Critical role of sphingosine-1-phosphate-induced signaling in vascular endothelial cell migration and vascular stabilization

Sphingosine-1-phosphate (S1P), a platelet-derived bioactive lipid, is a potent regulator of angiogenesis. However, the molecular mechanisms involved in S1P-induced angiogenic responses are not defined. Therefore we investigated the role of S1P-induced signaling in two critical steps of angiogenesis, endothelial cell (EC) migration and subsequent vascular stabilization by investment of mural cells. S1P-induced EC migration is mediated through S1P1 and S1P3 as respective antisense oligonucleotides potently inhibited the response. Both receptors activate Rho and Rac small GTPases. Inhibition of Rho by C3 blocks S1P-induced EC attachment, spreading, and migration, suggesting S1P signaling via Rho is important for cell migration. S1P induced Rho-dependent integrin αvβ3 and β1 activation, leading to effective cell migration and morphogenesis in 3D-fibrin matrix. Upon formation of new blood vessels, the integrity and stability of vasculature develops by the support and interaction with mural cells. Destabilization of mature vessels is often observed in cancer and vascular diseases. Therefore, we investigated the molecular basis of S1P-induced vascular stabilization. S1P activation of SIP1 induced activation and cell-surface trafficking of N-cadherin and it was required for proper EC-mural cell interaction. The activation of S1P1/Gi/Rac axis induced rapid polymerization of microtubules which delivers N-cadherin to polarized microdomains on the apical surface of EC. The strength of N-cadherin-dependent adhesion is further enhanced by regulated phosphorylation of p120ctn and N-cadherin by S1P1 signaling in EC. Furthermore N-cadherin forms an intracellular protein complex including catenins, focal adhesion kinase (FAK), paxillin, cortactin and β3 integrin. Perturbation of N-cadherin expression with small interfering RNA attenuated vascular stabilization in vitro and in vivo. The mechanisms uncovered in the current study may be useful to control angiogenesis and vessel stability during the disease progression and the development of novel approaches in therapeutic angiogenesis.