title: Reactive oxygen species and ERK 1/2 mediate monocyte chemotactic protein-1-stimulated smooth muscle cell migration.
abstract: Monocyte chemotactic protein-1 (MCP-1), a potent chemoattractant for monocytes, is thought to play a major role in atherosclerosis, but whether its atherogenic effects involve the direct modulation of vascular smooth muscle cell (SMC) functions remains unclear. This study examined the effects of MCP-1 on the migration of cultured A7r5 SMCs and the signaling pathways involved. Addition of recombinant MCP-1 stimulated SMC migration in modified Boyden chambers coated with type I collagen in a concentration-dependent manner, with 10(-9) M being maximally effective. Using untreated A7r5 cells, two MCP-1 receptors, CCR2 and CCR4, were detected and MCP-1 secretion was significantly increased by stimulation with platelet-derived growth factor. MCP-1-stimulated A7r5 migration was completely blocked by the NAD(P)H oxidase inhibitor, diphenylene iodonium (DPI), and dose-dependently inhibited by polyethylene glycol-conjugated superoxide dismutase (PEG-SOD), suggesting a role for reactive oxygen species (ROS) in this process. During MCP-1 stimulation, ROS production increased rapidly, then gradually decayed over 60 min, and this effect was markedly decreased by pretreatment with DPI or PEG-SOD. Interestingly, U0126 and PD98059, which inhibit activation of extracellular signal-regulated kinases 1/2 (ERK 1/2), significantly inhibited MCP-1-activated ROS generation. Furthermore, transfection of an active mutant of MEK1 (ERK 1/2 kinase) markedly increased superoxide production in rat aortic smooth muscle cells, as detected by dihydroethydium staining, suggesting that ERK 1/2 activation stimulates ROS generation. ERK 1/2 activation was increased for at least 30 min in cells incubated with MCP-1, and this effect was abolished by U0126 or DPI pretreatment. These results demonstrate that MCP-1 is a chemoattractant for SMCs and that MCP-1-stimulated migration requires both ROS production and ERK 1/2 activation in a positive activation loop, which may contribute to the atherogenic effects of MCP-1.
authors.affiliation: Department of Cell Biology and Anatomy, National Cheng Kung University Medical College, Taiwan.
authors.forename: I-Chung
authors.initials: IC
authors.lastname: Lo
authors.forename: Jun-Ming
authors.initials: JM
authors.lastname: Shih
authors.forename: Meei Jyh
authors.initials: MJ
authors.lastname: Jiang
mesh_terms: D000818:Animals; D001011:Aorta / Q000378:metabolism; D002070:Butadienes / Q000494:pharmacology; D002460:Cell Line; D002465:Cell Movement; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000378:metabolism*; D004396:Coloring Agents / Q000494:pharmacology; D017931:DNA Primers / Q000737:chemistry; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay; D005419:Flavonoids / Q000494:pharmacology; D015151:Immunoblotting; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism*; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000166:cytology*; D009570:Nitriles / Q000494:pharmacology; D009861:Onium Compounds / Q000494:pharmacology; D011092:Polyethylene Glycols / Q000737:chemistry; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D013696:Temperature; D013997:Time Factors; D014162:Transfection
chemical_list: D002070:Butadienes; D018932:Chemokine CCL2; D004396:Coloring Agents; D017931:DNA Primers; D004791:Enzyme Inhibitors; D005419:Flavonoids; D009570:Nitriles; D009861:Onium Compounds; D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; C113580:U 0126; D013481:Superoxides; D011092:Polyethylene Glycols; C007517:diphenyleneiodonium; D013482:Superoxide Dismutase; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one