Inhibition of rac1 reduces PDGF-induced reactive oxygen species and proliferation in vascular smooth muscle cells.
In vascular smooth muscle cells, reactive oxygen species (ROS) were known to mediate platelet-derived growth factor (PDGF)-induced cell proliferation and NADH/NADPH oxidase is the major source of ROS. NADH/NADPH oxidase is controlled by rac1 in non-phagocytic cells. In this study, we examined whether the inhibition of rac1 by adenoviral-mediated gene transfer of a dominant negative rac1 gene product (Ad.N17rac1) could reduce the proliferation of rat aortic vascular smooth muscle cells (RASMC) stimulated by PDGF via decreasing intracellular ROS. RASMC were stimulated by PDGF (80 ng/mL) with or without N-acetylcysteine 1 mM or infected with 100 mutiplicity of infection of Ad.N17rac1. Intracellular ROS levels were measured at 12 hr using carboxyl-2', 7'-dichlorodihydrofluorescein diacetate confocal microscopy. At 72 hr, cellular proliferation was evaluated by cell number counting and XTT assay. Compared with control, ROS levels were increased by 2-folds by PDGF. NAC and Ad.N17rac1 inhibited PDGF-induced increase of ROS by 77% and 65%, respectively. Cell number was increased by PDGF by 1.6-folds compared with control. NAC and Ad.N17rac1 inhibited PDGF-induced cellular growth by 45% and 87%, respectively. XTT assay also showed similar results. We concluded that inhibition of rac1 in RASMCs could reduce intracellular ROS levels and cellular proliferation induced by PDGF.
Kong
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Cardiology Division, College of Medicine, Hanyang University, Seoul, Korea. kskim@hanyang.ac.kr
Lee
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Kim
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KS
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D001013:Aorta, Thoracic / Q000166:cytology; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D015870:Gene Expression / Q000502:physiology; D018014:Gene Transfer Techniques; D009097:Multienzyme Complexes / Q000037:antagonists & inhibitors; D009131:Muscle, Smooth, Vascular / Q000166:cytology* / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000037:antagonists & inhibitors; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000235:genetics* / Q000378:metabolism
D009097:Multienzyme Complexes; D010982:Platelet-Derived Growth Factor; D017382:Reactive Oxygen Species; C023107:NADH oxidase; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D020830:rac1 GTP-Binding Protein