Mitogenic effect of oxidized low-density lipoprotein on vascular smooth muscle cells mediated by activation of Ras/Raf/MEK/MAPK pathway.
1. 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. 2. Both native-LDL (N-LDL) and OX-LDL induced a time- and concentration-dependent incorporation of [(3)H]-thymidine in VSMCs. 3. OX-LDL induced time- and concentration-dependent phosphorylation of p42/p44 MAPK. Pretreatment of these cells with pertussis toxin or U73122 attenuated the OX-LDL-induced responses. 4. 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 [(3)H]-thymidine incorporation and p42/p44 MAPK phosphorylation induced by OX-LDL. 5. Removal of Ca(2+) by BAPTA/AM or depletion of the internal Ca(2+) pool by thapsigargin significantly inhibited OX-LDL-induced [(3)H]-thymidine incorporation and p42/p44 MAPK phosphorylation. 6. OX-LDL-induced [(3)H]-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. 7. 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. 8. 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.
Cellular and Molecular Pharmacology Laboratory, Department of Physiology and Pharmacology, College of Medicine, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan. chuenmao@mail.cgu.tw
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Yang
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Chien
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Hsiao
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Pan
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Wang
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Chiu
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Lin
D000818:Animals; D016227:Benzoquinones; D002118:Calcium / Q000494:pharmacology; D004247:DNA / Q000096:biosynthesis / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D019833:Genistein / Q000494:pharmacology; D006801:Humans; D047029:Lactams, Macrocyclic; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D020935:MAP Kinase Signaling System / Q000187:drug effects; D019950:Mitogen-Activated Protein Kinase 1 / Q000187:drug effects / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000187:drug effects* / Q000378:metabolism; D008934:Mitogens / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D037342:Pertussis Toxin; D010766:Phosphorylation / Q000187:drug effects; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D011760:Pyrrolidinones / Q000494:pharmacology; D011809:Quinones / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D017828:Rifabutin / Q000031:analogs & derivatives; D019311:Staurosporine / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology; D018631:ras Proteins / Q000378:metabolism
D016227:Benzoquinones; D004791:Enzyme Inhibitors; D004963:Estrenes; D005419:Flavonoids; D047029:Lactams, Macrocyclic; D008077:Lipoproteins, LDL; D008934:Mitogens; D011760:Pyrrolidinones; D011809:Quinones; D010566:Virulence Factors, Bordetella; C099252:oxidized low density lipoprotein; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D017828:Rifabutin; C020754:herbimycin; D004247:DNA; D019833:Genistein; D037342:Pertussis Toxin; D011505:Protein-Tyrosine Kinases; D019908:Proto-Oncogene Proteins c-raf; D011493:Protein Kinase C; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D018631:ras Proteins; D019311:Staurosporine; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; D002118:Calcium