Vascular endothelial dysfunction with aging: endothelin-1 and endothelial nitric oxide synthase

Abstract

To determine whether impaired endothelium-dependent dilation (EDD) in older adults is associated with changes in the expression of major vasoconstrictor or vasodilator proteins in the vascular endothelium, endothelial cells (EC) were obtained from the brachial artery and peripheral veins of 56 healthy men, aged 18-78 yr. Brachial artery EC endothelin-1 (ET-1) [0.99 +/- 0.10 vs. 0.57 +/- 0.10 ET-1/human umbilical vein EC (HUVEC) intensity, P = 0.01] and serine 1177 phosphorylated endothelial nitric oxide synthase (PeNOS) (0.77 +/- 0.09 vs. 0.44 +/- 0.07 PeNOS/HUVEC intensity, P < 0.05) (quantitative immunofluorescence) were greater, and EDD (peak forearm blood flow to intrabrachial acetylcholine) was lower (10.2 +/- 0.9 vs. 14.7 +/- 1.7 ml.100 ml(-1).min(-1), P < 0.05) in older (n = 18, 62 +/- 1 yr) vs. young (n = 15, 21 +/- 1 yr) healthy men. EDD was inversely related to expression of ET-1 (r = -0.39, P < 0.05). Brachial artery EC eNOS expression did not differ significantly with age, but tended to be greater in the older men (young: 0.23 +/- 0.03 vs. older: 0.33 +/- 0.07 eNOS/HUVEC intensity, P = 0.08). In the sample with venous EC collections, EDD (brachial artery flow-mediated dilation) was lower (3.50 +/- 0.44 vs. 7.68 +/- 0.43%, P < 0.001), EC ET-1 and PeNOS were greater (P < 0.05), and EC eNOS was not different in older (n = 23, 62 +/- 1 yr) vs. young (n = 27, 22 +/- 1 yr) men. EDD was inversely related to venous EC ET-1 (r = -0.37, P < 0.05). ET-1 receptor A inhibition with BQ-123 restored 60% of the age-related impairment in carotid artery dilation to acetylcholine in B6D2F1 mice (5-7 mo, n = 8; 30 mo, n = 11; P < 0.05). ET-1 expression is increased in vascular EC of healthy older men and is related to reduced EDD, whereas ET-1 receptor A signaling tonically suppresses EDD in old mice. Neither eNOS nor PeNOS is reduced with aging. Changes in ET-1 expression and bioactivity, but not eNOS, contribute to vascular endothelial dysfunction with aging.

Fig. 1.

Endothelium-dependent dilation (forearm blood flow to intrabrachial artery infusion of ACh; 1.0, 2.0, 4.0, and 8.0 μg/100 ml forearm tissue; A) and endothelium-independent dilation (forearm blood flow to intrabrachial artery infusion of SNP; 0.5, 1.0, and 2.0 μg/100 ml forearm tissue; B) in young and older men. Values are means ± SE. *P < 0.01 vs. young.

Fig. 2.

Endothelium-dependent dilation [brachial artery flow-mediated dilation (FMD); percent change; A] and endothelium-independent dilation [brachial artery dilation with sublingual nitroglycerin (NTG); percent change; B] in young and older men. Values are means ± SE. *P < 0.001 vs. young.

Fig. 3.

Endothelin-1 (ET-1) (A), eNOS (B), and serine 1177 PeNOS protein expression (C) in endothelial cells obtained from brachial arteries in young and older men. Values are means ± SE. *P < 0.05 vs. young.

Fig. 4.

ET-1 (A), endothelial nitric oxide synthase (eNOS; B), and serine 1177 phosphorylated eNOS (PeNOS) protein expression (C) in endothelial cells obtained from antecubital veins in young and older men. Values are means ± SE. HUVEC, human umbilical vein endothelial cell. *P < 0.05 vs. young.

Fig. 5.

Endothelial-dependent dilation (acetylcholine 1 × 10⁻⁹ to 1 × 10⁻⁴ M) in untreated and BQ-123-treated carotid arteries from old mice (A) (dose-response curve of young mice is provided as a reference), and untreated and BQ-123-treated carotid arteries from young mice (B). C: endothelial-independent dilation (sodium nitroprusside, 1 × 10⁻¹⁰ to 1 × 10⁻⁴ M) of carotid arteries isolated from young and old mice. D: maximal dilation to acetylcholine in untreated and BQ-123-treated carotid arteries from young and old mice. E: sensitivity (IC₅₀) to acetylcholine-mediated dilation in untreated and BQ-123-treated carotid arteries from young and old mice. Values are means ± SE. *P < 0.05 vs. young untreated mice. ‡P < 0.05 vs. untreated old mice. †P < 0.05 vs. young untreated mice.