Rho-mediated phosphorylation of focal adhesion kinase and myosin light chain in human endothelial cells stimulated with sphingosine 1-phosphate, a bioactive lysophospholipid released from activated platelets

Since sphingosine 1-phosphate (Sph-1-P) is stored in abundant amounts in bl ood platelets and released extracellularly upon stimulation, it is importan t to clarify the effects of this bioactive lysophospholipid on vascular end othelial cells from the viewpoint of platelet-endothelial cell interactions . In this study, we investigated the effects of Sph-1-P on the cytoskeletal remodeling of human umbilical vein endothelial cells (HUVECs). Of a focal adhesion kinase (FAK) family of non-receptor protein-tyrosine kinases, HUVE Cs were found to express FAK, but scarcely proline-rich tyrosine kinase 2. Sph-1-P induced FAK tyrosine phosphorylation, myosin light chain phosphoryl ation, and the formation of stress fibers in HUVECs. The specific Rho inact ivator C3 transferase from Clostridium botulinum abolished all of these cyt oskeletal responses induced by Sph-1-P, while pertussis toxin only partly i nhibited FAK tyrosine phosphorylation, and hardly affected myosin light cha in phosphorylation and stress fiber formation. In contrast, Sph-1-P-induced intracellular Ca2+ mobilization was suppressed by pertussis toxin, but not at all by C3 exoenzyme. Our results suggest that Sph-1-P, a bioactive lipi d released from activated platelets, induces endothelial cell cytoskeletal reorganization, mainly through Rho-mediated signaling pathways.