title: Reactive oxygen species-mediated effects on vascular remodeling induced by human atrial natriuretic peptide T2238C molecular variant in endothelial cells in vitro.
abstract: OBJECTIVE
  T2238C ANP (atrial natriuretic peptide) gene variant has been associated with increased cardiovascular risk in humans and with a significant pharmacogenomic effect on cardiovascular disease outcome in hypertensive patients. We investigated the impact of T2238C ANP gene variant on oxidative stress production, cell proliferation and migration, angiogenesis and vascular remodeling in human umbilical vein endothelial cells in vitro.
  
  
  METHODS
  Differentially expressed genes in human umbilical vein endothelial cells exposed to either wild-type (TT2238) or mutant (CC2238) alpha-ANP were characterized by real time-PCR-macroarray analysis using human oxidative stress, angiogenesis and matrix arrays. Reactive oxygen species (ROS) production was determined by dihydroethidium and by evaluation of dichlorofluorescein content. NADPH oxidase gp91phox subunit was investigated by western blotting. Endothelial cell proliferation, migration and tube formation were characterized both in the presence and in the absence of NADPH oxidase inhibition.
  
  
  RESULTS
  Compared with TT2238, CC2238 alpha-ANP altered the redox state balance of the cells in a more significant manner, favoring ROS production and reducing antioxidative stress response. Gene expression of molecules involved in atherogenesis and vascular remodeling was enhanced. In contrast to TT2238 peptide, CC2238 was unable to stimulate cell proliferation and it markedly inhibited endothelial cell tube formation. NADPH oxidase inhibition restored the cell proliferative properties under CC2238 peptide exposure.
  
  
  CONCLUSIONS
  CC2238 alpha-ANP led to ROS accumulation and increased expression of genes related to atherosclerosis and vascular remodeling in human umbilical vein endothelial cells. As a consequence of NADPH-derived ROS, blunted endothelial cell proliferation and impaired endothelial cell tube formation were observed. These in-vitro effects may link the T2238C alpha-ANP variant to enhanced susceptibility to vascular damage in vivo.
authors.affiliation: Department of Cytology and Histology, University of Rome La Sapienza, Ospedale S. Andrea, Italy.
authors.forename: Stefania
authors.initials: S
authors.lastname: Scarpino
authors.forename: Simona
authors.initials: S
authors.lastname: Marchitti
authors.forename: Rosita
authors.initials: R
authors.lastname: Stanzione
authors.forename: Anna
authors.initials: A
authors.lastname: Evangelista
authors.forename: Sara
authors.initials: S
authors.lastname: Di Castro
authors.forename: Carmine
authors.initials: C
authors.lastname: Savoia
authors.forename: Giovanni
authors.initials: G
authors.lastname: Quarta
authors.forename: Sebastiano
authors.initials: S
authors.lastname: Sciarretta
authors.forename: Luigi
authors.initials: L
authors.lastname: Ruco
authors.forename: Massimo
authors.initials: M
authors.lastname: Volpe
authors.forename: Speranza
authors.initials: S
authors.lastname: Rubattu
mesh_terms: D009320:Atrial Natriuretic Factor / Q000235:genetics* / Q000378:metabolism; D015153:Blotting, Western; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism; D042783:Endothelial Cells / Q000502:physiology*; D004996:Ethidium / Q000031:analogs & derivatives; D005452:Fluoresceins; D020869:Gene Expression Profiling; D006801:Humans; D008562:Membrane Glycoproteins / Q000378:metabolism; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases / Q000378:metabolism; D009389:Neovascularization, Pathologic / Q000378:metabolism; D018384:Oxidative Stress; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D005452:Fluoresceins; D008562:Membrane Glycoproteins; D017382:Reactive Oxygen Species; C067883:dihydroethidium; C037631:2',7'-dichlorofluorescein; D009320:Atrial Natriuretic Factor; C056558:CYBB protein, human; D000074662:NADPH Oxidase 2; D019255:NADPH Oxidases; D004996:Ethidium; D006152:Cyclic GMP