Sph-1-P in the vascular system, information about the regulation of endothelial cell expression of EDG family Sph-1-P receptors, i.e., EDG-1 and EDG-3, is very important. In fact, the EDG-1 cDNA was originally isolated as a phorbol ester-inducible immediate early transcript from vascular endothelial cells; morphogenetic differentiation of vascular endothelial cells into capillary-like tubules was induced under the conditions [11]. Furthermore, recent data have shown that the mechanical force associated with blood flow, i.e., the fluid shear stress, modulates vascular structure and function, and plays an important role in the pathogenesis of vascular diseases, including atherosclerosis, hypertension, and restenosis [12]. Shear stress has been reported to affect endothelial cell gene expression, and EDG-1 has been cloned as one of two cDNAs encoding a G protein-coupled receptor from a cDNA library of human umbilical vein endothelial cells (HUVECs) exposed to fluid shear stress; EDG-1 mRNA levels increase markedly in response to fluid flow [13]. Accordingly, the modulation of Sph-1-P/EDG signaling may be involved in important vascular responses related to angiogenesis and blood flow conditions. In this study, we examined the possibility that EDG expression, and hence, responsiveness to Sph-1-P in vascular endothelial cells, may be modulated by tumor necrosis factor-a (TNF-a), which is a pleiotropic cytokine that