Angiotensin-(1-7) through receptor Mas mediates endothelial nitric oxide synthase activation via Akt-dependent pathways.
Angiotensin-(1-7) [Ang-(1-7)] causes endothelial-dependent vasodilation mediated, in part, by NO release. However, the molecular mechanisms involved in endothelial NO synthase (eNOS) activation by Ang-(1-7) remain unknown. Using Chinese hamster ovary cells stably transfected with Mas cDNA (Chinese hamster ovary-Mas), we evaluated the underlying mechanisms related to receptor Mas-mediated posttranslational eNOS activation and NO release. We further examined the Ang-(1-7) profile of eNOS activation in human aortic endothelial cells, which constitutively express the Mas receptor. Chinese hamster ovary-Mas cells and human aortic endothelial cell were stimulated with Ang-(1-7; 10(-7) mol/L; 1 to 30 minutes) in the absence or presence of A-779 (10(-6) mol/L). Additional experiments were performed in the presence of the phosphatidylinositol 3-kinase inhibitor wortmannin (10(-6) mol/L). Changes in eNOS (at Ser1177/Thr495 residues) and Akt phosphorylation were evaluated by Western blotting. NO release was measured using both the fluorochrome 2,3-diaminonaphthalene and an NO analyzer. Ang-(1-7) significantly stimulated eNOS activation (reciprocal phosphorylation/dephosphorylation at Ser1177/Thr495) and induced a sustained Akt phosphorylation (P<0.05). Concomitantly, a significant increase in NO release was observed (2-fold increase in relation to control). These effects were blocked by A-779. Wortmannin suppressed eNOS activation in both Chinese hamster ovary-Mas and human aortic endothelial cells. Our findings demonstrate that Ang-(1-7), through Mas, stimulates eNOS activation and NO production via Akt-dependent pathways. These novel data highlight the importance of the Ang-(1-7)/Mas axis as a putative regulator of endothelial function.
Sampaio
Walkyria Oliveira
WO
Department of Physiology, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Souza dos Santos
Robson Augusto
RA
Faria-Silva
Raphael
R
da Mata Machado
Leonor Tapias
LT
Schiffrin
Ernesto L
EL
Touyz
Rhian M
RM
D000803:Angiotensin I / Q000494:pharmacology / Q000502:physiology*; D000818:Animals; D001011:Aorta / Q000166:cytology; D016466:CHO Cells / Q000378:metabolism; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D042783:Endothelial Cells / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D006801:Humans; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D010446:Peptide Fragments / Q000494:pharmacology / Q000502:physiology*; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational / Q000502:physiology; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000502:physiology*; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology*; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000502:physiology*; D014162:Transfection
D010446:Peptide Fragments; D000090063:Proto-Oncogene Mas; D011518:Proto-Oncogene Proteins; D043562:Receptors, G-Protein-Coupled; D009569:Nitric Oxide; D000803:Angiotensin I; D052250:Nitric Oxide Synthase Type III; D051057:Proto-Oncogene Proteins c-akt; C118790:angiotensin I (1-7)