Reduction of angiotensin A and alamandine vasoactivity in the rabbit model of atherogenesis: differential effects of alamandine and Ang(1-7)

Abstract

Novel treatments are necessary to reduce the burden of cardiovascular disease (CVD). Alamandine binds to MrgD and is reported to induce vasodilation via stimulation of endothelial nitric oxide synthase (eNOS), but its role in atherogenic blood vessels is yet to be determined. To determine the vasoactive role of alamandine and its precursor AngA in diseased aorta, New Zealand White rabbits were fed a diet containing 1% methionine + 0.5% cholesterol + 5% peanut oil for 4 weeks (MC, n = 5) or control (n = 6). In abdominal aorta, alamandine (1 μM) was added 30 min before a dose-response curve to angiotensin II or AngA (1 nM-1 μM), and immunohistochemistry was used to identify MrgD receptors and eNOS. The thoracic aorta, renal, carotid and iliac arteries were mounted in organ baths. Rings were precontracted with phenylephrine, then a bolus dose of alamandine (1 μM) was added 10 min before a dose-response curve to acetylcholine (0.01 μM-10 μM). The MrgD receptor was localized to normal and diseased aorta and colocalized with eNOS. In control but not diseased blood vessels, alamandine enhanced acetylcholine-mediated vasodilation in the thoracic aorta and the iliac artery (P < 0.05) and reduced it in the renal artery (P < 0.05). In control abdominal aorta, AngA evoked less desensitization than AngII (P < 0.05) and alamandine reduced AngA-mediated vasoconstriction (P < 0.05). In MC, AngA constriction was markedly reduced vs. control (P < 0.05). The vasoactivity of alamandine and AngA are reduced in atherogenesis. Its role in the prevention of CVD remains to be validated.

Keywords: alamandine; angiotensin A; atherosclerosis; endothelial dysfunction.

Figure 1

Immunohistochemical localization of MrgD receptors (MrgD R) clearly shows positive immunostaining within atherosclerotic plaques and the endothelial layer. The first row shows 3-μm serially adjacent sections stained for RAM-11 (Macrophage), MrgD R and HHF-35 (smooth muscle cells). Inserted cropped sections show little or no staining for macrophage/MrgD R +/+ cells (arrows), yet HHF-35/MrgD R +/+ cells were abundant (cropped sections in 2nd row attached by dashed line as well as identified by letters A, B and C). In other plaques, cells expressing eNOS also expressed MrgD R (3rd row, letters D, E and F), and the endothelial layer also showed co-expression of both proteins (cropped sections, 3rd row). Small arterioles in the vaso vasorum showed positive endothelial MrgD R (cropped sections, 4th row), and negative controls also shown (−). All sections were photographed at an original 400× magnification.

Figure 2

In control rings, alamandine enhanced endothelium-dependent relaxation to acetylcholine in the (a) thoracic aorta and (b) iliac artery. Alamandine did not affect the carotid artery (c), but reduced endothelium-dependent relaxation to acetylcholine in the renal artery (d). In rings from animals fed the atherogenic diet (MC), alamandine failed to significantly affect endothelium-dependent relaxation to acetylcholine (e–h) (V = vehicle, Alam. = alamandine). *P < 0.05.

Figure 3

In control rings, AngA evoked less desensitization than AngII (a) and almandine reduced AngA-mediated vasoconstriction (b). Neither alamandine nor Ang(1-7) affected AngII-mediated vasoconstriction (c). In MC, AngA vasoconstriction was reduced compared with control (d), and neither almandine nor Ang(1-7) affected this constriction (e). Interestingly, Ang(1-7) but not alamandine increased AngII-mediated vasoconstriction (f). *P < 0.05.