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). (Hypertension. 2007; 50: 1093- 1098.).