Citations

Endothelium-dependent contractions to acetylcholine and endothelium-independent contractions to oxygen-derived free radicals in the aorta of the spontaneously hypertensive rat (SHR) are mediated by an unidentified product of the cyclooxygenase pathway of arachidonic acid metabolism. To determine the role of thromboxane A2 (TXA2) or prostaglandin H2 (PGH2) in these contractions, rings of the thoracic aorta of SHR were suspended in organ chambers for measurement of isometric force. Acetylcholine caused endothelium-dependent contractions in quiescent rings from SHR aortas. Oxygen-derived free radicals generated with xanthine plus xanthine oxidase caused contractions in rings without endothelium. Dazoxiben (thromboxane synthetase inhibitor) did not affect contractions evoked by acetylcholine. AH 23,848, SQ 29,548, or R 68,070 (TXA2/PGH2 receptor antagonists) inhibited contractions to U 46,619 (a TXA2/PGH2 receptor agonist), acetylcholine, and oxygen-derived free radicals. Acetylcholine stimulated the release of prostacyclin from Wistar-Kyoto (WKY) rat and SHR aortas but not the release of other prostaglandins (PGE2, PGF2 alpha, TXA2). Oxygen-derived free radicals did not stimulate the release of prostaglandins from either SHR or WKY rat aortas. These results demonstrate that stimulation of TXA2/PGH2 receptors probably by PGH2 might be involved in endothelium-dependent contractions. Oxygen-derived free radicals, which might be an endothelium-derived contracting factor or factors, ultimately cause contraction by stimulation of TXA2/PGH2 receptors.

To determine if oxygen-derived free radicals are mediators of endothelium-dependent contractions to acetylcholine in the aorta of spontaneously hypertensive rats (SHR), the mechanism of contraction to xanthine plus xanthine oxidase was studied. Rings, with and without endothelium, of thoracic aorta from normotensive Wistar-Kyoto (WKY) rats and SHR were suspended in organ chambers for isometric tension recording. Oxygen-derived free radicals caused concentration-dependent contractions; these contractions were twice as large in the aortas of SHR than in WKY rats. Deferoxamine reversed the response to xanthine oxidase to a small relaxation. Either allopurinol, superoxide dismutase, or catalase, or the combination of superoxide dismutase plus catalase reduced the contractions. Diltiazem inhibited the response to xanthine oxidase; in contrast, phentolamine plus propranolol did not affect it. Indomethacin and meclofenamate, but not tranylcypromine or dazoxiben blocked the contractions. Endothelium-dependent contractions to acetylcholine in aortas from the SHR were not affected by deferoxamine or superoxide dismutase plus catalase. These data suggest that hydroxyl radicals cause contractions in the rat aorta, which are dependent on extracellular calcium and mediated by activation of the cyclooxygenase in the vascular smooth muscle. The augmented contractions in the hypertensive strain are due to an increased reactivity of the smooth muscle to oxygen-derived free radicals. However, the lack of effect of the scavengers on endothelium-dependent contractions to acetylcholine suggests that the endothelium-derived contracting factor is chemically different from oxygen-derived free radicals.

The response of isolated blood vessels to a variety of vasoactive agonists is modulated by the presence of endothelial cells. Indeed, these cells can release both dilator and constrictor substances. The major endothelium-derived relaxing factor may be nitric oxide, which activates soluble guanylate cyclase in the smooth muscle, although the endothelial cells also secrete an unidentified hyperpolarizing factor. Among the natural stimuli for the release of endothelium-derived relaxing factors are circulating hormones, platelet products, thrombin, shear stress, and certain autacoids. Endothelium-derived relaxing factors may contribute to the regulation of the release of atrial natriuretic factor and renin. The endothelial cells can also release constricting factors; among the likely candidates are superoxide anions or the peptide endothelin. In hypertensive blood vessels, the ability to release endothelium-derived relaxing factors but not endothelium-derived contracting factors is blunted.

Aggregating platelets release serotonin, which induces contraction of most vascular smooth muscle by activation of S2-serotoninergic receptors. Serotonin released in the circulation may contribute to the increase in peripheral resistance of hypertension as the responsiveness of blood vessels from hypertensive animals and humans to the vasoconstrictor action of the monoamine is augmented. The data obtained with the new antihypertensive agent ketanserin may favor that interpretation. Ketanserin is a selective S2-serotoninergic antagonist with additional alpha 1-adrenergic blocking properties. In humans, it has a terminal half-life of 12 to 25 hours and is eliminated predominantly by the liver. The hemodynamic profile of ketanserin is that of a vasodilator drug with actions on both resistance and capacitance vessels. On short-term intravenous administration, it lowers blood pressure in hypertensive patients with minimal reflex changes in cardiovascular function. When given orally long term to hypertensive patients, ketanserin causes a sustained reduction in arterial blood pressure, comparable to that obtained with either beta-adrenergic blockers or diuretics. Several studies have shown a greater efficacy in older (greater than 60 years of age) than in younger patients independent of starting pressure. Side effects mainly consist of dizziness, somnolence, and dry mouth, but they are usually not severe. The mechanism underlying the antihypertensive effect of ketanserin is unclear. It cannot be attributed to either S2-serotoninergic or alpha 1-adrenergic blockade alone, but an interaction between the two effects appears to be required.

The purposes of the present study were to examine the natural course of the impairment of endothelium-dependent relaxations during a regeneration and tissue repair process after balloon endothelium removal and to elucidate the cellular mechanism(s) underlying it. Twenty-three male Yorkshire pigs underwent balloon endothelium removal along the proximal portion of either the left anterior descending or circumflex coronary artery and were then maintained on a regular chow for 4, 8, 16, or 24 weeks. Endothelium-dependent responses were examined in vitro in rings taken from the control and previously denuded arteries studied in parallel. Morphometric analysis revealed that intimal thickening developed only at the previously denuded area. In the previously denuded arteries with regenerated endothelium, the endothelium-dependent relaxations to UK 14304 (a selective alpha 2-adrenergic agonist), serotonin, and aggregating platelets were impaired 4 weeks after endothelium removal and remained so throughout the study. The endothelium-dependent relaxations to thrombin and adenosine diphosphate became depressed 8 weeks after endothelium removal and those to bradykinin became depressed 16 weeks after endothelium removal, while those to the calcium ionophore A23187 were maintained throughout the study. Endothelium-dependent relaxations to all vasoactive agents were unaltered in the control arteries. In the control arteries, pertussis toxin, an inhibitor of certain G proteins, markedly inhibited the endothelium-dependent relaxations to UK 14304 and serotonin and partially inhibited those to thrombin and aggregating platelets. The responses inhibited by the toxin in control arteries were significantly reduced in the reduced in the previously denuded arteries with regenerated endothelium. The inhibitory effect of pertussis toxin was markedly reduced in those arteries with regenerated endothelium. In quiescent rings, the presence of normal endothelium inhibited the contractions caused by serotonin and aggregating platelets; this endothelium-dependent depression was markedly impaired in the previously denuded arteries throughout the study. Direct relaxation of the coronary smooth muscle to nitric oxide or sodium nitroprusside or direct contraction to KCl or serotonin were comparable between the control and previously denuded arteries. These experiments indicate that endothelium-dependent relaxations progressively worsen after regeneration of the endothelium and that the dysfunction of a pertussis toxin-sensitive G protein partly account for the endothelial dysfunction in the chronic regenerated state.

Experiments were performed to determine the effect of ouabain on the release of relaxing factor(s) from cultured endothelial cells, and its action on the effect of the relaxing factor(s) on arterial smooth muscle. A column of porcine aortic endothelial cells grown on microcarrier beads in suspension culture was perfused with modified Krebs-Ringer bicarbonate solution. The release of relaxing factor(s) by the endothelial cells was detected under bioassay conditions by measuring the relaxing activity of the perfusate overflowing a ring of canine coronary artery (without endothelium) contracted with prostaglandin F2 alpha. Incubation of the endothelial cells with ouabain did not affect the relaxation of the bioassay ring under basal conditions or upon stimulation of the endothelial cells with ADP but impaired the relaxation induced by bradykinin or the calcium ionophore A23187. Incubation of the bioassay ring with ouabain reduced the relaxation under basal conditions as well as the relaxation induced by ADP but did not affect the relaxation observed upon stimulation with bradykinin and A23187 and the endothelium-independent relaxations induced by nitric oxide. These experiments suggest that cultured porcine aortic endothelial cells release two endothelium-derived relaxing factors; one is released under basal conditions and upon stimulation with adenosine diphosphate and the other (which presumably is nitric oxide) upon stimulation with bradykinin and the calcium ionophore A23187.

Vasoconstrictor responses are augmented in porcine coronary arteries in hypercholesterolemia and atherosclerosis, leading to an occurrence of coronary vasospasm in the latter condition. The role of the endothelium in the vascular hyperreactivity in hypercholesterolemic and atherosclerotic coronary arteries was examined, particularly in response to aggregating and related vasoactive substances. Male Yorkshire pigs underwent balloon endothelial denudation of the left anterior descending coronary artery (LAD) and 2% high-cholesterol feeding for 10 weeks. Electron microscopic examination demonstrated a full lining of endothelial cells in the LAD and the left circumflex coronary artery (LCX). Endothelium-dependent responses were examined in vitro. In cholesterol-fed animals, endothelium-dependent relaxations to aggregating platelets, serotonin, ADP, bradykinin, thrombin, and the calcium ionophore A23187 were depressed in LAD (atherosclerosis), while the relaxations to aggregating platelets, serotonin and ADP were depressed in LCX (hypercholesterolemia). Serotonin-induced contractions were endothelium-dependently augmented in atherosclerotic LAD; the endothelium-dependent component of the contractions was inhibited by blockers of cyclooxygenase. Bioassay studies demonstrated a depressed release of endothelium-derived relaxing factor(s) from the atherosclerotic LAD in response to serotonin. These experiments indicate that the endothelium-dependent relaxations to aggregating platelets and related vasoactive substances are severely impaired in atherosclerosis and moderately impaired in hypercholesterolemia. Since coronary atherosclerosis was induced by a combination of balloon endothelial injury (and regeneration) and high-cholesterol feeding in this study, the combined effects of those factors must account for the severely impaired responses in atherosclerosis. The depressed release of the endothelium-derived relaxing factor(s) and the concomitant release of vasoconstrictor product(s) of cyclooxygenase appear to be responsible for the impaired relaxations.

The role of the endothelium was examined in the response to aggregating platelets in cerebral arteries from normal and hypercholesterolemic animals. Male Yorkshire pigs were fed either a normal diet or a 2% high-cholesterol diet for 10 weeks. Endothelium-dependent responses were examined in vitro. In rings of basilar arteries from control animals aggregating platelets caused endothelium-dependent relaxations, which were significantly inhibited by apyrase, an adenosine diphosphatase and triphosphatase, but were augmented by methiothepin, a combined S1- and S2-serotonergic blocker. In quiescent rings platelets induced contractions that were inhibited by the presence of the endothelium; these contractions were significantly inhibited by methiothepin, but not by ketanserin (an S2-serotonergic blocker) or dazoxiben (a thromboxane-synthetase blocker) in the presence or absence of SQ29548 (a thromboxane-receptor blocker). Adenosine diphosphate but not serotonin caused endothelium-dependent relaxations. In cholesterol-fed animals the endothelium-dependent relaxations in response to aggregating platelets and adenosine diphosphate were impaired. These experiments indicate that 1) the endothelium inhibits the vasoconstrictor effect of aggregating platelets in porcine cerebral arteries; 2) platelet-induced relaxations are achieved mainly by a purinergic mechanism, while platelet-induced contractions are mediated by activation of S1-serotonergic receptors with little contribution of thromboxanes; and 3) hypercholesterolemia impairs the endothelium-dependent relaxations in response to aggregating platelets due to the impaired responses to adenosine diphosphate.

Experiments were designed in a bioassay system to analyze the effect of elevated (from 5.9 mM to 7.5-45.9 mM) extracellular K+ concentration on the release of endothelium-derived relaxing factor. Segments of canine femoral artery with endothelium (donor segment) were mounted in an organ bath and perfused with modified Krebs-Ringer bicarbonate solution; the effluent from the donor segment was used to superfuse a canine coronary artery ring without endothelium (bioassay tissue). Elevation of perfusate K+ concentration by 1.6-15 mM by intraluminal infusion of potassium chloride upstream of the donor segment evoked further contractions of bioassay rings contracted with prostaglandin F2 alpha. In contrast, the bioassay rings progressively relaxed when increasing concentrations of potassium chloride (10-40 mM) were added extraluminally to the organ bath where the perfused segment was mounted. Extraluminal application of phenylephrine or prostaglandin F2 alpha did not evoke relaxations in the bioassay ring. Removal of the endothelium from the donor segment or selective exposure of the segment (but not the bioassay ring) to Ca2+-deficient solution prevented the K+-induced relaxations. Treatment of the donor segment and the bioassay ring with inhibitors of known endogenous vasoactive substances (acetylcholine, norepinephrine, adenine nucleotides, and prostanoids) had no significant effect on the relaxation of the bioassay ring evoked by extraluminal application of potassium chloride. Simultaneous measurements of changes in isometric force in the donor segment and bioassay ring revealed that extraluminal elevation of K+ concentration relaxed the segments as well and that the relaxations could not be prevented by simultaneous intraluminal infusion of potassium chloride.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in the isometric tension of isolated strips of cutaneous, femoral, mesenteric, pulmonary, and muscle arteries and veins were recorded at 37°C in an organ bath. Acetylcholine (5 x 10 -8 and 10 -7 g/ml) caused relaxation of strips from the saphenous veins, the femoral veins, and all of the arteries after contraction by norepinephrine released from nerve terminals by electrical stimulation (2-5 Hz); in the pulmonary and mesenteric veins, acetylcholine caused a further increase in tension. Pulmonary artery and mesenteric vein strips were incubated with [ 3 H] norepinephrine and mounted for superfusion (3 ml/min) and isometric tension recording. Electrical stimulation increased the tension and the total radioactivity released in both preparations. Acetylcholine (2 x 10 -7 g/ml) depressed the contractions of the pulmonary artery strips but augmented those of the mesenteric vein strips; it diminished the efflux of radioactivity in both, indicating that acetylcholine inhibits adrenergic neurotransmission. In the absence of sympathetic stimulation, acetylcholine (5 x 10 -10 -10 -5 g/ml) caused all vein strips to contract; the most common reaction in artery strips was a slight relaxation (at 10 -9 -10 -8 g/ml) followed by a contraction (at 5 x 10 -8 -10 -5 g/ml). During contractions caused by norepinephrine, acetylcholine caused a further increase in tension in vein strips but a relaxation in artery strips. Atropine abolished the effects of acetylcholine. The results of this study suggest the presence in vascular smooth muscle of both excitatory and inhibitory cholinergic receptors.