The antithrombotic effect of angiotensin-(1-7) involves mas-mediated NO release from platelets

Mol Med. 2008 Jan-Feb;14(1-2):28-35. doi: 10.2119/2007-00073.Fraga-Silva.

Abstract

The antithrombotic effect of angiotensin(Ang)-(1-7) has been reported, but the mechanism of this effect is not known. We investigated the participation of platelets and receptor Mas-related mechanisms in this action. We used Western blotting to test for the presence of Mas protein in rat platelets and used fluorescent-labeled FAM-Ang-(1-7) to determine the specific binding for Ang-(1-7) and its displacement by the receptor Mas antagonist A-779 in rat platelets and in Mas(-/ -) and Mas(+/+) mice platelets. To test whether Ang-(1-7) induces NO release from platelets, we used the NO indicator DAF-FM. In addition we examined the role of Mas in the Ang-(1-7) antithrombotic effect on induced thrombi in the vena cava of male Mas(-/ -) and Mas(+/+) mice. The functional relevance of Mas in hemostasis was evaluated by determining bleeding time in Mas(+/+) and Mas(-/ -) mice. We observed the presence of Mas protein in platelets, as indicated by Western Blot, and displacement of the binding of fluorescent Ang-(1-7) to rat platelets by A-779. Furthermore, in Mas(+/+) mouse platelets we found specific binding for Ang-(1-7), which was absent in Mas(-/ -) mouse platelets. Ang-(1-7) released NO from rat and Mas(+/+) mouse platelets, and A-779 blocked this effect. The NO release stimulated by Ang-(1-7) was abolished in Mas(-/ -) mouse platelets. Ang-(1-7) inhibited thrombus formation in Mas(+/+) mice. Strikingly, this effect was abolished in Mas(-) (/) (-)mice. Moreover, Mas deficiency resulted in a significant decrease in bleeding time (8.50 +/- 1.47 vs. 4.28 +/- 0.66 min). This study is the first to show the presence of Mas protein and specific binding for Ang-(1-7) in rat and mouse platelets. Our data also suggest that the Ang-(1-7) antithrombotic effect involves Mas-mediated NO release from platelets. More importantly, we showed that the antithrombotic effect of Ang-(1-7) in vivo is Mas dependent and that Mas is functionally important in hemostasis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Angiotensin I / metabolism
  • Angiotensin I / pharmacology*
  • Angiotensin II / analogs & derivatives
  • Angiotensin II / pharmacology
  • Animals
  • Bleeding Time
  • Blood Coagulation / drug effects
  • Blood Platelets / drug effects
  • Blood Platelets / metabolism*
  • Male
  • Mice
  • Mice, Inbred C57BL
  • Mice, Knockout
  • Nitric Oxide / metabolism*
  • Peptide Fragments / metabolism
  • Peptide Fragments / pharmacology*
  • Proto-Oncogene Mas
  • Proto-Oncogene Proteins / deficiency
  • Proto-Oncogene Proteins / drug effects
  • Proto-Oncogene Proteins / metabolism*
  • Rats
  • Receptors, G-Protein-Coupled / deficiency
  • Receptors, G-Protein-Coupled / drug effects
  • Receptors, G-Protein-Coupled / metabolism*
  • Vasoconstrictor Agents / pharmacology
  • Vasodilator Agents / metabolism
  • Vasodilator Agents / pharmacology*

Substances

  • 7-Ala-angiotensin (1-7)
  • Peptide Fragments
  • Proto-Oncogene Mas
  • Proto-Oncogene Proteins
  • Receptors, G-Protein-Coupled
  • Vasoconstrictor Agents
  • Vasodilator Agents
  • Angiotensin II
  • Nitric Oxide
  • Angiotensin I
  • angiotensin I (1-7)