Effect of hypercholesterolemia on vascular reactivity in the rabbit. II. Influence of treatment with dipyridamole on endothelium-dependent and endothelium-independent responses in isolated aortas of control and hypercholesterolemic rabbits

The effects of cholesterol-feeding in the presence of dipyridamole (0.60 g daily) on contractile responses and on endothelium-dependent and endothelium-independent relaxations in isolated rabbit aortas are described. The investigations were performed simultaneously with those described in Part I (Circ Res 1986; 58:552-564), where the effects of cholesterol feeding on vascular reactivity in rabbit arteries (n = 8 in each group) selected at random from the same group of animals was studied. In the hypercholesterolemic rabbits treated with dipyridamole for 8 or 16 weeks, both the increases in plasma cholesterol and the formation of fatty streaks were significantly less pronounced than in the hypercholesterolemic rabbits not receiving the drug. Segments of the isolated arteries were mounted in organ chambers for isometric tension recording. The contractions caused by acetylcholine, prostaglandin F2 alpha, norepinephrine, clonidine, and serotonin and the endothelium-independent relaxations to nitroglycerin were not significantly altered by the hypercholesterolemia in rabbits treated with dipyridamole, even after 16 weeks of treatment. Thus, the decreased responses to norepinephrine, clonidine, and nitroglycerin and the augmented responses to serotonin noted in aortas of hypercholesterolemic rabbits in Part I were absent in the dipyridamole-treated hypercholesterolemic animals. The endothelium-dependent relaxations to ATP and acetylcholine were not affected after 8 weeks of hypercholesterolemia in presence of dipyridamole, while after 16 weeks the relaxations to ATP and acetylcholine were attenuated only in the more severely affected arteries. The effects of hypercholesterolemia + dipyridamole on endothelium-dependent relaxations were significantly less pronounced than those induced by hypercholesterolemia alone.(ABSTRACT TRUNCATED AT 250 WORDS)     

Vascular reactivity during the progression of atherosclerotic plaque. A study in Watanabe heritable hyperlipidemic rabbits

The effects of varying degrees of atherosclerotic plaque on vascular responsiveness in aortas of Watanabe heritable hyperlipidemic (WHHL) rabbits and New Zealand White (normal cholesterolemic) rabbits were studied. Ring segments from the aortic arch and thoracic aorta were mounted in organ chambers for isometric tension recording and measurement of endothelium-derived relaxing factor. WHHL rabbits were divided into three groups according to age: group 1, 3-5 months; group 2, 6-9 months; and group 3, 12-14 months. Atherosclerotic changes (expressed as a percent of total surface area) in the aortic arches in groups 1, 2, and 3 were 11 +/- 3% (mild), 28 +/- 6% (moderate), and 54 +/- 8% (severe) respectively; only occasional plaques were present in the thoracic aorta in all groups. Maximal contractions elicited with phenylephrine progressively decreased with increasing degrees of atherosclerotic plaque. Contractions evoked by histamine were augmented in all groups of WHHL rabbits when compared with controls, whereas those to serotonin were augmented only in vessels with mild atherosclerosis. As the severity of the intimal lesions increased, endothelium-dependent relaxations to acetylcholine, ATP, and calcium ionophore A23187 progressively decreased. Endothelium-independent relaxation to nitroglycerin was virtually complete in all segments. However, vessels with severe atherosclerosis were less sensitive to this agent as illustrated by a significant increase in the ED50 value. Scanning electron microscopy revealed a predominant loss of endothelial cells in the central regions of fibrous plaques. Thus, in WHHL rabbits, hypercholesterolemia and atherosclerosis result in an increased responsiveness of vascular smooth muscle to histamine and serotonin. Endothelium-mediated relaxation of vascular smooth muscle is reduced with the progression of atherosclerosis primarily due to a loss of endothelial cells.     

Mechanisms of abnormal endothelium-dependent vascular relaxation in atherosclerosis: implications for altered autocrine and paracrine functions of EDRF

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.     

Atherosclerosis impairs endothelium-dependent vascular relaxation to acetylcholine and thrombin in primates

To test the hypothesis that atherosclerosis impairs endothelium-dependent vascular relaxation, we examined the effect of the endothelium-dependent vasodilators acetylcholine and thrombin and the endothelium-independent vasodilator nitroglycerin on iliac arteries from normal cynomolgus monkeys and cynomolgus monkeys with diet-induced atherosclerosis. Rings of iliac artery were suspended in organ chambers at their optimal length for generating tension. After preconstriction with prostaglandin F2 alpha, cumulative concentration-response curves to acetylcholine, thrombin, and nitroglycerin were examined. The presence of endothelium was confirmed in each vessel by scanning electron microscopy. Atherosclerotic vessels showed morphologic evidence of moderate to severe atherosclerosis. Acetylcholine produced a maximal relaxation of 65 +/- 10% in the normal group and 27 +/- 10% in atherosclerotic vessels (P less than 0.05). Thrombin (10.0 U/ml) produced relaxation of 39 +/- 9% in the normal group and 13 +/- 7% in atherosclerotic iliac arteries (P less than 0.05). Nitroglycerin relaxed both normal and atherosclerotic blood vessels to an equal extent; maximal relaxation was 92 +/- 4% in normal vessels and 98 +/- 2% in atherosclerotic vessels. To determine if hypercholesterolemia alone produces an abnormality in endothelium-dependent relaxation, we performed two additional studies. First, because veins are exposed to hypercholesterolemia, but do not develop atherosclerosis, we studied relaxation responses to acetylcholine and thrombin in veins from normal monkeys and monkeys with diet-induced atherosclerosis. Veins from normal and atherosclerotic monkeys relaxed to a similar extent upon exposure to the endothelium-dependent vasodilators acetylcholine and thrombin.(ABSTRACT TRUNCATED AT 250 WORDS)     

Dietary antioxidants preserve endothelium-dependent vessel relaxation in cholesterol-fed rabbits.

Recent evidence suggests that dietary therapy with lipid-soluble antioxidants may be beneficial for patients with atherosclerotic vascular disease but the potential mechanism(s) for these observations remain obscure. Abnormalities in endothelium-dependent control of vascular tone develop early in the course of atherosclerosis and may result from oxidative modification of low density lipoproteins. We examined the role of dietary antioxidants in preserving normal endothelial cell vasodilator function in cholesterol-fed rabbits with particular attention to possible effects on serum lipoproteins, low density lipoprotein oxidation, and atherogenesis. Male New Zealand White rabbits were fed diets containing no additive (controls), 1% cholesterol (cholesterol group), or 1% cholesterol chow supplemented with either beta-carotene (0.6 g/kg of chow) or alpha-tocopherol (1000 international units/kg of chow) for a 28-day period. After dietary therapy, thoracic aortae were harvested for assay of vascular function and for pathologic examination and tissue antioxidant levels. Compared to controls, acetylcholine- and A23187-mediated endothelium-dependent relaxations were significantly impaired in vessels from the cholesterol group (P < 0.001), whereas vessels from animals treated with beta-carotene or alpha-tocopherol demonstrated normal endothelium-dependent arterial relaxation. Preservation of endothelial function was associated with vascular incorporation of alpha-tocopherol and beta-carotene but was unrelated to plasma lipoprotein levels, smooth muscle cell function, or the extent of atherosclerosis. Increased low density lipoprotein resistance to ex vivo copper-mediated oxidation was observed only in the alpha-tocopherol group. Our results suggest that dietary antioxidants may benefit patients with atherosclerosis by preserving endothelial vasodilator function through a mechanism related to vascular tissue antioxidant content and not reflected by assay of low density lipoprotein resistance to ex vivo oxidation.     

Arginine restores cholinergic relaxation of hypercholesterolemic rabbit thoracic aorta

BACKGROUND. Reduced synthesis of endothelium-derived relaxing factor (EDRF) may explain impaired endothelium-dependent vasodilation in hypercholesterolemia. Accordingly, we designed studies to determine if endothelium-dependent relaxation in hypercholesterolemic rabbits may be restored by supplying L-arginine, the precursor of EDRF. METHODS AND RESULTS. Normal or hypercholesterolemic rabbits received intravenous L-arginine (10 mg/kg/min) or vehicle for 70 minutes. Subsequently, animals were killed, thoracic aortas were harvested, and vascular rings were studied in vitro. Rings were contracted by norepinephrine and relaxed by acetylcholine chloride or sodium nitroprusside. Vasorelaxation was quantified by determining the maximal response (expressed as percent relaxation of the contraction) and the ED50 (dose of drug inducing 50% relaxation; expressed as -log M). In vessels from hypercholesterolemic animals receiving vehicle, there was a fivefold rightward shift in sensitivity to acetylcholine compared with normal animals (p = 0.05, n = 5 in each group). In vessels from hypercholesterolemic animals, L-arginine augmented the maximal response to acetylcholine (83 +/- 16% versus 60 +/- 15%, p = 0.04 versus vehicle) and increased the sensitivity to acetylcholine (ED50 value: 6.7 +/- 0.2 versus 6.2 +/- 0.2, p less than 0.05 versus vehicle). Arginine did not affect maximal and EC50 responses to acetylcholine in vessels from normal animals. Arginine did not potentiate endothelium-independent responses in either group. CONCLUSIONS. We conclude that the endothelium-dependent relaxation is normalized in hypercholesterolemic rabbit thoracic aorta by in vivo exposure to L-arginine, the precursor for EDRF.     

Putative mechanisms of the impairment of endothelium-dependent relaxation of the aorta with atheromatous plaque in heritable hyperlipidemic rabbits

Attenuation of acetylcholine-induced endothelium-dependent relaxation of thoracic aortas excised from Watanabe heritable hyperlipidemic (WHHL) rabbits linearly correlated with the percent area coated with atheromatous plaque. To elucidate mechanisms related to this reduced endothelium-dependent relaxation in the presence of atherosclerosis, the acetylcholine-induced release of endothelium-derived relaxing factor (EDRF) was assessed functionally as a percent relaxation of the precontracted detector strips obtained from the tunica media beneath the intact intima or the atheromatous plaque in the same aortic ring preparation. Relaxations of the normal detectors to effluents containing EDRF of thoracic aortas during stimulation by acetylcholine (3 x 10(-6) M) in heterozygous and homozygous WHHL rabbits were 73 +/- 5% and 59 +/- 9% (p less than 0.01) of the phenylephrine-induced precontraction, respectively. Relaxations of the atherosclerotic detectors to effluents (EDRF) through the aortas during stimulation by acetylcholine (3 x 10(-6) M) in heterozygous and homozygous WHHL rabbits were 16 +/- 4% and 14 +/- 5%, respectively--values significantly smaller than those seen in the normal detectors. When superoxide dismutase was added to the perfusate of the donors from homozygous and heterozygous WHHL rabbits, atherosclerotic detectors relaxed by effluents stimulated by acetylcholine to 73% and 65% (p less than 0.01 versus before the addition of superoxide dismutase) of the normal detector, respectively. Relaxations induced by sodium nitroprusside as well as the contractions by acetylcholine, phenylephrine, and KCl (118 mM) were comparable in detector strips from the normal and atherosclerotic portions. Thus, not only is the amount of EDRF released by acetylcholine reduced in the presence of atherosclerosis, the tunica media beneath the atheromatous plaque is also to some extent responsible for the superoxide-induced inactivation of EDRF.     

Effect of hypercholesterolemia on vascular reactivity in the rabbit. I. Endothelium-dependent and endothelium-independent contractions and relaxations in isolated arteries of control and hypercholesterolemic rabbits

We studied the effects of hypercholesterolemia on vascular responsiveness in different arteries isolated from rabbits: control groups of rabbits and groups receiving the atherogenic diet consisted of eight animals each. In the arteries, 16 weeks of cholesterol-rich (0.3%) diet evoked intimal lesions which were more pronounced than those noted after 8 weeks of hypercholesterolemia; the aortic arch was affected significantly more by the lesions than the abdominal aorta and the pulmonary artery. Segments of the arteries were mounted in organ chambers for isometric tension recording or for measurement of the endothelium-derived relaxant factor. Contractions caused by acetylcholine and prostaglandin F2 alpha were not altered by the hypercholesterolemia; those evoked by serotonin were moderately augmented only in the aortic arch of hypercholesterolemic rabbits. As the degree of intimal lesion formation increased, the contractions to norepinephrine and clonidine were progressively inhibited. The endothelium-independent relaxations to nitroglycerin were inhibited in only the most severely affected arteries; the endothelium-dependent relaxations to acetylcholine and adenosine triphosphate were progressively inhibited as the degree of fatty streak formation augmented. Thus, in the aortic arch, the relaxations to 3 X 10(-6) M acetylcholine, expressed as percent of the initial contraction, decreased from 86.7 +/- 3.3% in control tissues to 16.3 +/- 8.6% in the 16-week hypercholesterolemic vessels; in the abdominal aortas these relaxations averaged 93.5 +/- 2.2% in control vessels and 72.0 +/- 6.9% in the hypercholesterolemic tissues. The acetylcholine-induced release of endothelium-derived relaxant factor from the abdominal aorta was not significantly affected by the hypercholesterolemia. We conclude from these studies that in arteries obtained from hypercholesterolemic rabbits: the contractions caused by serotonergic mechanisms tend to be augmented, while those to alpha-adrenergic activation are decreased, the endothelium-independent relaxations are modified only in the more severely affected arteries, and the endothelium-dependent relaxations are progressively inhibited as the degree of fatty streak formation augments, probably because a step subsequent to the release of endothelium-derived relaxant factor is altered.     

Endothelium-dependent vascular relaxation is abnormal in the coronary microcirculation of atherosclerotic primates

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.     

Decreased endothelium-dependent relaxation in New Zealand genetic hypertensive rats

The relaxation response to endothelium-dependent (acetylcholine and the calcium ionophore A23187) and independent (sodium nitroprusside) vasodilators was examined in isolated aortic ring segments from age-matched genetically hypertensive (GH) and normotensive (N) rats (New Zealand strain). Tissues were initially contracted with methoxamine to achieve similar levels of contractile force. The IC20, IC40 and IC50 values for acetylcholine, A23187 and sodium nitroprusside were shifted significantly to the right (P less than 0.05) in aortic rings from GH rats compared to the corresponding values in N rats. The maximal relaxation achieved by acetylcholine and A23187 was significantly depressed in aortas from GH rats (P less than 0.05). Sodium nitroprusside elicited the maximal relaxation in both groups of tissues. These results demonstrate that there exists a generalized defect in the relaxant ability of vascular smooth muscle from GH rats. In addition, our findings suggest that this defect is coupled with a decreased responsiveness to endothelium-dependent vasodilators in this particular animal model of hypertension.     

Segmental impairment of endothelium-mediated relaxation in thoracic aortas from atherosclerotic rabbits. Comparison to cholesterol infiltration and energy metabolism

Three sets of parameters, (i) relaxation to acetylcholine (Ach), ATP and NaNO2, (ii) cholesterol content in aortic tissue, and (iii) energy metabolism were compared in normal and atherosclerotic rabbits, fed 1% cholesterol for eight weeks. A special protocol was envisaged to permit a strict comparison between Ach, ATP and NaN O2 at different levels of thoracic aorta, in each rabbit. A gradual impairment of the endothelium-dependent relaxation to Ach and ATP was found at different levels of the thoracic aorta from hypercholesteromic rabbits. By contrast, NaNO2--endothelium-independent--maintained its relaxing power quite normally at all aortic levels. A close correlation was evident between the impairment of aorta relaxation to Ach and the cholesterol infiltration in the vessel wall, being the correlation coefficient -0.85 (P less than 0.001). A correlation was also evident for ATP, but to a lower degree, being the correlation coefficient -0.61 (P less than 0.01). Energy metabolism and related parameters (ATP, ADP, AMP, adenosine, inosine, GTP, GDP, guanosine, NAD, NADP, total adenylate nucleotides and adenylate energy charge) were not modified by the cholesterol diet. These data show that the gradual impairment of endothelium-dependent relaxation, decreasing down the thoracic aorta of hypercholesterolemic rabbits, and correlated with cholesterol content in the aortic wall, may be considered as an index of a very early atherogenic damage, prior to variation in the parameters of energy metabolism and purine turnover.     

Impaired endothelium-dependent relaxation to aggregating platelets and related vasoactive substances in porcine coronary arteries in hypercholesterolemia and atherosclerosis

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.     

Preservation of endothelium-dependent vascular relaxation in cholesterol-fed rabbit by treatment with the calcium blocker PN 200110

We tested the effects of low doses of a dihydropyridine calcium antagonist, PN 200110, on endothelium-dependent vascular relaxation in rabbits fed a 1% cholesterol diet. The drug was given orally, 1 mg/day, and control rabbits received placebo. Plasma total cholesterol after 10 weeks, was similar in the placebo- and PN 200110-treated groups. The respective values averaged 2140 +/- 116 (n = 14; mean +/- SEM) and 2012 +/- 115 mg/dl (n = 13). In placebo-treated rabbits, sudanophilic aortic lesions covered 52 +/- 5% of the intimal surface, and the aortic cholesterol concentration was 72 +/- 6 mg/g protein. Corresponding values in aortas from PN 200110-treated rabbits were significantly lower [36 +/- 5% (P less than 0.03) and 52 +/- 3 mg/g protein (P less than 0.03)]. Maximal endothelium-dependent cholinergic relaxation of aortic strips in untreated (n = 14) and treated cholesterol-fed rabbits (n = 13) differed significantly (P less than 0.01) and averaged 31 +/- 4% and 61 +/- 7% of the value in normocholesterolemic controls (n = 13). We conclude that cholesterol feeding suppresses endothelium-dependent relaxation evoked by acetylcholine, and that PN 200110 reduces the severity of atherosclerosis and impairment of endothelium-dependent relaxation.     

Effect of lysophosphatidylcholine on atherosclerotic rabbit arteries

We report here on the effect of an endothelium-dependent vascular smooth muscle relaxant, lysophosphatidylcholine (LPC) on rabbit aortic strips and on hemodynamic changes by LPC in atherosclerotic animals. Cyclic GMP changes induced by LPC in atherosclerotic vessels were also determined. Atherosclerosis was produced by feeding a high cholesterol and saturated fatty acid diet. LPC was injected into the left atrium and coronary flow was measured by radioactive microspheres; in vitro, relaxation of precontracted aortic strips by lysophosphatidylcholine was also recorded. LPC failed to increase coronary flow in the presence of atherosclerosis. In isolated aortic strips, dose-response curves with acetylcholine and LPC showed diminished relaxation in atherosclerotic preparations, and cyclic GMP production following LPC was reduced. The results demonstrate that vascular relaxation by LPC, together with its ability to activate guanylate cyclase is dependent on the functional and morphological integrity of the vascular wall.     

Endothelium-dependent responses in isolated blood vessels of lower vertebrates

Endothelium-dependent relaxations to acetylcholine have been identified in mammalian arteries and veins. To determine the occurrence of such relaxations in other classes of vertebrates, rings of descending aortas of turtles, cayman and bullfrogs and ventral aortas of trout were suspended for isometric force measurements. Acetylcholine and the calcium ionophore A23187 initiated concentration-dependent relaxations in aortas from cayman and bullfrogs contracted with norepinephrine. These relaxations were not affected by meclofenamate, were reversed by methylene blue and abolished by endothelium removal. Acetylcholine caused concentration-dependent contractions in aortas (with and without endothelium) from trout and turtles; these tissues contracted minimally to norepinephrine. In the aortas of the trout contracted with acetylcholine, the calcium ionophore A23187 initiated endothelium-dependent relaxations which were reversed by methylene blue and abolished by meclofenamate. A23187 contracted turtle aortas; an effect reduced by endothelium removal. These data demonstrate endothelium-dependent relaxations and contractions in blood vessels of reptiles, amphibians and teleosts. Thus, endothelium-dependent modulation of the responses of the vascular smooth muscle represents a cardiovascular regulatory mechanism which appears early in vertebrate phylogeny.     

A triglyceride-rich fat emulsion and free fatty acids but not very low density lipoproteins impair endothelium-dependent vasorelaxation.

OBJECTIVE: To investigate the effects of triglycerides and free fatty acids on endothelium-dependent and endothelium-independent vasorelaxation. METHODS: Femoral arterial rings from rats were studied in organ baths. The vascular segments were constricted with phenylephrine after 20 min of preincubation with the triglyceride-rich fat emulsion Intralipid, free fatty acids (16:0, 18:1, 18:3) bound to bovine serum albumin, or very low density lipoproteins. Endothelium-dependent and endothelium-independent relaxations were determined after administration of acetylcholine and nitric oxide donors, respectively. RESULTS: Preincubation with Intralipid caused a concentration-dependent impairment of endothelium-dependent but not endothelium-independent relaxation. Very low density lipoproteins did not affect vascular function. All free fatty acids impaired endothelium-dependent relaxation, whereas endothelium-independent relaxation was unaffected. Administration of the antioxidant vitamin C partly reversed the impairment of the endothelium-dependent relaxation induced by Intralipid and free fatty acids. CONCLUSIONS: The present study demonstrates that the triglyceride-rich fat emulsion Intralipid and individual FFAs impair endothelium-dependent relaxation of arterial rings from rat, whereas triglycerides in the form of VLDL do not affect endothelial function. The finding that the antioxidant vitamin C partly reverses this impairment indicates the involvement of oxidative mechanisms.     

Endothelium-dependent relaxation in experimental atherosclerosis in the rabbit

The effect of feeding a diet supplemented with lipids and containing 2% cholesterol on the endothelium-dependent relaxation of rabbit aorta to acetylcholine was assessed. The effect of feeding a standard rabbit diet after an initial period of 2% cholesterol feeding was assessed also. Age-matched male, New Zealand white rabbits were fed either a 2% cholesterol diet or a standard rabbit diet. The animals were anesthetized with pentobarbitone sodium (25 mg/kg) and killed either at the beginning of the study (0 weeks) or at 4, 8, or 10 weeks. The animals in the reversal study were fed the 2% cholesterol diet for 6 weeks and killed after an additional 14 and 32 weeks on standard diet. The extent of atherosclerosis in the aorta was assessed by Sudan Red staining, estimation of tissue cholesterol, and light and electron microscopy. The relaxation response to acetylcholine was measured in rings of the thoracic aorta following precontraction with norepinephrine (-6.0 log mol/l). The relaxation was significantly impaired in aortas from rabbits fed the 2% cholesterol diet compared to aortas from animals fed the standard diet. The impairment of relaxation was apparent as early as 4 weeks after the start of the 2% cholesterol diet and remained impaired over the next 6 weeks. No improvement in endothelium-dependent relaxation was seen in rabbits on the reversal diet for 14 and 32 weeks. Thus, endothelium-dependent relaxation is attenuated in animals fed a 2% cholesterol diet, and the loss of relaxation persists for at least 32 weeks after the animals are returned to a standard diet.     

精氨酸对高胆固醇血症兔冠状动脉功能的效应

目的评价左-精氨酸对高胆固醇血症兔模型冠状动脉内皮舒张功能的效应。方法雄性日本大耳白兔36只随机分为3组:对照组、高胆固醇血症组、左-精氨酸组,每组12只。两周后,处死兔,取兔冠状动脉做成动脉环。结果与对照组比较,高胆固醇血症组、左-精氨酸组兔胆固醇水平明显增高;高胆固醇血症组兔冠状动脉环在KCl预收缩基础上对内皮依赖的(乙酰胆碱累积浓度10-8～10-4mol/L)效应减弱;左-精氨酸组兔对乙酰胆碱的舒血管反应明显减低,在高浓度(乙酰胆碱10-4mol/L)下表现为冠状动脉环的收缩(P<0.05)。结论左-精氨酸不能改善高胆固醇血症兔冠状动脉的内皮舒张功能。     

High sodium intake increases vascular superoxide formation and promotes atherosclerosis in apolipoprotein E-deficient mice

Hypertension is a major risk factor for atherosclerosis. We tested the hypothesis whether high salt intake aggravates endothelial dysfunction and promotes atherosclerosis in apolipoprotein E-deficient mice (ApoE^-/^- mice) and their littermate controls (C57Bl/6 mice). The role of increased oxidative stress was also examined. A high-salt diet (NaCl 7%) for 12 weeks increased blood pressure and induced cardiac hypertrophy and albuminuria more pronouncedly in ApoE^-/^- mice compared with C57Bl/6. Endothelium-dependent vascular relaxation in response to acetylcholine was almost maximally impaired in ApoE^-/^- mice during a normal sodium diet. A high-salt diet did not further impair NO-mediated vascular relaxation. A high-salt diet also markedly attenuated endothelium-dependent relaxation in C57Bl/6 mice. Preincubation with the superoxide scavenger Tiron normalized endothelial function almost completely in both mice strains indicating the central role of increased oxidative stress in the pathogenesis. Aortic superoxide production and the extent of atherosclerotic lesions were greater in ApoE^-/^- mice on a normal-salt diet compared with C57Bl/6. The high-salt diet increased vascular superoxide formation and promoted atherosclerosis in ApoE^-/^- mice. Changes in dietary salt intake did not influence serum lipids in either mouse strains. Our findings suggest a detrimental role for high salt intake in the development of atherosclerosis and underscore the importance of increased oxidative stress in the pathogenesis salt-induced vascular damage.     

Low level hyperlipidemia impairs endothelium-dependent relaxation of porcine coronary arteries by two mechanisms. Functional change in endothelium and impairment of endothelium-dependent relaxation by two mediators.

We evaluated the effect of a low level of hyperlipidemia and the effects of in vitro exposure to atherogenic lipoproteins (LDL, VLDL) on the vascular responsiveness of isolated porcine coronary arteries. Firstly we studied the change in vascular responsiveness induced by feeding a cholesterol-rich diet to pigs for 4 and 9 weeks (C4 and C9 pigs). The serum cholesterol level in pigs fed a cholesterol-rich diet reached 218.5 +/- 32.9 mg/dl compared with 85.5 +/- 8.4 mg/dl in the controls. Segments of the left descending coronary artery were examined. The contraction induced by KCl or prostaglandin F2 alpha was not altered significantly by hypercholesterolemia nor was the relaxation induced by the Ca2+ ionophore, A23187, or by nitroglycerin. Endothelium-dependent relaxation (EDR) evoked by high, but not low, concentrations of bradykinin was reduced in the C4 pigs as compared with those in normal animals. EDRs evoked by bradykinin, substance P, and serotonin were significantly reduced in C9 pigs. Histologically, as observed by light and electron microscopy, fatty changes or intimal thickenings were not seen in the coronary arteries of the C4 pigs. Minimal changes (intimal thickening and fragmentation of internal elastic lamina) were observed only in parts of arteries of the C9 pigs. Secondly, the direct effects of LDL and VLDL on vascular responsiveness were studied. Although preincubation with LDL inhibited the EDR caused by exposure to bradykinin and A23187 in the coronary arteries of normal and cholesterol-fed pigs, preincubation with LDL inhibited the arterial relaxation induced by exposure to substance P or serotonin in both the C4 and the C9 pigs, but not in the control animals. The degree of inhibition was especially marked in the C9 pigs. The inhibitory effect of VLDL on EDR was weaker than that of LDL. Indomethacin (5 microM) did not alter this inhibitory effect of lipoproteins. Neither LDL nor VLDL had any effect on the vascular relaxation induced by nitroglycerin. These results are consistent with the idea that endothelium-dependent arterial relaxation is attenuated even at the very early stage of cholesterol-induced atherosclerosis. Atherogenic lipoproteins may further impair the decreased EDR in the arteries of hyperlipidemic pigs by two factors: one released on stimulation with bradykinin and the calcium ionophore A23187, the other released on stimulation with substance P and serotonin.