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Angiotensin-converting-enzyme inhibition counteracts angiotensin II-mediated endothelial cell dysfunction by modulating the p38/SirT1 axis

Marampon, Francescoa,b,*; Gravina, Giovanni L.a,*; Scarsella, Lucaa; Festuccia, Claudioa; Lovat, Francescac; Ciccarelli, Carmelaa; Zani, Bianca M.a; Polidoro, Lorellab; Grassi, Davideb; Desideri, Giovambattistab; Evangelista, Stefanod; Ferri, Claudiob

doi: 10.1097/HJH.0b013e3283638b32

Objective: Oxidative stress has been linked to endothelial dysfunction and angiotensin II stimulates the reactive oxygen species production contributing to several cardiovascular diseases. We have studied the chain of events induced by angiotensin-converting-enzyme (ACE) activation in vascular umbilical vein endothelial cells (HUVECs) by using an ACE inhibitor such as zofenoprilat.

Methods: We used specific assay to measure the superoxide anion production, tetrazolium bromide (MTT) assay for cell viability, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay for cell apoptosis, and western blot for protein analysis in the study.

Results: Zofenoprilat counteracts the superoxide anion production and cell apoptosis induced by angiotensin I treatment by blocking the extrinsic caspase cascade, NF-kB and p38 activation. p38 inhibitor SB203580 reverted the angiotensin II oxidant effects while the p38 constitutively activation, by MKK6 transfection, abrogated the zofenoprilat effects. Characterizing the zofenoprilat downstream effector we found that zofenoprilat reverted the SirT-1 downregulation induced by angiotensin II. p38 activation by angiotensin II was strictly correlated with SirT1 protein downregulation; SB203580 significantly prevented SirT1 downregulation induced by angiotensin II while the p38 constitutive activation abolished SIRT1 protein basal levels. p38 directly bound SirT1 sequestering it in the cytoplasm. SirT1 inhibition by sirtinol annulled zofenoprilat action while SirT1 overexpression reverted the cytotoxic effects of angiotensin II. Finally, zofenoprilat negatively controlled angiotensin I receptor protein expression through SirT1.

Conclusion: The p38-SirT1 axis is found markedly relevant in modulating the cardiovascular benefit deriving from ACE-inhibitors and might represent a novel target for innovative drugs in cardiovascular prevention.

aDepartment of Applied Clinical Sciences and Biotechnology, University of L’Aquila

bDepartment MeSVA, University of L’Aquila, L’Aquila, Italy

cDepartment of Molecular Immunology, Virology, and Medical Genetics, The Ohio State University, Columbus, Ohio, USA

dPreclinical Development Department, Menarini Ricerche, Firenze, Italy

*Francesco Marampon and Giovanni L. Gravina contributed equally to the writing of this article.

Correspondence to Francesco Marampon, Department of Applied Clinical Sciences and Biotechnology, University of L’Aquila, Via Vetoio, Coppito 2, L’Aquila, Italy. E-mail:

Received 14 February, 2013

Revised 24 April, 2013

Accepted 23 May, 2013

© 2013 Wolters Kluwer Health | Lippincott Williams & Wilkins