title: Activation of c-Ha-ras by benzo(a)pyrene in vascular smooth muscle cells involves redox stress and aryl hydrocarbon receptor.
abstract: Repeated cycles of vascular injury by benzo(a)pyrene (BaP) increase the onset and progression of atherosclerotic lesions in laboratory animals. This atherogenic response is partly mediated by activation of cis-acting antioxidant/electrophile response elements that enhance c-Ha-ras transcription in vascular smooth muscle cells (vSMCs). Activation of antioxidant/electrophile responsive cis-acting elements may depend on metabolism of BaP by cytochrome P450s to intermediates that induce oxidative stress and modulate gene expression. To test this hypothesis, we evaluated mitogen-activated c-Ha-ras expression in vSMCs treated with BaP or its metabolic intermediates alone, and in combination with agents that modulate cellular redox status. BaP (0.3 and 3 microM), BaP-3, 6-quinone (0.3 microM), or hydrogen peroxide (50 microM) enhanced serum-activated c-Ha-ras. Ellipticine (0.01 nM), a known inhibitor of cytochrome P450 metabolism and aryl hydrocarbon receptor (AhR) antagonist, inhibited c-Ha-ras induction by BaP (3 microM). Serum challenge of G(0) synchronized cultures of vSMCs with DL-buthionine-(S,R)-sulfoximine (0.1 mM), a depletor of cellular glutathione, increased c-Ha-ras mRNA levels during the early phase of the mitogenic response. Combined BaP/DL-buthionine-(S, R)-sulfoximine challenge was cytotoxic to the cells and inhibited c-Ha-ras expression, whereas up-regulation of antioxidant capacity by N-acetylcysteine (0.5 mM) precluded BaP-induced ras expression. BaP increased formation of reactive oxygen species and depleted cellular glutathione, but these changes did not correlate with the kinetics of c-Ha-ras induction. BaP did not enhance c-Ha-ras expression in vSMCs from AhR knockout mice, although aryl hydrocarbon hydroxylase activity was constitutively expressed in these cells. These results suggest that c-Ha-ras activation in vSMCs by BaP involves a redox-sensitive mechanism that is coupled to AhR receptor-dependent functions.
authors.affiliation: Department of Physiology and Pharmacology, College of Veterinary Medicine, Texas A&M University, College Station 77843-4466, USA.
authors.forename: J K
authors.initials: JK
authors.lastname: Kerzee
authors.forename: K S
authors.initials: KS
authors.lastname: Ramos
mesh_terms: D000818:Animals; D001564:Benzo(a)pyrene / Q000494:pharmacology*; D002478:Cells, Cultured; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects*; D011905:Genes, ras; D051379:Mice; D008810:Mice, Inbred C57BL; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000378:metabolism; D010084:Oxidation-Reduction / Q000187:drug effects; D018336:Receptors, Aryl Hydrocarbon / Q000187:drug effects / Q000378:metabolism*
chemical_list: D018336:Receptors, Aryl Hydrocarbon; D001564:Benzo(a)pyrene