Sphingosine 1-phosphate (S1P) is a bioactive sphingolipid metabolite involved in many critical cellular processes including proliferation, survival, and migration, as well as angiogenesis and allergic responses. S1P levels inside cells are tightly regulated by the balance between its synthesis by sphingosine kinases and degradation. S1P is interconvertible with ceramide, which is a critical mediator of apoptosis. It has been postulated that the ratio between S1P and ceramide determines cell fate. Activation of sphingosine kinase by a variety of agonists increases intracellular S1P, which in turn can function intracellularly as a second messenger or be secreted out of the cell and act extracellularly by binding to and signaling through S1P receptors in autocrine and/or paracrine manners. Recent studies suggest that this “inside-out” signaling by S1P may play a role in many human diseases, including cancer, atherosclerosis, inflammation, and autoimmune disorders such as multiple sclerosis. In this review we summarize metabolism of S1P, mechanisms of sphingosine kinase activation, and S1P receptors and their downstream signaling pathways and examine relationships to multiple disease processes. In particular, we describe recent preclinical and clinical trials of therapies targeting S1P signaling, including 2-amino-2-propane-1,3-diol hydrochloride (FTY720, fingolimod), S1P receptor agonists, sphingosine kinase inhibitors, and anti-S1P monoclonal antibody.

“Inside-Out” Signaling of Sphingosine-1-Phosphate: Therapeutic Targets

HDL is a major atheroprotective factor, but the mechanisms underlying this effect are still obscure. HDL binding to scavenger receptor-BI has been shown to activate eNOS, although the responsible HDL entities and signaling pathways have remained enigmatic. Here we show that HDL stimulates NO release in human endothelial cells and induces vasodilation in isolated aortae via intracellular Ca2+ mobilization and Akt-mediated eNOS phosphorylation. The vasoactive effects of HDL could be mimicked by three lysophospholipids present in HDL: sphingosylphosphorylcholine (SPC), sphingosine-1-phosphate (S1P), and lysosulfatide (LSF). All three elevated intracellular Ca2+ concentration and activated Akt and eNOS, which resulted in NO release and vasodilation. Deficiency of the lysophospholipid receptor S1P3 (also known as LPB3 and EDG3) abolished the vasodilatory effects of SPC, S1P, and LSF and reduced the effect of HDL by approximately 60%. In endothelial cells from S1P3-deficient mice, Akt phosphorylation and Ca2+ increase in response to HDL and lysophospholipids were severely reduced. In vivo, intra-arterial administration of HDL or lysophospholipids lowered mean arterial blood pressure in rats. In conclusion, we identify HDL as a carrier of bioactive lysophospholipids that regulate vascular tone via S1P3-mediated NO release. This mechanism may contribute to the vasoactive effect of HDL and represent a novel aspect of its antiatherogenic function.

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HDL induces NO-dependent vasorelaxation via the lysophospholipid receptor S1P3

http://www.scripps.edu/chun/PDFs/Sanna%20JBC%202004.pdf

Sphingosine 1-Phosphate (S1P) Receptor Subtypes S1P1 and S1P3, Respectively, Regulate Lymphocyte Recirculation and Heart Rate

Sphingosine-1-phosphate (S1P) is a biologically active metabolite of plasma-membrane sphingolipids that is essential for immune-cell trafficking. Its concentration is increased in many inflammatory conditions, such as asthma and autoimmunity. Much of the immune function of S1P results from the engagement of a family of G-protein-coupled receptors (S1PR1-S1PR5). Recent findings on the role of S1P in immunosurveillance, the discovery of regulatory mechanisms in S1P-mediated immune-cell trafficking and new advances in understanding the mechanism by which S1P affects immune-cell function indicate that the alliance between S1P and its receptors has a fundamental role in immunity.

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The alliance of sphingosine-1-phosphate and its receptors in immunity

At sites of vascular injury, platelets come into contact with the subendothelial extracellular matrix which triggers their activation and the formation of a hemostatic plug. This process is crucial for normal hemostasis, but may also lead to pathological thrombus formation causing diseases such as myocardial infarction or stroke. The initial capture of flowing platelets is mediated by the interaction of the glycoprotein (GP) Ib-V-IX complex with von Willebrand factor (vWF) immobilized on exposed collagens. This interaction allows the binding of the collagen receptor GPVI to its ligand and to initiate cellular activation, a process that is reinforced by locally produced thrombin and soluble mediators released from platelets. These events lead to the shift of beta1 and beta3 integrins on the platelet surface from a low to a high affinity state, thereby enabling them to bind their ligands and to mediate firm adhesion, spreading, coagulant activity, and aggregation. This review summarizes the most important structural and functional properties of these adhesion receptors and briefly discusses their potential as targets for antithrombotic therapy.

Cell Adhesion Mechanisms in Platelets

Objectives: Sphingosine 1-phosphate (Sph-1-P), a bioactive lipid derived from activated platelets, may play an important role in coronary artery spasm and hence the pathogenesis of ischemic heart diseases, since we reported that a decrease in coronary blood flow was induced by this lysophospholipid in an in vivo canine heart model [Cardiovasc Res 46 (2000) 119] In this study, metabolism related to and cellular responses elicited by Sph-1-P were examined in human coronary artery smooth muscle cells (CASMCs) Methods and results: [3H]Sphingosine (Sph), incorporated into CASMCs, was converted to [3H]Sph-1-P intracellularly, but its stimulation-dependent formation and extracellular release were not observed Furthermore, the cell surface Sph-1-P receptors of S1P family (previously called EDG) were found to be expressed in CASMCs Accordingly, Sph-1-P seems to act as an extracellular mediator in CASMCs Consistent with Sph-1-P-elicited coronary vasoconstriction in vivo, Sph-1-P strongly induced CASMC contraction, which was inhibited by JTE-013, a newly-developed specific antagonist of S1P2 (EDG-5) Furthermore, C3 exoenzyme or Y-27632 inhibited the CASMC contraction induced by Sph-1-P, indicating Rho involvement Finally, exogenously-added [3H]Sph-1-P underwent a rapid degradation Since lipid phosphate phosphatases, ectoenzymes capable of dephosphorylating Sph-1-P, were expressed in CASMCs, Sph-1-P may be dephosphorylated by the ectophosphatases Conclusions: Sph-1-P, derived from platelets and dephosphorylated on the cell surface, may induce the contraction of coronary artery smooth muscle cells through the S1P2/Rho signaling

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Sphingosine 1-phosphate induces contraction of coronary artery smooth muscle cells via S1P2.

Interaction between von Willebrand factor and glycoprotein Ib activates Src kinase in human platelets: role of phosphoinositide 3-kinase

In this study, we examined the metabolism of [3H]N-hexanoylsphingosine [C6-ceramide (Cer)] in neutrophils, erythrocytes, platelets and mononuclear cells. [3H]C6

Ceramide 1-phosphate formation in neutrophils.

To analyze the involvement in allergic reactions of platelets and sphingosine 1-phosphate (Sph-1-P), a lysophospholipid mediator released from activated platelets, the effects of Sph-1-P and a supernatant prepared from activated platelets on mast cell line RBL-2H3 were examined. Sph-1-P strongly inhibited the migration of both non-stimulated and fibronectin-stimulated RBL-2H3 cells, which was reversed by JTE-013, a specific antagonist of G protein-coupled Sph-1-P receptor S1P(2); S1P(2) was confirmed to be expressed in these cells. A similar anti-motility effect of Sph-1-P was observed in a phagokinetic assay. Consistent with these results, treatment of RBL-2H3 cells with Sph-1-P resulted in a rounded cell morphology, which was blocked by JTE-013. Under the present conditions, Sph-1-P failed to induce intracellular Ca(2+) mobilization or histamine degranulation, responses postulated to be elicited by intracellular Sph-1-P. Importantly, the Sph-1-P effect, i.e., the regulation of RBL-2H3 cell motility, was mimicked by the supernatant (both with and without boiling) prepared from activated platelets, and this effect of the supernatant was also blocked by JTE-013. Our results suggest that the motility of mast cells can be regulated by Sph-1-P and also platelets (which release Sph-1-P), via cell surface receptor S1P(2) (not through intracellular Sph-1-P actions, postulated previously in the same cells).

Toshiro Takafuta

Papers

Sphingosine 1-phosphate induces contraction of coronary artery smooth muscle cells via S1P2.

Interaction between von Willebrand factor and glycoprotein Ib activates Src kinase in human platelets: role of phosphoinositide 3-kinase

Ceramide 1-phosphate formation in neutrophils.

Sphingosine 1-Phosphate Inhibits Migration of RBL-2H3 Cells via S1P2: Cross-Talk between Platelets and Mast Cells

Evaluation of platelet function under high shear condition in the small-sized collagen bead column.