Role of sphingosine 1-phosphate in the mitogenesis induced by oxidized lowdensity lipoprotein in smooth muscle cells via activation of sphingomyelinase, ceramidase, and sphingosine kinase

Oxidized LDL (oxLDL) have been implicated in diverse biological events lead ing to the development of atherosclerotic lesions. We previously demonstrat ed that the proliferation of cultured vascular smooth muscle cells (SMC) in duced by oxLDL is preceded by an increase in neutral sphingomyelinase activ ity, sphingomyelin turnover to ceramide, and stimulation of mitogen-activat ed protein kinases (Auge, N., Escargueil-Blanc, I., Lajoie-Mazenc, I., Sue, I., Andrieu-Abadie, N., Pieraggi, M. T., Chatelut, M., Thiers, J. C., Jaff rezou, J. P., Laurent, G., Levade, T., Negre-Salvayre, A., and Salvayre, R. (1998) J. Biol. Chem. 273, 12893-12900). Since ceramide can be converted t o other bioactive metabolites, such as the well established mitogen sphingo sine l-phosphate (S1P), we investigated whether additional ceramide metabol ites are involved in the oxLDL-induced SMC proliferation. We report here th at incubation of SMC with oxLDL increased the activities of both acidic and alkaline ceramidases as well as sphingosine kinase, and elevated cellular sphingosine and S1P. Furthermore, the mitogenic effect of oxLDL was inhibit ed by D-erythro-2-(N-myristoylamino)-1-phenyl-1- propanol and N,N-dimethyls phingosine which are inhibitors of ceramidase and sphingosine kinase, respe ctively, These findings suggest that S1P is a key mediator of the mitogenic effect of oxLDL. In agreement with this conclusion, exogenous addition of sphingosine stimulated the proliferation of cultured SMC, and this effect w as abrogated by dimethylsphingosine but not by fumonisin B1, an inhibitor o f the acylation of sphingosine to ceramide. Exogenous S1P also promoted SMC proliferation. Altogether, these results strongly suggest that the mitogen ic effect of oxLDL in SMC involves the combined activation of sphingomyelin ase(s), ceramidase(s), and sphingosine kinase, resulting in the turnover of sphingomyelin to a number of sphingolipid metabolites, of which at least S 1P is critical for mitogenesis.