Sphingosine-1-phosphate promotes lymphangiogenesis by stimulating S1P1/Gi/PLC/Ca2+ signaling pathways

Abstract

The lymphatic system plays pivotal roles in mediating tissue fluid homeostasis and immunity, and excessive lymphatic vessel formation is implicated in many pathological conditions, which include inflammation and tumor metastasis. However, the molecular mechanisms that regulate lymphatic vessel formation remain poorly characterized. Sphingosine-1-phosphate (S1P) is a potent bioactive lipid that is implicated in a variety of biologic processes such as inflammatory responses and angiogenesis. Here, we first report that S1P acts as a lymphangiogenic mediator. S1P induced migration, capillarylike tube formation, and intracellular Ca 2؉ mobilization, but not proliferation, in human lymphatic endothelial cells (HLECs) in vitro. Moreover, a Matrigel plug assay demonstrated that S1P promoted the outgrowth of new lymphatic vessels in vivo. HLECs expressed S1P1 and S1P3, and both RNA interference-mediated down-regulation of S1P1 and an S1P1 antagonist significantly blocked S1P-mediated lymphangiogenesis. Furthermore, pertussis toxin, U73122, and BAPTA-AM efficiently blocked S1P-induced in vitro lymphangiogenesis and intracellular Ca 2؉ mobilization of HLECs, indicating that S1P promotes lymphangiogenesis by stimulating S1P1/G i /phospholipase C/Ca 2؉ signaling pathways. Our results suggest that S1P is the first lymphangiogenic bioactive lipid to be identified, and that S1P and its receptors might serve as new therapeutic targets against inflammatory diseases and lymphatic metastasis in tumors. (Blood. 2008; 112:1129-1138) IntroductionSphingosine-1-phosphate (S1P) has been implicated in a wide spectrum of biologic processes, including the promotion of cell growth and survival, migration and differentiation, platelet aggregation, inflammatory responses, and angiogenesis. 1 S1P is generated by the phosphorylation of sphingosine through a process mediated by sphingosine kinase 1 (SphK1) and SphK2. S1P acts both intracellularly as a second messenger 2 and extracellularly as a ligand for a family of G-protein-coupled S1P receptors. 3 S1P1 couples stringently to the G i protein family, whereas S1P2 and S1P3 couple to the G i , G q , and G 12/13 protein families. Multiple interconnections of S1P signaling through S1P1 and S1P3 induce vascular endothelial cell proliferation, migration, morphogenesis, cytoskeletal reorganization, and adherens junction assembly, whereas signaling via S1P2 negatively regulates S1P-mediated multiple responses of vascular endothelial cells. 4 The defective vascular maturation observed in S1P1-deficient mice highlights a fundamental role for S1P signaling on vasculogenesis. 5 Neutralization of the action of extracellular S1P shows significant inhibition of angiogenesis, tumor growth, metastasis, and lymphocyte transmigration, indicating that S1P is an important pathological regulator of inflammation and angiogenesis. [6][7][8] Lymphatic vessels play important roles in mediating tissue fluid homeostasis and immunity. 9 Although lymphangiogenesis, the formation of new lymphatic vessels from preexisting ves...

Phenotype-based high-content chemical library screening identifies statins as inhibitors of in vivo lymphangiogenesis

“…Our results demonstrate that the 3D sprouting of LECs into the collagen hydrogel (Movie S1) is morphologically similar to in vivo vessel sprouting as recently depicted in the mouse retina (25). We used a combination of VEGF-A, bFGF and S1P to promote lymphatic sprout formation since these factors are involved in pathological tumor and lymph node lymphangiogenesis (26,27), were previously validated in vitro (28), and yielded a more pronounced sprout formation in our assay than the individual growth factors. An advantage of our 3D assay is the automated image acquisition and analysis, which resolves these bottlenecks of conventional high-throughput screens.…”

Developmental and pathological lymphangiogenesis: from models to human disease

“…Recently, Sphingosine-1-phosphate (S1P), a potent bioactive lipid that is implicated in a variety of biologic processes such as inXammatory responses and angiogenesis, was reported as a lymphangiogenic mediator (Yoon et al 2008). S1P induces the migration and capillary-like tube formation of lymphatic endothelial cells in vitro and lymphangiogenesis in vivo.…”

Sphingosine 1-phosphate receptor 1 regulates the directional migration of lymphatic endothelial cells in response to fluid shear stress

“…S1P is a known chemoattractant for a variety of cell types [28,31,32,53,54]. Furthermore, activation of S1PR1 by S1P promotes lymphangiogenesis via Gai-dependent signalling [33]. However, the full recirculation of cell culture media in the IFC takes 5 min, whereas the HLMVECs migrate on the timescale of multiple hours.…”

“…It has been previously shown that S1P is required for lymphangiogenesis and EC motility [33]. We therefore hypothesized that S1P might itself play a necessary role in the inward migration triggered by impinging flow.…”

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