Objectives: 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) α-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 α-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.
Conclusion: CC2238 α-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 α-ANP variant to enhanced susceptibility to vascular damage in vivo.
aDepartment of Cytology and Histology, IInd School of Medicine, University of Rome La Sapienza, Ospedale S. Andrea, Rome, Italy
bIRCCS Neuromed, Polo Molisano University of Rome La Sapienza, Pozzilli (Is), Italy
cDepartment of Cardiology, IInd School of Medicine, University of Rome La Sapienza, Ospedale S. Andrea, Italy
dSan Pietro Fatebenefratelli Hospital Research Center, Rome, Italy
Received 12 January, 2009
Revised 23 March, 2009
Accepted 20 April, 2009
Correspondence to Speranza Rubattu, MD, Department of Cardiology, IInd School of Medicine, University of La Sapienza, Rome, Italy Tel: +39 06 33775561; fax: +39 06 33775061; e-mail: firstname.lastname@example.org