POTENTIAL ROLE FOR CERAMIDE IN MITOGEN-ACTIVATED PROTEIN-KINASE ACTIVATION AND PROLIFERATION OF VASCULAR SMOOTH-MUSCLE CELLS INDUCED BY OXIDIZED LOW-DENSITY-LIPOPROTEIN

Proliferation of vascular smooth muscle cells (SMC) is a hallmark in t he pathogenesis of atherosclerotic lesions. Mildly oxidized low densit y lipoproteins (UV-oxLDL), which are mitogenic to cultured AG-08133A S MC, activate the sphingomyelin (SM)-ceramide pathway. We report here t he following. (i) UV-oxLDL elicited a biphasic and sustained activatio n of MBP kinase activity, phosphorylation and nuclear translocation of p44/42 mitogen-activated protein kinase (MAPK), and [H-3]thymidine in corporation, which were inhibited by PD-098059, a MAPK kinase inhibito r. (ii) The use of preconditioned media (from SMC pre-activated by UV- oxLDL) transferred to native SMC and blocking antibodies against growt h factors suggest that UV-oxLDL-induced activation of MAPK and [H-3]th ymidine incorporation seem to be independent of any autocrine secretio n of growth factors. (iii) UV-oxLDL-induced activation of a neutral sp hingomyelinase, SM hydrolysis, ceramide production, and [H-3]thymidine incorporation were inhibited by two serine-protease inhibitors (serpi ns), suggesting that a serpin-sensitive proteolytic pathway is involve d in the activation of the SM-ceramide signaling pathway. (iv) UV-oxLD L-induced MAPK activation and [H-3]thymidine incorporation were mimick ed by ceramide generated in the plasma membrane by bacterial sphingomy elinase treatment or by addition of the permeant C-2-ceramide. Serpins did not inhibit the MAPK activation and [H-3]thymidine incorporation induced by C-2-ceramide, indicating that activation of the MAPK and [H -3]thymidine incorporation is subsequent to the stimulation of the SM- ceramide pathway. Taken together, these data suggest that mitogenic co ncentrations of UV-oxLDL are able to stimulate the SM-ceramide pathway through a protease-dependent mechanism and activate p44/42 MAPK, lead ing to proliferation of vascular SMC.