Angiotensin-(1-7) counterregulates angiotensin II signaling in human endothelial cells.
Angiotensin (Ang)-(1-7), acting through the Mas receptor, opposes the actions of Ang II. Molecular mechanisms for this are unclear. Here we sought to determine whether Ang-(1-7) influences Ang II signaling in human endothelial cells, focusing specifically on Src homology 2-containing inositol phosphatase 2 (SHP-2) and its interaction with c-Src. Ang II-induced phosphorylation of c-Src, extracellular signal regulated kinase (ERK)1/2, and SHP-2 and activation of NAD(P)H oxidase were assessed in the absence and presence of Ang-(1-7) (10(-6) mol/L, 15 minutes) by immunoblotting and lucigenin-enhanced chemiluminescence, respectively. (D-Ala(7))-Ang I/II (1-7) (Ang fragment 1-7 receptor antagonist) was used to block Ang-(1-7) effects. Association between SHP-2 and c-Src was assessed by immunoprecipitation/immunoblotting studies. Ang II significantly increased activation of c-Src, ERK1/2, and NAD(P)H oxidase and reduced phosphorylation of SHP-2 (P<0.05) in human endothelial cells. These effects were abrogated in cells pre-exposed to Ang-(1-7). Ang fragment 1-7 receptor antagonist pretreatment blocked the negative modulatory actions of Ang-(1-7) on Ang II-induced signaling. Ang-(1-7) alone did not significantly alter phosphorylation of c-Src, ERK1/2, and SHP-2 and had no effect on basal activity of NAD(P)H oxidase. SHP-2 and c-Src were physically associated in the basal state. This association was increased by Ang-(1-7) and blocked by Ang fragment 1-7 receptor antagonist. Our findings demonstrate that, in human endothelial cells, Ang-(1-7) negatively modulates Ang II/Ang II type 1 receptor-activated c-Src and its downstream targets ERK1/2 and NAD(P)H oxidase. We also show that SHP-2-c-Src interaction is enhanced by Ang-(1-7). These phenomena may represent a protective mechanism in the endothelium whereby potentially deleterious effects of Ang II are counterregulated by Ang-(1-7).
Sampaio
Walkyria O
WO
Department of Physiology, Federal University of Minas Gerais, Belo Horizonte, Minas Gerais, Brazil.
Henrique de Castro
Carlos
C
Santos
Robson A S
RA
Schiffrin
Ernesto L
EL
Touyz
Rhian M
RM
D000803:Angiotensin I / Q000494:pharmacology*; D000804:Angiotensin II / Q000031:analogs & derivatives / Q000037:antagonists & inhibitors* / Q000494:pharmacology; D000081247:CSK Tyrosine-Protein Kinase; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006801:Humans; D019255:NADPH Oxidases / Q000378:metabolism; D010446:Peptide Fragments / Q000494:pharmacology*; D010766:Phosphorylation; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11 / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D019061:src-Family Kinases
C092006:7-Ala-angiotensin (1-7); D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D000804:Angiotensin II; D000803:Angiotensin I; D019255:NADPH Oxidases; D011505:Protein-Tyrosine Kinases; D000081247:CSK Tyrosine-Protein Kinase; D019061:src-Family Kinases; C000631346:CSK protein, human; D048049:Extracellular Signal-Regulated MAP Kinases; C516724:PTPN11 protein, human; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; C118790:angiotensin I (1-7)