The Induction of Proliferative Vascular Smooth Muscle Cell Phenotypes by Benzo(a)pyrene Is Characterized by Up-Regulation of Inositol Phospholipid Metabolism and c-Ha-ras Gene Expression

Abstract

Polycyclic aromatic hydrocarbons, such as benzo(a)pyrene (BaP), have been implicated in the initiation and progression of vascular lesions of atherosclerotic morphology in laboratory animals. In the present study, experiments were conducted to define the impact of repeated cycles of BaP challengein vivoon the phenotypic expression of rat vascular (aortic) smooth muscle cells (SMCs). After eight weekly injections of male Sprague–Dawley rats with BaP (10 mg/kg) or vehicle, SMCs were isolated from the thoracic aorta and established in culture to define patterns of phenotypic expression. The mitogenic responsiveness of five independent strains of BaP cells was markedly enhanced relative to control counterparts. Increased proliferation rates in BaP cells correlated with increased synthesis and secretion of proline-containing proteins. Upregulation of mitogenicity in BaP cells did not involve autocrine factors since conditioned media from growth-arrested, as well as cycling cultures of, BaP cells stimulated DNA synthesis to a lesser extent than conditioned media from control counterparts. Levels of inositol phosphate metabolites, primarily products of phosphatidylinositol (PtdIns)-3-kinase activity, were increased relative to controls in cultures of BaP cells under constitutive and serum-stimulated conditions. LY294002, an inhibitor of PtdIns-3-kinase, inhibited DNA synthesis and cell proliferation in control cells to a greater extent than in BaP counterparts. The proliferative phenotype of BaP cells was also characterized by marked up-regulation of c-Ha-ras protooncogene expression. Based on these data we conclude that repeated cycles of BaP challengein vivoinduce highly proliferative vascular smooth muscle cell phenotypes characterized by up-regulation of PtdIns metabolism and c-Ha-ras gene expression.