Mitogenic effect of oxidized low-density lipoprotein on vascular smooth muscle cells mediated by activation of Ras/Raf/MEK/MAPK pathway

摘要

class="enumerated" style="list-style-type:decimal">It has been demonstrated that oxidized low-density lipoprotein (OX-LDL) is a risk factor in atherosclerosis by stimulating vascular smooth muscle cell (VSMC) proliferation. However, the mechanisms of OX-LDL-induced cell proliferation are not completely understood. Therefore, we investigated the effect of OX-LDL on cell proliferation associated with mitogen-activated protein kinase (MAPK) activation in rat cultured VSMCs.Both native-LDL (N-LDL) and OX-LDL induced a time- and concentration-dependent incorporation of [³H]-thymidine in VSMCs.OX-LDL induced time- and concentration-dependent phosphorylation of p42/p44 MAPK. Pretreatment of these cells with pertussis toxin or attenuated the OX-LDL-induced responses.Pretreatment with PMA for 24 h, preincubation with a PKC inhibitor staurosporine or the tyrosine kinase inhibitors, genistein and herbimycin A for 1 h, substantially reduced [³H]-thymidine incorporation and p42/p44 MAPK phosphorylation induced by OX-LDL.Removal of Ca²⁺ by BAPTA/AM or depletion of the internal Ca²⁺ pool by thapsigargin significantly inhibited OX-LDL-induced [³H]-thymidine incorporation and p42/p44 MAPK phosphorylation.OX-LDL-induced [³H]-thymidine incorporation and p42/p44 MAPK phosphorylation was inhibited by PD98059 (an inhibitor of MEK1/2) and SB203580 (an inhibitor of p38 MAPK) in a concentration-dependent manner.Overexpression of dominant negative mutants of Ras (H-Ras-15A) and Raf (Raf-N4) significantly suppressed MEK1/2 and p42/p44 MAPK activation induced by OX-LDL and PDGF-BB, indicating that Ras and Raf may be required for activation of these kinases.These results suggest that the mitogenic effect of OX-LDL is mediated through a PTX-sensitive G protein-coupled receptor that involves the activation of the Ras/Raf/MEK/MAPK pathway similar to that of PDGF-BB in rat cultured VSMCs.