calcimycin and Arteriosclerosis

The objective was to test the hypothesis that dietary copper inhibits atherosclerosis by inducing superoxide dismutase (SOD) and potentiating nitric oxide (NO). New Zealand White rabbits were fed either a cholesterol diet (n = 8) or a cholesterol diet containing 0.02% copper acetate (n = 8) for 13 weeks. We found that the intimal area was significantly smaller in the animals supplemented with copper (P < 0.005), although integrated plasma cholesterol levels were not significantly different. This was associated with a significant increase in aortic copper content (P < 0.05), SOD activity (P < 0.05) and Cu/Zn SOD mRNA (P < 0.05) and a significant decrease in nitrotyrosine content (P < 0.05). Furthermore, there was a positive correlation between aortic copper content and SOD activity (P < 0.005, R(2) = 0.83) and a negative correlation between aortic superoxide dimutase activity and nitrotyrosine content (P < 0.005, R(2) = 0.93). In organ bath experiments, the relaxation of precontracted carotid artery rings to calcium ionophore was greater in animals supplemented with copper. No difference in response to sodium nitroprusside was observed. These data suggest that in the cholesterol-fed rabbit, copper supplements inhibit the progression of atherosclerosis by increasing SOD expression, thereby reducing the interaction of NO with superoxide, and hence potentiating NO-mediated pathways that may protect against atherosclerosis.

Topics: Animals; Aorta, Thoracic; Arteriosclerosis; Calcimycin; Carotid Arteries; Cholesterol; Copper; Dietary Supplements; Ionophores; Muscle, Smooth; Nitric Oxide; Oxidative Stress; Rabbits; Superoxide Dismutase; Tyrosine

Atheroma macrophages internalize large quantities of lipoprotein-derived lipids. While most emphasis has been placed on cholesterol, lipoprotein-derived fatty acids may also play important roles in lesional macrophage biology. Little is known, however, about the trafficking or metabolism of these fatty acids. In this study, we first show that the cholesterol-fatty acyl esterification reaction, catalyzed by acyl-CoA:cholesterol acyltransferase (ACAT), competes for the incorporation of lipoprotein-derived fatty acids into cellular phospholipids. Furthermore, conditions that inhibit trafficking of cholesterol from late endosomes/lysosomes to the endoplasmic reticulum (ER), such as the amphipathic amine U18666A and the Npc1+/- mutation, also inhibit incorporation of lipoprotein-derived fatty acids into phospholipids. The biological relevance of these findings was investigated by studying the suppression of agonist-induced prostaglandin E(2) (PGE(2)) and leukotriene C(4)/D(4)/E(4) production during lipoprotein uptake by macrophages, which has been postulated to involve enrichment of cellular phospholipids with non-arachidonic fatty acids (NAAFAs). We found that eicosanoid suppression was markedly enhanced when ACAT was inhibited and prevented when late endosomal/lysosomal lipid trafficking was blocked. Moreover, PGE(2) suppression depended entirely on acetyl-LDL-derived NAAFAs, not on acetyl-LDL-cholesterol, and was not due to decreased cPLA(2) activity per se. These data support the following model: lipoprotein-derived NAAFAs traffic via the NPC1 pathway from late endosomes/lysosomes to a critical pool of phospholipids. In competing reactions, these NAAFAs can be either esterified to cholesterol or incorporated into phospholipids, resulting in suppression of eicosanoid biosynthesis. In view of recent evidence suggesting dysfunctional cholesterol esterification in late lesional macrophages, these data predict that such cells would have highly suppressed eicosanoid synthesis, thus affecting eicosanoid-mediated cell signaling in advanced atherosclerosis.

Topics: Androstenes; Animals; Arteriosclerosis; Calcimycin; Cholesterol; Dinoprostone; Eicosanoids; Endoplasmic Reticulum; Endosomes; Enzyme Inhibitors; Esterification; Fatty Acids; Humans; Intracellular Signaling Peptides and Proteins; Leukotrienes; Lipoproteins; Lipoproteins, LDL; Lysosomes; Macrophages; Macrophages, Peritoneal; Mice; Mice, Inbred C57BL; Mutation; Niemann-Pick C1 Protein; Niemann-Pick Diseases; Phospholipases A; Phospholipids; Proteins; Recombinant Proteins; Signal Transduction; Sterol O-Acyltransferase

Current consensus suggests that lysophosphatidylcholine is the major detrimental factor in oxidized low-density lipoprotein that may contribute to the alterations of vasomotor responses associated with atherosclerosis. This study investigated the influences of lysophosphatidylcholine and major lipid components in oxidized low-density lipoprotein on vascular relaxation. We also determine if there was any interaction between these phospholipid components on relaxation. Porcine coronary artery rings were incubated with lysophosphatidylcholine, phosphatidylcholine or sphingomyelin. After contraction by the thromboxane A2 mimetic U46619, rings were relaxed with bradykinin and calcium ionophore A23187. Lysophosphatidylcholine with a higher proportion of stearoyl-lysophosphatidylcholine to palmitoyl-lysophosphatidylcholine ratio caused greater reduction of relaxational responses. While phosphatidylcholine and sphingomyelin had no effect on vascular relaxation, they reduced the ability of lysophosphatidylcholine to impair vascular relaxation. Our results thus suggested that the effectiveness of oxidized low-density lipoprotein at inhibiting vasodilatory responses may be determined by the relative proportion of different types of lysophosphatidylcholine as well as the amount of other phospholipid components: phosphatidylcholine and sphingomyelin.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Animals; Arteriosclerosis; Bradykinin; Calcimycin; Calcium; Cattle; Coronary Vessels; Dose-Response Relationship, Drug; Endothelium, Vascular; Ionophores; Lipoproteins, LDL; Liver; Lysophosphatidylcholines; Oxygen; Phosphatidylcholines; Phospholipids; Sphingomyelins; Swine; Thromboxane A2; Vasoconstrictor Agents

This study was to investigate the effects of chronic exercise on vasodilatation and endothelial intracellular calcium (EC [Ca2+]i) signaling in atherosclerotic animals.. For 8 weeks, male New Zealand White rabbits were fed rabbit chow with or without the addition of 2% cholesterol. They were further divided into control and exercise groups. Animals in the exercise groups ran on a leveled treadmill at 0.88 km/h for 10 to 60 minutes gradually for 5 days per week for a total of 8 weeks. At the end of experiments, femoral arteries were dissected, loaded with fura 2-AM, and mounted in a tissue flow chamber. PE-precontracted vessel specimens were exposed to acetylcholine (ACh). The EC [Ca2+]i elevation and vasorelaxation were determined simultaneously under an epifluorescence microscope equipped with a ratio-imaging capability. Our results showed the following: (1) high cholesterol diet feeding caused lipid deposition on vascular surface, reduced the ACh-evoked EC [Ca2+]i elevation, and impaired endothelium-dependent and endothelium-independent vascular responses, but chronic exercise had the opposite effects; (2) ACh-induced vasorelaxation was associated with EC [Ca2+]i elevation in all groups; and (3) vasorelaxation at high levels of EC [Ca2+]i elevation decreased in hypercholesterolemia.. Our data suggest that hypercholesterolemia induces vascular structural changes and impairs EC [Ca2+]i signaling and vasodilatation, whereas chronic exercise partially reverses these adverse effects.

Topics: Acetylcholine; Animals; Arteriosclerosis; Azo Compounds; Calcimycin; Calcium; Calcium Signaling; Citrate (si)-Synthase; Coloring Agents; Endothelium, Vascular; Femoral Artery; Hypercholesterolemia; In Vitro Techniques; Lipid Metabolism; Lipids; Male; Muscle Contraction; Muscle, Skeletal; Nitroprusside; Phenylephrine; Physical Conditioning, Animal; Rabbits; Tunica Intima; Vasodilation; Vasodilator Agents

Tetrahydrobiopterin (BH4) is of fundamental importance for the normal function of endothelial NO synthase. The purpose of this study was to investigate the effects of hyperlipidemia on vascular BH4 levels and the effect of supplementation with sepiapterin in the presence and absence of N-acetylcysteine (NAC).. New Zealand White rabbits were fed normal chow (normocholesterolemic [NC] group) or hyperlipidemic chow (hyperlipidemic [HL] group) for 8 to 10 weeks. Mean cholesterol levels were 1465+/-333 and 53+/-17 mg/dL in the HL and NC group, respectively. Markedly diminished BH4 levels were found in the HL group compared with the NC group, but these levels could be restored after 6 hours of incubation with sepiapterin. Peak relaxations to acetylcholine and A23187 were impaired in the HL group. Supplementation with sepiapterin resulted in a further diminution of relaxation in the HL but not NC group. Incubation with NAC for 6 hours failed to raise BH4 levels, whereas NAC in conjunction with sepiapterin raised BH4 levels approximately 221-fold. However, this increase did not improve relaxations to A23187 and acetylcholine.. Prolonged exposure to sepiapterin impairs vasorelaxation in hyperlipidemia despite repletion of endogenous BH4. Antioxidant thiols do not correct this impairment. These studies have implications for the use of sepiapterin in the correction of vasomotor tone in atherosclerosis.

Topics: Acetylcholine; Acetylcysteine; Animals; Antioxidants; Aorta, Thoracic; Arteriosclerosis; Biopterins; Calcimycin; Cholesterol; Diet; Endothelium, Vascular; Free Radical Scavengers; Hyperlipidemias; Ionophores; Male; Muscle, Smooth, Vascular; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Oxidation-Reduction; Oxygen; Pteridines; Pterins; Rabbits; Sulfhydryl Compounds; Vasodilator Agents

We have determined whether the anti-atherosclerotic effect of a 3-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase inhibitor (fluvastatin) is mediated through nitric oxide (NO) as well as affecting plasma lipids. NO related vascular responses, endothelial nitric oxide synthase (eNOS) mRNA and superoxide anion (O(2)(-)) release were examined in vascular walls of oophorectomized female rabbits fed 0.5% cholesterol chow for 12 weeks with or without fluvastatin (2 mg/kg per day). Serum lipid profile was not different between two groups. NO dependent responses stimulated by acetylcholine and calcium ionophore A23187 and tone related basal NO response induced by N(G)-monomethyl-L-arginine acetate (L-NMA); nitric oxide synthase inhibitor were all improved by fluvastatin treatment. Endothelium independent vasorelaxation induced by nitroglycerin was not different between the two groups of rabbits' arteries. Fluvastatin treatment increased cyclic GMP concentration in aorta of rabbits. eNOS mRNA expression and O(2)(-) release were measured in aorta using competitive reverse transcription-polymerase chain reaction (RT-PCR) and with lucigenin analogue, 2-methyl-3,7-dihydroimidazol [1,2-a]pyrazine-3-one (MCLA) chemiluminescence methods. eNOS mRNA in the endothelial cells of aorta was significantly up-regulated and O(2)(-) production was significantly reduced in fluvastatin treated rabbit aorta. Anti-macrophage staining area, but not anti-smooth muscle cell derived actin stained area in the aorta was also reduced by fluvastatin treatment. Conclusion, fluvastatin, a HMG-CoA reductase inhibitor, retards the initiation of atherosclerosis formation through the improvement of NO bioavailability by both up-regulation of eNOS mRNA and decrease of O(2)(-) production in vascular endothelial cells, and this means that part of the anti-atherosclerotic effect of fluvastatin may be due to nonlipid factors.

Topics: Acetylcholine; Animals; Antioxidants; Aorta; Arteriosclerosis; Calcimycin; Calcium; Endothelium, Vascular; Enzyme Induction; Enzyme Inhibitors; Fatty Acids, Monounsaturated; Female; Fluvastatin; Free Radical Scavengers; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Indoles; Ionophores; Lipids; Lipoproteins, LDL; Luminescent Measurements; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type III; Nitroarginine; Ovariectomy; Rabbits; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Superoxide Dismutase; Superoxides

Hyperlipidemia may increase endothelial damage and promote accelerated atherogenesis in graft coronary vasculopathy. To study the effects of hypercholesterolemia on coronary endothelial dysfunction, intimal hyperplasia, and lipid content, a porcine model of heterotopic heart transplantation, allowing nonacute rejection without immunosuppressive drugs, was used. A high cholesterol diet was fed to donor and recipient swine 1 month before and after transplantation. The endothelial function of coronary arteries of native and transplanted hearts from cholesterol-fed animals was studied in organ chambers 30 days after implantation and compared with endothelial function in arteries from animals fed a normal diet. The total serum cholesterol increased 3-fold in donors and recipients. Endothelium-dependent relaxations to serotonin, to the alpha(2)-adrenergic agonist UK14,304, and to the direct G-protein activator sodium fluoride were decreased significantly in allografted hearts compared with native hearts from both groups. Relaxations to the calcium ionophore A23187 and bradykinin were decreased significantly in allografts from animals fed the high cholesterol diet. The prevalence of intimal hyperplasia was significantly increased in coronary arteries from hypercholesterolemic swine. There was a significant increase in the lipid content of allograft arteries of hypercholesterolemic recipients. Hypercholesterolemia causes a general coronary endothelial dysfunction, increases the prevalence of intimal hyperplasia, and augments the incorporation of lipids in the vascular wall after heart transplantation. Hyperlipidemia accelerates graft coronary atherosclerosis through its effects on the endothelium.

Topics: Adrenergic alpha-Agonists; Animals; Arteriosclerosis; Biological Transport; Brimonidine Tartrate; Calcimycin; Calcium; Cholesterol, HDL; Cholesterol, LDL; Coronary Vessels; Diet, Atherogenic; Dinoprost; Dose-Response Relationship, Drug; Endothelium, Vascular; Erythrocyte Count; Female; Free Radical Scavengers; Heart Transplantation; Hematocrit; Hemoglobins; Hypercholesterolemia; Hyperplasia; Ionophores; Male; Myocardium; Postoperative Period; Potassium Chloride; Quinoxalines; Serotonin; Swine; Transplantation, Homologous; Tunica Intima; Vasodilation

Dehydroepiandrosterone (DHEA) is speculated to have an antiatherosclerotic effect, although the mechanism of action remains unclear. The objective of the current study was to determine whether the antiatherosclerotic effect of DHEA is related to its conversion to estrogen and to define the role of nitric oxide (NO) in the antiatherosclerotic effect of DHEA. Forty-eight oophorectomized rabbits were divided into 5 groups and fed the following diets for 10 weeks: group 1, a regular rabbit diet plus 1% cholesterol (a high-cholesterol diet [HCD]); group 2, an HCD plus 0.3% DHEA; group 3, an HCD plus 0.3% DHEA and fadrozole (2.0 mg x kg(-1) x d(-1)), a specific aromatase inhibitor; group 4, an HCD plus 17beta-estradiol (20 microg x kg(-1) x d(-1)); and group 5, a regular diet. Atherosclerotic lesions, lipid deposition in aortic vessels, and basal and stimulated NO release were measured in the aforementioned groups of rabbits. NO release was measured by using an NO-selective electrode as well as by measuring vascular responses and the plasma NO metabolites nitrite and nitrate. The plasma total cholesterol level was increased, but there were no significant differences in lipid profile in the 4 groups of rabbits that were fed the HCD. The area occupied by atherosclerosis in the thoracic aorta was diminished by approximately 60% in the DHEA-treated rabbits (group 2) compared with the HCD group of rabbits (group 1); there was a corresponding 80% decrease in the estradiol group (group 4) but only a 30% decrease in the DHEA plus fadrozole group (group 3). In the aortas of rabbits from groups 1 and 3, the acetylcholine-induced and tone-related basal NO-mediated relaxations were diminished compared with those of the controls (group 5). However, these relaxations were restored in the aortas of group 2 and 4 rabbits, and an increase in NO release was observed in groups 2 and 4 compared with groups 1 and 3, as measured by an NO-selective electrode. Injection of neither solvent (20% ethanol/distilled water) nor fadrozole significantly affected the atherosclerotic area or the NO-related responses described above. We conclude that approximately 50% of the total antiatherosclerotic effect of DHEA was achieved through the conversion of DHEA to estrogen. NO may also play a role in the antiatherosclerotic effect of DHEA and 17beta-estradiol.

Topics: Acetylcholine; Adjuvants, Immunologic; Animals; Aorta, Thoracic; Aromatase; Arteriosclerosis; Calcimycin; Cholesterol, Dietary; Cyclic GMP; Dehydroepiandrosterone; Diet, Atherogenic; Electrodes; Endothelium, Vascular; Estrogens; Female; Ionophores; Nitrates; Nitric Oxide; Nitrites; Rabbits; Triglycerides; Vasodilation; Vasodilator Agents

In an experimental model of atherosclerosis we investigated whether rabbits fed an atherogenic diet (0.25% cholesterol, 3% coconut oil) develop endothelial dysfunction accompanied with increased infarct mass compared to normal fed rabbits and, whether hypercholesterolemia would interfere with the beneficial outcome of ischemic preconditioning observed in normal rabbits. After four weeks on either a normal or an atherogenic diet, New Zealand White rabbits (n=7 in each group) were subjected to 30 min of myocardial ischemia by occlusion of a branch of the left anterior descending coronary artery (LAD) followed by 2 hours of reperfusion (infarct studies). For ischemic preconditioning experiments, LAD was additionally occluded twice for 5 min followed by 10 min reperfusion before the long-lasting (30 min) ischemia. Infarct mass was evaluated by triphenyl-tetrazolium staining. Besides the assessment of aortic endothelium-dependent function and NO-release, aortic and cardiac vessels were inspected for atherosclerotic lesions. Total cholesterol serum levels in rabbits on an atherogenic diet were significantly higher (15.3+/-2.7 mmol/L) than those on a standard diet (0.65+/-0.08 mmol/L). The aortas and heart vessels were without any histological evidence of atherosclerosis, whereas endothelial dysfunction and significantly reduced calcium-ionophore stimulated endothelial NO-release were found in isolated aortic rings of hypercholesterolemic animals. Rabbits on a standard diet showed an infarct mass (related to the area at risk) of 41+/-33%, which was reduced to 21+/-2% by ischemic preconditioning (49% decrease, p<0.05). In rabbits on an atherogenic diet, infarct mass was significantly increased to 63+/-3% (52% increase versus standard diet). Interestingly, hypercholesterolemia did not affect the beneficial influence of ischemic preconditioning; infarct mass (21+/-3%, p<0.05 vs hypercholesterolemia) was similar to rabbits on a standard diet with ischemic preconditioning. Our results show that experimental hypercholesterolemia increases infarct mass in nonpreconditioned hearts but it does not interfere with the reduction of infarct mass elicited by preconditioning. This may suggest that NO produced by the endothelium is not a prime factor in the cardioprotective mechanism of preconditioning.

Topics: Animals; Aorta, Thoracic; Arteriosclerosis; Calcimycin; Cholesterol; Diet, Atherogenic; Disease Models, Animal; Endothelium, Vascular; Enzyme Inhibitors; Free Radical Scavengers; Hypercholesterolemia; Ionophores; Ischemic Preconditioning, Myocardial; Male; Myocardial Infarction; NG-Nitroarginine Methyl Ester; Nitric Oxide; Polyethylene Glycols; Potassium Chloride; Rabbits; Superoxide Dismutase; Vasodilation

Superoxide anion plays important roles in vascular disease states. Increased superoxide production contributes to reduced nitric oxide (NO) bioactivity and endothelial dysfunction in experimental models of vascular disease. We measured superoxide production by NAD(P)H oxidase in human blood vessels and examined the relationships between NAD(P)H oxidase activity, NO-mediated endothelial function, and clinical risk factors for atherosclerosis. Endothelium-dependent vasorelaxations and direct measurements of vascular superoxide production were determined in human saphenous veins obtained from 133 patients with coronary artery disease and identified risk factors. The predominant source of vascular superoxide production was an NAD(P)H-dependent oxidase. Increased vascular NAD(P)H oxidase activity was associated with reduced NO-mediated vasorelaxation. Furthermore, reduced endothelial vasorelaxations and increased vascular NAD(P)H oxidase activity were both associated with increased clinical risk factors for atherosclerosis. Diabetes and hypercholesterolemia were independently associated with increased NADH-dependent superoxide production. The association of increased vascular NAD(P)H oxidase activity with endothelial dysfunction and with clinical risk factors suggests an important role for NAD(P)H oxidase-mediated superoxide production in human atherosclerosis. The full text of this article is available at http://www.circresaha.org.

Topics: Acetylcholine; Aged; Analysis of Variance; Arteriosclerosis; Calcimycin; Endothelium, Vascular; Female; Humans; Ionophores; Male; Middle Aged; NADH, NADPH Oxidoreductases; NADPH Oxidases; Nitroprusside; Risk Factors; Saphenous Vein; Superoxides; Vasodilator Agents; Vasomotor System

Thrombin activation requires assembly of a prothrombinase complex of activated coagulation factors on an anionic phospholipid surface, classically provided by activated platelets. We have previously shown that anionic phosphatidylserine is exposed by rat vascular smooth muscle cells (VSMCs) undergoing apoptosis after serum withdrawal. In this study, using a chromogenic assay, we have shown thrombin generation by apoptotic VSMCs expressing c-myc (VSMC-myc) with an area under the thrombin-generation curve (AUC) of 305 +/- 17 nmol x min/L and a peak thrombin (PT) of 154 +/- 9 nmol/L. The thrombin-generating potential of the apoptotic VSMC-myc cells was greater than that of unactivated platelets (P = .003 for AUC; P = .0002 for PT) and similar to calcium-ionophore activated platelets (AUC of 332 +/- 15 nmol x min/L, P = .3; PT of 172 +/- 8 nmol/L, P = .2). Thrombin activation was also seen with apoptotic human VSMCs (AUC of 211 +/- 8 nmol x min/L; PT of 103 +/- 4 nmol/L) and was inhibited by annexin V (P < .0001 for AUC and PT). VSMC-myc cells maintained in serum generated less thrombin than after serum withdrawal (P = .0002 for AUC and PT). VSMCs derived from human coronary atherosclerotic plaques that apoptose even in serum also generated thrombin (AUC of 260 +/- 2 nmol x min/L; PT of 128 +/- 4 nmol/L). We conclude that apoptotic VSMCs possess a significant thrombin-generating capacity secondary to phosphatidylserine exposure. Apoptotic cells within atherosclerotic plaques may allow local thrombin activation, thereby contributing to disease progression.

Topics: Adenovirus E1A Proteins; Animals; Annexin A5; Aorta, Thoracic; Apoptosis; Arteriosclerosis; Blood Platelets; Calcimycin; Calcium Chloride; Cells, Cultured; Coronary Artery Disease; Culture Media, Serum-Free; DNA, Complementary; Genes, myc; Hirudins; Humans; Ionophores; Muscle, Smooth, Vascular; Platelet Activation; Proto-Oncogene Proteins c-myc; Rats; Rats, Sprague-Dawley; Recombinant Fusion Proteins; Thrombin; Transfection

We examined the vascular structure and endothelium-dependent relaxation in two genetic models of hypercholesterolemia: apolipoprotein E (apoE)-knockout mice and combined apoE/LDL receptor-double-knockout mice. Intimal area was increased markedly in proximal segments of thoracic aortas from apoE/LDL receptor-knockout mice [0.13 +/- 0.03 (mean +/- SE) mm2] compared with normal (C57BL/6J) mice (0.002 +/- 0.002 mm2, P < .05). Despite intimal thickening, the vascular lumen was not smaller in the aortas of apoE/LDL receptor-knockout mice (0.52 +/- 0.03 mm2) than in normal mice (0.50 +/- 0.03 mm2). In apoE-deficient mice, intimal thickening was minimal or absent, even though the concentration of plasma cholesterol was only modestly less than that in the double-knockout mouse (14.9 +/- 1.1 vs 18.0 +/- 1.2 mmol/L, respectively, P < .05). Relaxation of the aorta was examined in vitro in vascular rings precontracted with U46619. In normal mice, acetylcholine produced relaxation, which was markedly attenuated by the nitric oxide synthase inhibitor NG-nitro-L-arginine (100 microM). Relaxation to acetylcholine and the calcium ionophore A23187 was normal in apoE-deficient mice (in which lesions were minimal) but greatly impaired in the proximal segments of thoracic aortas of apoE/LDL receptor-deficient mice, which contained atherosclerotic lesions. Vasorelaxation to nitroprusside was similar in normal and apoE-knockout mice, with modest but statistically significant impairment in atherosclerotic segments of apoE/LDL receptor-knockout mice. In distal segments of the thoracic aorta of apoE/LDL receptor-deficient mice, atherosclerotic lesions were minimal or absent, and the endothelium-dependent relaxation to acetylcholine and calcium ionophore was normal. Thus, in apoE/LDL receptor-knockout mice (a genetic model of hyperlipidemia), there is vascular remodeling with preservation of the aortic lumen despite marked intimal thickening, with impairment of endothelium-dependent relaxation to receptor- and nonreceptor-mediated agonists. Atherosclerosis may be accelerated in the apoE/LDL receptor-double-knockout mouse compared with the apoE-knockout strain alone. We speculate that other factors, such as the absence of LDL receptors, may contribute to the differences in the extent of atherosclerosis in these two models of hyperlipidemia.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Acetylcholine; Animals; Aorta, Thoracic; Aortic Diseases; Apolipoproteins E; Arteriosclerosis; Calcimycin; Calcium; Disease Models, Animal; Endothelium, Vascular; Enzyme Inhibitors; Female; Hypercholesterolemia; Ionophores; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle Relaxation; Nitric Oxide Synthase; Nitroarginine; Receptors, LDL; Superoxide Dismutase; Vasoconstrictor Agents

Tissue destruction in atherosclerosis is partly due to uncontrolled protease and oxygen radical release. In this study we investigated the release of elastase and myeloperoxidase, as well as the production of reactive oxygen species by polymorphonuclear leukocytes (PMNLs) obtained from patients with obliterative atherosclerotic of the lower legs. In addition we measured the plasma concentration of xanthine oxidase. PMNLs of atherosclerotic patients have a greater ability to increase elastase and myeloperoxidase release after their stimulation with formyl-methionin-leucyl-phenylalanin (fMLP) and calcium ionophore, A23187, independently of their age, than PMNLs of healthy middle-aged subjects. Similarly to healthy elderly subjects there was an increased superoxide anion (O2-) production under basal condition in both atherosclerotic patient age-groups. The activation of PMNLs with fMLP and A23187 enhanced O2- formation both in healthy subjects and in patients with atherosclerotic disease of the lower legs, however the increase was significantly less in the latter group. No biochemical parameters showed significant correlation with patient's risk factors, however myeloperoxidase production was significantly higher in less severe stage of the disease (P < 0.05). We found that patients with atherosclerotic disease of the lower legs have higher plasma xanthine oxidase level than control subjects. This study indicates an other piece of evidence suggesting the activation and involvement of neutrophils in the pathogenesis of atherosclerosis of the lower legs. The similar tendencies in the reactivity of neutrophils during aging and in atherosclerosis suggest that atherosclerosis may be an early aging process.

Topics: Adult; Aged; Arteriosclerosis; Calcimycin; Cell Degranulation; Elastin; Humans; Hydrogen Peroxide; Ionophores; Leukocyte Elastase; Middle Aged; N-Formylmethionine Leucyl-Phenylalanine; Neutrophils; Pancreatic Elastase; Peroxidase; Respiratory Burst; Superoxides; Xanthine Oxidase

The lipoxygenase product 15-hydroxyeicosatetraenoic acid (15-HETE) was shown to be the most important eicosanoid formed in the atherosclerotic rabbit aorta. The aim of the present study was to compare the effects of 15-HETE and its hydroperoxy precursor 15-HpETE with those of other vasoconstrictor and vasodilator agents in arteries from rabbits fed either a control or a cholesterol-rich diet for 16 and 30 weeks. 5-Hydroxytryptamine (5-HT) aggregated platelets and thrombin caused contractions of isolated rabbit aortas. The contractile responses elicited by platelets from control animals were similar to those evoked by platelets from atherosclerotic rabbits. After 16 weeks of hypercholesterolemia, the contractile responses were either augmented (5-HT), unchanged (platelets) or reduced (thrombin). After 30 weeks of hypercholesterolemia, the responses to all contractile agents used had decreased. In both aortas and pulmonary arteries the endothelium-dependent relaxations to the calcium ionophore, A23167, and to acetylcholine were progressively lost and the endothelium-independent relaxations to nitroglycerin were reduced by the progressing hypercholesterolemia. The 15-lipoxygenase metabolites contracted the isolated thoracic aorta and pulmonary artery from control rabbits and to a lesser extent those of the cholesterol-fed rabbits. After raising the tone in these vessels with prostaglandin F2 alpha PGF2 alpha) or noradrenaline, 15-HpETE induced relaxations which were not significantly influenced by the development of fatty streaks. Our data illustrate that the contractions of the blood vessel wall to 15-HETE, like those to other vasoconstrictors, are markedly reduced by developing atherosclerosis. In contrast, the relaxations to 15-HpETE in the rabbit arteries remain unaltered after 16 to 30 weeks of hypercholesterolemia. This is unlike the reactions to other vasodilators, which are markedly reduced.

Topics: Acetylcholine; Animals; Arachidonate 15-Lipoxygenase; Arteriosclerosis; Calcimycin; Dinoprost; Hydroxyeicosatetraenoic Acids; Hypercholesterolemia; In Vitro Techniques; Leukotrienes; Lipid Peroxides; Male; Muscle, Smooth, Vascular; Nitroglycerin; Platelet Aggregation; Rabbits; Thrombin; Vasoconstrictor Agents; Vasodilator Agents

The mechanism by which women in the reproductive age group are protected from developing coronary heart disease (CHD) as compared with men of similar age is not known. To elucidate whether there is a gender difference in the rate of atherosclerosis formation, we investigated the rate of development of atherosclerosis in both male and female rabbits fed an identical diet consisting of 2% cholesterol for 10 and 15 weeks. The extent of atherosclerosis was correlated with the amount of basal and stimulated release of nitric oxide (NO) from endothelium-intact aortic rings obtained from these animals. Under identical dietary conditions, the female rabbits fed a high cholesterol diet (HCD) for 10 weeks developed very little atherosclerosis (10% surface involvement) as compared with male rabbits (42% surface involvement). However, no significant gender differences in atherosclerosis were observed after 15 weeks of the HCD. The serum cholesterol, high and low density lipoprotein (HDL and LDL) cholesterol were similar in animals fed the HCD for 10 and 15 weeks. The basal release of NO from endothelium-intact aortic rings was significantly greater in control females as compared with males. The magnitude of endothelium-dependent relaxation of aortic rings obtained from both male and female rabbits fed the HCD were impaired to a similar extent, and this impairment correlated with the duration of hyperlipidemia but not with the extent of atherosclerosis. The arginine content of aortic rings were not different between males (257 +/- 52 nmol/g wet weight) and females (345 +/- 62 nmol/g wet weight) or between control and hyperlipidemic groups (males 312 +/- 69; females 301 +/- 65 nmol/g wet weight). Although the precise mechanism for the slower rate of development of atherosclerosis in the female rabbits as compared with males is not clear, the greater basal release of NO in females before they were fed a hyperlipidemic diet, as well as other factors, may be involved. The impairment of endothelium-dependent relaxation in hyperlipidemic animals is not due to a decrease in the availability of arginine, the substrate for NO.

Topics: Acetylcholine; Animals; Arginine; Arteriosclerosis; Calcimycin; Estradiol; Estrone; Female; In Vitro Techniques; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; omega-N-Methylarginine; Rabbits; Sex Factors; Vasodilation

The objectives of this study were to determine if psychosocial stress impairs dilation through endothelium-derived relaxing factor (EDRF)-mediated mechanisms and if this effect is long lasting. Monkeys were fed an atherogenic diet for 36 mo while in one of three experimental conditions: (a) stable social groups ("unstressed," n = 6); (b) unstable social groups for the first half of the experiment and stable groups for the second half ("early stress," n = 8); and (c) stable groups for the first half of the experiment and unstable groups for the second half ("late stress," n = 6). Iliac arteries were studied in organ chambers containing Krebs' buffer and 10(-6) M indomethacin. Arteries from the late stress group had impaired dilation (shift of the dose-response curve down and to the right) to acetylcholine and the calcium ionophore A23187 (for both, P < 0.05), but not to nitroprusside (P > 0.05), compared with unstressed or early stress monkeys. NG-methyl-L-arginine reduced the dose-response curve to both acetylcholine and A23187 in the unstressed group and resulted in similar vascular responses among all three groups (P > 0.05). We conclude that current, but not previous, exposure to chronic stress impairs endothelium-mediated dilation of atherosclerotic iliac arteries of cynomolgus monkeys through an EDRF-mediated mechanism.

Topics: Acetylcholine; Animals; Arginine; Arteriosclerosis; Calcimycin; Diet, Atherogenic; Dose-Response Relationship, Drug; Iliac Artery; In Vitro Techniques; Indomethacin; Macaca fascicularis; Male; Muscle, Smooth, Vascular; Nitroprusside; omega-N-Methylarginine; Social Behavior; Stress, Psychological; Vasodilation

During neointima formation, which is an early and essential step in the development of atherosclerosis, endothelium-independent relaxations (nitroglycerin, 3-morpholinosydnonimine) are preserved, whereas muscarinic endothelium-dependent relaxation becomes impaired. The present study was undertaken to determine the selectivity of this impairment. The neointima was induced by positioning a nonocclusive, soft silicone collar around the left carotid artery of rabbits. The contralateral artery served as a control. Seven days later, vascular rings were mounted in organ chambers, contracted with phenylephrine (0.35 microM), and cumulative dose-relaxation curves were made. Intima-bearing vessels were less sensitive to acetylcholine, confirming the original observation. In contrast, the dose-relaxation curves for substance P and for the calcium ionophore A23187 were not altered in the presence of neointima. The curve for ATP was even shifted to the left. These results suggest that the nitric oxide synthase: cyclic GMP system remains intact in intima-bearing vessels and that the diminished endothelium-dependent relaxations are due to a selective alteration of the muscarinic receptors.

Topics: Adenosine Triphosphate; Animals; Arteriosclerosis; Calcimycin; Dose-Response Relationship, Drug; Male; Nitric Oxide; Rabbits; Receptors, Muscarinic; Substance P; Vasodilation

The aim of our study was to examine the release of various lipid and peptide contracting autacoids by aortae of normal and atherosclerotic rabbits. Leukotriene (LT) E4, an enzymatic derivative of LTC4, thromboxane (Tx) B2, and endothelin-1 (ET-1) were measured by radioimmunoassay techniques in aortic preparations of normal and cholesterol-fed rabbits. Intact aortae of normal rabbits incubated with the calcium ionophore A23187 for 1 h at 37 degrees C released LTE4 and TxB2 (22 +/- 3.5 and 14.8 +/- 2 pg/mg of tissue, respectively, mean +/- SEM, n = 33). Removal of aortic endothelium was associated with a significant reduction in LTE4 (44%) and TxB2 (58%) release. In aortic preparations from cholesterol-fed rabbits, the release of LTE4 was significantly enhanced (41 +/- 8 pg/mg of tissue, mean +/- SEM, n = 27) whereas TxB2 was not significantly altered. No detectable amounts of ET-1 were measured after 1 h of incubation. However, at 4 h, an endothelium-dependent release of ET-1 from normal aortae was demonstrated. In atherosclerotic aortae, ET-1 release was significantly higher than in controls (10 +/- 1.3 vs. 5 +/- 0.5 pg/cm2, mean +/- SEM, n = 16). We conclude that enhanced formation of vasoconstrictor autacoids may contribute to altered vasomotion of atherosclerotic blood vessels.

Topics: Animals; Aorta; Arteriosclerosis; Calcimycin; Cholesterol, Dietary; Endothelins; Indoles; Indomethacin; Leukotriene Antagonists; Leukotriene E4; Male; Rabbits; SRS-A; Thromboxane B2

The present investigation was undertaken to characterize the relaxing responsiveness in aortic strips from rats with arteriosclerosis, which was produced by excess vitamin D2 (VD) administration followed by treatment with or without a high-cholesterol diet for 6 weeks (VD + CHOL and VD group, respectively). This arteriosclerotic aorta was characterized by medial calcification and intimal cell proliferation. Helical strips of thoracic aorta were suspended in oxygenated Krebs-Henseleit solution. Under precontraction with noradrenaline, endothelium-dependent relaxations to acetylcholine were significantly attenuated in the VD and VD + CHOL as compared with the control. Relaxation to calcium ionophore A23187 was also significantly attenuated in the VD + CHOL. However, the relaxing responses to acetylcholine and A23187 in aortas from rats fed a high-cholesterol diet alone remained unaffected. Nitroglycerin caused an equal degree of relaxation in aortas from control and arteriosclerotic rats. There was a significant negative correlation between the relaxing response to acetylcholine and the calcium content in the aorta. These results indicate that in arteriosclerotic rat aortas, the endothelium-dependent relaxation to acetylcholine is impaired in proportion to the degree of calcification, and such impairment is facilitated by cholesterol feeding but can not be attributed to hypercholesterolemia or vascular cholesterol deposition.

Topics: Acetylcholine; Animals; Aorta; Arteriosclerosis; Calcimycin; Calcium; Cholesterol; Cholesterol, Dietary; Endothelium, Vascular; Ergocalciferols; In Vitro Techniques; Male; Nitroglycerin; Rats; Rats, Inbred Strains; Vasodilation

Estradiol is known to exert a protective effect against the development of atherosclerosis, but the mechanism of this hormonal action is unknown. One of the early events in the development of atherosclerosis is the adhesion of macrophages to endothelial cells, and nitric oxide (NO) inhibits this process. We show that basal release of NO is greater with endothelium-intact aortic rings from female rabbits than those from males. Oophorectomy diminishes both circulating estradiol concentration and basal release of NO to levels seen in male rabbits. These data establish that basal NO release from endothelium-intact aortic rings depends on circulating estradiol concentration and offer an explanation for the protective effect of estradiol against the development of atherosclerosis.

Topics: Acetylcholine; Animals; Aorta; Arginine; Arteriosclerosis; Calcimycin; Cholesterol; Citrulline; Estrogens; Female; In Vitro Techniques; Male; Nitric Oxide; Nitroglycerin; Ovariectomy; Rabbits; Sex Factors; Triglycerides

A diet containing 0.3% cholesterol was given to male New Zealand rabbits for 16 weeks; this produced atherosclerotic lesions (fatty streaks) on 80% of the intimal surface of the thoracic aorta and on 45% of the intimal surface of the abdominal aorta. The endothelium-dependent relaxations induced by acetylcholine, substance P and ionophore A23187 were inhibited in the atherosclerotic aortas. Besides the endothelium-independent relaxations induced by nitroglycerine, the relaxations induced by atrial natriuretic peptide (ANF) were also significantly reduced in the more atherosclerotic thoracic aorta. In bioassay experiments it was found that acetylcholine and substance P caused a smaller release of endothelium-derived relaxing factor (EDRF) from atherosclerotic thoracic aortas than from control thoracic aortas: the EDRF released by the vasodilators evoked less relaxation in atherosclerotic detector abdominal aortas than in control detector abdominal aortas. Nitric oxide evoked significantly less transient relaxation in the atherosclerotic thoracic and abdominal aortas than in the respective control tissues. The data indicate that as experimental atherosclerosis in the rabbit progresses, both vascular activity and EDRF release become affected; this leads to a complete loss of endothelium-dependent relaxation in the more atherosclerotic blood vessels.

Topics: Acetylcholine; Animals; Aorta, Abdominal; Aorta, Thoracic; Arteriosclerosis; Calcimycin; Cholesterol, Dietary; Male; Muscle Contraction; Muscle Relaxation; Nitric Oxide; Nitroglycerin; Rabbits; Substance P

Atherosclerosis impairs endothelium-dependent relaxation of large conduit arteries. Because coronary resistance vessels are spared from the development of overt atherosclerosis, endothelium-dependent responses were examined in these vascular segments. Malaysian cynomolgus monkeys (n = 6) were made atherosclerotic by being fed a 0.7% cholesterol diet for 18 months. Control monkeys (n = 6) were fed a standard diet. Coronary microvessels (122-220 microns) were studied in a pressurized (20 mm Hg), no-flow state using a video-imaging apparatus. Relaxations of microvessels, preconstricted with the thromboxane analogue U46619, were determined in response to acetylcholine, bradykinin, the calcium ionophore A23187, adenosine, and sodium nitroprusside. Microvascular relaxations to bradykinin and A23187 were reduced in atherosclerotic monkeys compared with controls, whereas acetylcholine produced additional contraction in atherosclerotic monkeys. Responses of preconstricted microvessels to adenosine and sodium nitroprusside were identical in atherosclerotic and control animals. Indomethacin did not alter responses in control or atherosclerotic animals. Histologic examination revealed neither intimal thickening nor plaque formation in microvessels of this size class despite marked changes in conduit arteries. Electron microscopy showed minor alterations of endothelial cell morphology in microvessels of atherosclerotic animals. In conclusion, long-term hypercholesterolemia markedly impairs endothelium-dependent vascular relaxation in the coronary microcirculation where overt atherosclerosis does not develop. These changes in endothelial cell function may significantly alter regulation of myocardial perfusion by neurohumoral stimuli.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Acetylcholine; Animals; Arteriosclerosis; Bradykinin; Calcimycin; Cholesterol, Dietary; Coronary Vessels; Endothelium, Vascular; Indomethacin; Macaca fascicularis; Male; Microcirculation; Microscopy, Electron; Prostaglandin Endoperoxides, Synthetic; Saponins; Vasodilation

We examined the hypothesis that impaired endothelium-dependent vasodilation in atherosclerosis is associated with decreased synthesis of nitrogen oxides by the vascular endothelium. The descending thoracic aortae of rabbits fed either normal diet, a high cholesterol diet for 2-5 wk (hypercholesterolemic, HC), or a high cholesterol diet for 6 mo (atherosclerotic, AS) were perfused in a bioassay organ chamber with physiologic buffer containing indomethacin. Despite a dramatic impairment in the vasodilator activity of endothelium-dependent relaxing factor (EDRF) released from both HC and AS aortae (assessed by bioassay), the release of nitrogen oxides (measured by chemiluminescence) from these vessels was not reduced, but markedly increased compared to NL. Thus, impaired endothelium-dependent relaxation in atherosclerosis is neither due to decreased activity of the enzyme responsible for the production of nitrogen oxides from arginine nor to arginine deficiency. Because the production of nitrogen oxides increased in response to acetylcholine in both hypercholesterolemic and atherosclerotic vessels, impairments in signal transduction are not responsible for abnormal endothelium-dependent relaxations. Impaired vasodilator activity of EDRF by cholesterol feeding may result from loss of incorporation of nitric oxide into a more potent parent compound, or accelerated degradation of EDRF.

Topics: Animals; Aorta; Arginine; Arteriosclerosis; Calcimycin; Cholesterol; Diet, Atherogenic; Luminescent Measurements; Nitric Oxide; Nitrogen Oxides; omega-N-Methylarginine; Rabbits; Time Factors

A biochemical correlation between vascular cholesterol metabolism and eicosanoid biosynthesis has not been fully elucidated. To assess the effects of cholesteryl ester (CE) accretion on eicosanoid synthesis, we studied eicosanoid metabolism in cultured rabbit aortic smooth muscle cells (SMC) following lipid-enrichment by incubation with cationized LDL (cLDL). SMC exposed to cLDL synthesized 50% less immunoreactive 6-keto-PGF1 alpha than untreated cells when exposed to the calcium ionophore, A-23187. In addition, cLDL-treatment reduced arachidonate acid (AA)-induced prostacyclin (PGI2) production sevenfold. Components of cLDL decreased eicosanoid biosynthesis in the following rank-order: linoleate greater than cholesterol greater than apo-cLDL. Lipid-enriched cells incorporated amounts of [1-14C]AA into phosphatidylcholine and phosphatidylethanolamine equal to control cells, but subsequent exposure to ionophore released significantly less radioactivity as free arachidonate (AA), with proportionally less conversion to eicosanoids. Ionophore released equivalent amounts of AA from all phospholipids, suggesting specificity for uptake, but not release of AA by cellular phospholipases. Cells enriched in CE had an eightfold decrease in percentage of phospholipid-derived AA relative to linoleate as compared to controls. Taken together, our data demonstrate that SMC metabolism of cLDL leads to cholesterol and CE accretion concomitant with diminished production of eicosanoids. Potential mechanisms for this effect include competitive inhibition of eicosanoid production by linoleate derived from LDL, direct inhibition of phospholipase A2 activity by cholesterol, and decrease in cyclooxygenase activity. These findings may have pathophysiological significance in that a reduction in PGI2 synthetic capacity of arterial SMC may exacerbate CE deposition since PGI2 promotes intracellular CE hydrolysis.

Topics: Animals; Aorta; Arteriosclerosis; Calcimycin; Cells, Cultured; Cholesterol; Cholesterol Esters; Cholesterol, LDL; Chromatography, High Pressure Liquid; Eicosanoic Acids; Lipid Metabolism; Muscle, Smooth, Vascular; Phospholipids; Prostaglandin-Endoperoxide Synthases; Rabbits; Sterol O-Acyltransferase

Mammalian tissues contain 5-, 12- and 15-lipoxygenases. Only the 15-lipoxygenase can act on linoleic acid, the predominant essential fatty acid of tissues and plasma, producing 13-hydroxyoctadecadienoic acid (13-HODE). Intracellular production of 13-HODE renders endothelial cells resistant to platelet adhesion, while its hydroperoxy precursor, 13-HPODE, synergises with the platelet anti-aggregatory factor prostacyclin. We have found that a 15-lipoxygenase activity is induced in aortas of cholesterol-fed and Watanabe Heritable Hyperlipidemic (WHHL) rabbits. Aortic tissue from WHHL rabbits incubated with 3H-linoleic acid produced a major metabolite identified as 13-HODE, which was formed with an efficiency comparable to the synthesis 15-HETE from arachidonic acid. These findings indicate that the increased aortic 15-lipoxygenase in vascular tissue is capable of producing 13-HODE in vivo. Since platelet adhesion is increased in atherogenesis, and thrombogenesis is a major complication of advanced atherosclerosis, it is suggested that induction of this enzyme may be a protective response to hypercholesterolemia.

Topics: Animals; Antithrombins; Aorta; Arachidonate 15-Lipoxygenase; Arachidonate Lipoxygenases; Arachidonic Acids; Arteriosclerosis; Calcimycin; Chromatography, High Pressure Liquid; Hydroxyeicosatetraenoic Acids; In Vitro Techniques; Linoleic Acids; Rabbits

We studied the effects of beta-migrating very low density lipoprotein (beta-VLDL) on the vascular responses of isolated thoracic aortic preparations taken from normal and hypercholesterolemic rabbits. The endothelium-dependent relaxation induced by acetylcholine or adenosine triphosphate (ATP) was attenuated in the arteries from hypercholesterolemic rabbits that were fed a cholesterol-rich diet for 12 weeks. In these aortas, the lesional circumference of the atherosclerotic plaques (fatty streaks) was only 12.18 +/- 1.98%. The relaxation induced by the Ca2+ ionophore A23187 or nitroglycerin was not altered. Preincubation with beta-VLDL significantly inhibited the relaxation due to acetylcholine, ATP, or A23187, especially in the aortas of hypercholesterolemic rabbits. However, beta-VLDL did not alter the response to nitroglycerin. Preincubation with high density lipoprotein had no significant effect on vessel relaxation. These results indicated that endothelium-dependent relaxation was already inhibited in the early stages of atherosclerosis, and that the atherogenic lipoprotein, beta-VLDL, further inhibited endothelium-dependent relaxation in atherosclerotic aortas. It may be that beta-VLDL also plays a role in determining the level of vascular tonus in atherosclerosis.

Topics: Acetylcholine; Adenosine Triphosphate; Animals; Aorta, Thoracic; Aortic Diseases; Arteriosclerosis; Calcimycin; Dose-Response Relationship, Drug; Endothelium, Vascular; Hypercholesterolemia; In Vitro Techniques; Lipoproteins, LDL; Male; Muscle Contraction; Muscle Relaxation; Nitric Oxide; Nitroglycerin; Norepinephrine; Rabbits

The present studies were performed to determine if abnormal endothelium-dependent vascular relaxation in atherosclerosis is due to decreased production or release of endothelium-derived relaxing factor (EDRF) by atherosclerotic rabbit vessels or if atherosclerotic vessels are less sensitive to the relaxing effects of EDRF. EDRF release was quantified using two approaches, by the response of bioassay detector vessels and also by the activation of guanylate cyclase within cultured endothelial cells. Using these assays, atherosclerotic vessels were found to release significantly less EDRF than normal vessels in response to both receptor- and nonreceptor-mediated stimuli. Relaxations of normal and atherosclerotic vessels to luminally applied EDRF (derived from normal rabbit aortas stimulated by the calcium ionophore, A23187) and nitric oxide, a putative EDRF, were also studied. Atherosclerotic vessels were more sensitive to EDRF than normal vessels, and equally sensitive to nitric oxide. Additional studies performed in organ chambers failed to demonstrate augmented constriction of atherosclerotic vessels in response to acetylcholine in the presence or absence of methylene blue or LY83583, compounds which inhibit the effect of EDRF. We conclude that decreased EDRF release is the principal underlying mechanism responsible for abnormal endothelium-dependent vascular relaxation in atherosclerosis.

Topics: Acetylcholine; Aminoquinolines; Animals; Aorta, Thoracic; Aortic Diseases; Arteriosclerosis; Biological Assay; Calcimycin; Enzyme Activation; Guanylate Cyclase; Humans; Methylene Blue; Nitric Oxide; Rabbits; Vasodilation

We evaluated the hypothesis that leukotriene (LT) B4, a potent chemotactic and endothelium-permeabilizing autacoid, may be a factor in the progression of the atherosclerotic lesion. Human vascular fragments, freshly obtained at vascular surgery from saphenous veins, atrial appendages, normal aortas and atherosclerotic lesions (histologically classified as fibrous plaques, fatty plaques and complicated lesions) were incubated at 37 degree C with mechanical agitation, for various times ranging between 5 and 60 minutes, sequentially, with buffer (basal), ionophore A23187 (IONO), and arachidonic acid (AA). After each step medium was assayed for LTB4 production by radioimmunoassay. Incubations were also performed in a chamber allowing selective exposure of the endothelial surface to the medium. Basal production (15 minute incubation, ng/g, mean +/- SD) was lowest for fibrous plaques (0.9 +/- 0.2, n = 38) and saphenous veins (1.0 +/- 0.3, n = 32) and much higher (P less than 0.01) for fatty plaques (7.3 +/- 2.2, n = 35) and complicated lesions (5.3 +/- 3.0, n = 28). A virtual abolition of production was observed after boiling of the vascular fragment. Production was increased in the presence of AA and IONO (in atheromas: 11.5 +/- 4.2 AND 14.3 +/- 5.3, respectively, with 15 minutes incubation), and decreased after incubation with 5-lipooxigenase inhibitors. Peak of ionophore or arachidonic acid-stimulated production in kinetic studies was reached at 20 minutes. LTB4 production was able to reach the luminal surface in both basal and stimulated conditions. There was no relationship with concomitantly measured prostacyclin production (mainly endothelial), while a correlation was found between LTB4 levels and degree of white cell infiltration in the tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Arachidonic Acid; Arachidonic Acids; Arteries; Arteriosclerosis; Calcimycin; Humans; In Vitro Techniques; Leukotriene B4; Muscle, Smooth, Vascular; Reference Values; Veins

The dependence of vascular relaxation on an intact endothelium and the relationship between relaxation and cyclic GMP accumulation were determined in coronary arteries isolated from cardiac transplantation patients with or without coronary atherosclerosis. In nonatherosclerotic arteries, the endothelium-dependent agent acetylcholine produced concentration-related relaxations. In atherosclerotic arteries, endothelium-dependent relaxations were abolished with acetylcholine, partly suppressed with substance P and histamine, and completely preserved with the ionophore A23187. In these arteries, the endothelium-independent agent nitroglycerin remained fully active. Accumulation of cyclic GMP in atherosclerotic strips was suppressed with acetylcholine but unattenuated with A23187 and nitroglycerin. In aortas from rabbits with diet-induced atherosclerosis, there was likewise an impaired cholinergic relaxation and cyclic GMP accumulation in the presence of preserved responses to A23187 and nitroglycerin. The results demonstrate that impaired cholinergic responses in atherosclerotic arteries reflect a muscarinic defect and not an inability of endothelium to release endothelial factor or smooth muscle to respond to it.

Topics: Acetylcholine; Animals; Aorta; Arteriosclerosis; Calcimycin; Coronary Vessels; Cyclic GMP; Endothelium; Histamine; Humans; Nitric Oxide; Nitroglycerin; Rabbits; Receptors, Muscarinic; Substance P; Vasodilation; Vasodilator Agents

We have previously found that vascular relaxation to acetylcholine and thrombin is markedly impaired in vessels from monkeys with diet-induced atherosclerosis. In the present study, we found that both normal and atherosclerotic vessels relaxed completely to the calcium ionophore A23187, which stimulates release of the endothelium-derived relaxing factor by nonreceptor-mediated mechanisms. Atherosclerotic vessels, however, were less sensitive to this agent. The finding that responses to the calcium ionophore were impaired in atherosclerosis suggests that abnormal endothelium-dependent relaxation in atherosclerotic vessels is not related entirely to alterations of thrombin and muscarinic receptors but may also be due to abnormal endothelium-derived relaxing factor production or transfer from the endothelium to the underlying vascular smooth muscle. Neither normal nor atherosclerotic iliac arteries constricted in response to acetylcholine when studied in the nonpreconstricted state. Constriction to acetylcholine in these vessels was not unmasked by removal of the endothelium. Thus, the smooth muscle of iliac vessels from monkeys contains few functioning muscarinic receptors. Impaired relaxation of atherosclerotic vessels to acetylcholine is not due to enhanced muscarinic-mediated constriction or to production of an endothelium-derived constricting factor. In vivo studies were performed to determine if alpha-adrenergic coronary vascular constriction is enhanced in the presence of atherosclerosis. In anesthetized monkeys, myocardial oxygen consumption was increased by two mechanisms, aortic occlusion and phenylephrine infusion. During both aortic occlusion and phenylephrine infusion, decreases in coronary vascular resistance were similar in control and atherosclerotic monkeys.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Acetylcholine; Animals; Aorta; Arteriosclerosis; Biological Products; Blood Vessels; Calcimycin; Cholesterol, Dietary; Coronary Vessels; Endothelium; Macaca fascicularis; Myocardium; Nitric Oxide; Oxygen Consumption; Phenylephrine; Receptors, Adrenergic, alpha; Vasoconstriction; Vasodilation