title: Regulation of fibroblast growth factor-2 expression in pulmonary arterial smooth muscle cells involves increased reactive oxygen species generation.
abstract: We have previously demonstrated increased fibroblast growth factor-2 (FGF-2) expression in a lamb model of increased pulmonary blood flow secondary to congenital heart disease, which may contribute to the associated increases in pulmonary arterial muscularization. However, the mechanisms underlying these increases in FGF-2 expression remain to be identified. Initially, we found that exogenous FGF-2 increased endogenous FGF-2 promoter activity and protein levels in ovine pulmonary arterial smooth muscle cells (PASMC). Furthermore, we found that these increases in FGF-2 expression were mediated by increases in superoxide levels via NADPH oxidase activation. In addition, FGF-2-mediated increases in FGF-2 expression and PASMC proliferation were attenuated by inhibition of phosphatidylinositol 3-kinase, Akt, and NADPH oxidase. Increases in FGF-2 expression could be stimulated by other factors known to increase reactive oxygen species (ROS) signaling in PASMC (endothelin-1 and transforming growth factor-beta1), whereas antioxidants attenuated these increases. Deletion constructs localized the growth factor- and ROS-sensitive region within the proximal 103 bp of the FGF-2 promoter, and sequence analysis identified a putative hypoxia response element (HRE), a DNA binding site for the ROS-sensitive transcription factor hypoxia-inducible factor-1alpha (HIF-1alpha). Stabilization of HIF-1alpha increased FGF-2 promoter activity, whereas mutation of the putative HRE attenuated FGF-2-induced FGF-2 promoter activity. Furthermore, FGF-2 increased HIF-1alpha protein levels and consensus HRE promoter activity in PASMC via antioxidant-sensitive mechanisms. Thus we conclude that FGF-2 can stimulate its own expression in PASMC via NADPH oxidase-mediated activation of ROS-sensitive transcription factors, including HIF-1alpha. This positive feedback mechanism may contribute to pulmonary vascular remodeling associated with increased pulmonary blood flow.
authors.affiliation: Vascular Biology Center, Medical College of Georgia, Augusta, Georgia, USA.
authors.forename: Stephen M
authors.initials: SM
authors.lastname: Black
authors.forename: Jennifer M
authors.initials: JM
authors.lastname: DeVol
authors.forename: Stephen
authors.initials: S
authors.lastname: Wedgwood
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D000975:Antioxidants / Q000494:pharmacology; D019898:Autocrine Communication; D049109:Cell Proliferation; D004305:Dose-Response Relationship, Drug; D019332:Endothelin-1 / Q000378:metabolism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D009154:Mutation; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D051058:Oncogene Protein v-akt / Q000037:antagonists & inhibitors / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011401:Promoter Regions, Genetic; D011651:Pulmonary Artery / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D020218:Response Elements; D012756:Sheep; D013997:Time Factors; D014162:Transfection; D053773:Transforming Growth Factor beta1 / Q000378:metabolism; D015854:Up-Regulation
chemical_list: D000975:Antioxidants; D019332:Endothelin-1; D004791:Enzyme Inhibitors; D016222:Fibroblast Growth Factor 2; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D019255:NADPH Oxidases; D051058:Oncogene Protein v-akt; D019869:Phosphatidylinositol 3-Kinases; D000081082:Phosphoinositide-3 Kinase Inhibitors; D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D053773:Transforming Growth Factor beta1