Role of S-1-P receptors and human vascular smooth muscle cell migration in diabetes and metabolic syndrome

Background: Sphingosine-1-phosphate (S-1-P) is a bioactive sphingolipid released from activated platelets that stimulates migration of vascular smooth muscle cells (VSMC) in vitro. S-1-P is associated with oxidized low-density lipoprotein (oxLDL) and is important in vessel remodeling. S-1-P will activate multiple G protein-coupled receptors (S-1-PR 1 to 5), which can regulate multiple cellular functions, including cell migration. The aim of this study is to examine the role of S-1-PR signaling during smooth muscle cell migration in response to S-1-P. Methods: Human VSMCs were cultured in vitro. Expression of S-1-PR 1 to 5 was determined in conditions mirroring diabetes (40 mM glucose) and metabolic syndrome (25 mM glucose with 20 μM linoleic acid and 20 μM oleic acid). Linear wound and Boyden microchemotaxis assays of migration were performed in the presence of S-1-P with and without siRNA against S-1-PR 1 to 5. Assays were performed for activation of ERK1/2, p38^MAPK and JNK. Results: Human VSMCs express S-1-PR1, S-1-PR2, and S-1-PR3. There was no significant expression of S-1-PR4 and S-1-PR5. The expression of S-1-PR1 and S-1-PR3 is enhanced under high glucose conditions and metabolic syndrome conditions. Migration of VSMC in response to S-1-P is enhanced 2-fold by diabetes and 4-fold by metabolic syndrome. In diabetes, S-1-PR1 expression is enhanced, while S-1-PR2 and S-1-PR3 expression are both maintained. In metabolic syndrome, S-1-PR1 and 3 expressions are enhanced and that of S-1-PR2 is reduced. siRNA to S-1-PR1 results in a 2-fold reduction in S-1-P-mediated cell migration under all conditions. siRNA to S-1-PR2 enhanced cell migration only under normal conditions, while siRNA S-1-PR3 decreased migration in metabolic syndrome only. Down-regulation of S-1-PR1 reduced ERK1/2 activation in response to S-1-P, while that of S-1-PR2 had no effect under normal conditions. In diabetes, down-regulation of S-1-PR1 reduced activation of all three MAPKs. In metabolic syndrome, down-regulation of S-1-PR1 and S-1-PR3 reduced activation of all three MAPKs. Conclusion: S-1-PR 1, 2, and 3 regulate human VSMC migration and their expression level and function are modulated by conditions simulating diabetes and metabolic syndrome.