Pro-Inflammatory Role of S1P₃ in Macrophages

Sphingosine kinase 1 and its product, sphingosine 1-phosphate (S1P), as well as their receptors, have been implicated in inflammatory responses. The functions of receptors S1P₁ and S1P₂ on cell motility have been investigated. However, the function of S1P₃ has been poorly investigated. In this study, the roles of S1P₃ on inflammatory response were investigated in primary perito-neal macrophages. S1P₃ receptor was induced along with sphingosine kinase 1 by stimulation of lipopolysaccharide (LPS). LPS treatment induced inflammatory genes, such iNOS, COX-2, IL-1β, IL-6 and TNF-α. TY52156, an antagonist of S1P₃ suppressed the induction of inflammatory genes in a concentration dependent manner. Suppression of iNOS and COX-2 induction was further confirmed by western blotting and NO measurement. Suppression of IL-1β induction was also confirmed by western blotting and ELISA. Caspase 1, which is responsible for IL-1β production, was similarly induced by LPS and suppressed by TY52156. Therefore, we have shown S1P₃ induction in the inflammatory conditions and its pro-inflammatory roles. Targeting S1P₃ might be a strategy for regulating inflammatory diseases.

Analysis of a Sphingosine 1-phosphate Receptor hS1P3 in Rat Hepatoma Cells

To examine intracellular signaling of human S1P3 (hS1P3), a sphingosine 1-phosphate (S1P) receptor in plasma membrane, hS1P3 DNA was transfected into RH7777 rat hepatoma cell line, and the inhibition of forskolin-induced cAMP accumulation and activation of MAP kinases by S1P were tested. In hS1P3 transformants, S1P inhibited forskolin-induced activation of adenylyl cyclase activity by about 80% and activated MAP kinases in dose-dependent and pertussis-toxin (PTX) sensitive manners. In oocytes expressing hS1P3 receptor, S1P evoked Cl conductance. These data suggested that PTX-sensitive G proteins are involved in hS1P3-mediated signaling, especially the positive action of S1P in cell proliferation. The potential advantages of rat hepatoma cells for the research of sphingosine 1-phosphate receptor are discussed.