Sphingosine 1-phosphate mediates chemotaxis of human primary fibroblasts via the S1P-receptor subtypes S1P₁ and S1P₃ and Smad-signalling.

The sphingolipid sphingosine 1-phosphate (S1P) induces chemotaxis of primary fibroblasts. Thus, S1P exhibited a chemotactic effect in a concentration-dependent manner from 10⁻⁶ to 10⁻⁹ M; higher concentrations resulted in a loss of migration, and lower amounts were ineffective to evoke movement toward a concentration gradient of S1P. In congruence with the migratory response, S1P caused an extension of lamellipodia at the cell periphery of human fibroblasts and a rearrangement of the cytoskeleton. These effects were visible by phalloidin staining of actin filaments as well as focal adhesion turnover. As the molecular mechanism of S1P-mediated migration of fibroblasts has not been well characterized, we investigated whether S1P-receptors are involved in the chemotactic response. Indeed, inhibition of G(i) signalling markedly reduced motility towards S1P, suggesting an involvement of S1P-receptor subtypes. Moreover, downregulation of S1P₁ and S1P₃ indicated that these S1P-receptor subtypes are responsible for the chemotactic action of the bioactive sphingolipid. After having identified a crosstalk between Smad-proteins and S1P-signalling, we investigated whether Smad-activation is involved in the chemotactic response induced by S1P. Indeed S1P caused a Smad-activation via the S1P receptor subtypes S1P₁ and S1P₃. Moreover, downregulation of Smad3 diminished the ability of S1P to mediate a chemotactic response in fibroblasts, indicating a crosstalk between TGF-β- and S1P-signalling.