Cholesterol feeding increases plasma and aortic tissue cholesterol oxide levels in parallel: further evidence for the role of cholesterol oxidation in atherosclerosis.

To determine the relationship between plasma and arterial wall oxysterols, plasma and aortic tissue from 7 New Zealand White rabbits fed a high cholesterol (1%) diet for 6 weeks was compared to plasma and aortic tissue from 7 normocholesterolemic rabbits fed standard rabbit chow. Cholesterol and cholesterol oxide fractions were isolated and analyzed by gas chromatography. Normocholesterolemic plasma and aortic tissue contained low levels of cholest-5-ene-3 beta, 7 alpha-diol, cholesta-3,5-dien-7-one, 5,6 alpha-epoxy-5 alpha-cholestan-3 alpha-ol, cholest-5-ene-3 beta, 7 beta-diol, and 5 alpha-cholestane-3 beta, 5,6 beta-triol while hypercholesterolemic plasma and atherosclerotic aorta contained significantly higher levels (P less than 0.05) of these products. Furthermore, 5,6 beta-epoxy-5 alpha-cholestan-3 beta-ol not found in normocholesterolemic plasma or aortic tissue was present in substantial amounts in both hypercholesterolemic plasma and atherosclerotic aortic tissue. Cholest-5-ene-3 beta,25-diol and 3 beta-hydroxycholest-5-ene-7- one not present in normocholesterolemic aorta were present in the atherosclerotic aorta. The oxysterol chromatographic patterns of normocholesterolemic plasma and normocholesterolemic aortic tissue were similar to each other as were the oxysterol chromatographic patterns of hypercholesterolemic plasma and atherosclerotic aortic tissue. The chromatographic patterns between the normocholesterolemic and hypercholesterolemic samples differed however. Possible absorption of the low levels of cholesterol oxides present in the cholesterol feed could account for the elevation of only some of the oxysterols. We conclude that cholesterol oxides exist at some basal level in normocholesterolemia and that these levels are increased by cholesterol-feeding which results in hypercholesterolemia. Our findings demonstrate that there is a strong relationship between plasma and aortic arterial wall levels of cholesterol oxides and suggest that in addition to exogenous sources, formation of cholesterol oxides proceeds via free radical oxidation acting upon elevated cholesterol levels resulting in the accumulation of these potentially cytotoxic and atherogenic products.     

A new HMG-CoA reductase inhibitor, pitavastatin remarkably retards the progression of high cholesterol induced atherosclerosis in rabbits

BACKGROUND: The remarkable anti-atherosclerotic effects of 3-hydroxy-3-methyl-glutaryl-CoA reductase inhibitor have not been demonstrated in diet induced severe hyperlipidemia in rabbit model. OBJECTIVE: We have investigated the effect of pitavastatin, a newly developed statin, on atherosclerosis in rabbits. METHODS AND RESULTS: Oophorectomized female NZW rabbits were fed 0.3% cholesterol chow for 12 weeks with or without pitavastatin (0.1mg/kg per day) (Gp.NK and HCD). The level of serum cholesterol was decreased in Gp.NK compared with Gp.HCD (772.8 +/- 70.2 versus 1056.9 +/- 108.3 mg/d), whereas no significant alterations were observed in triglyceride and HDL-cholesterol. NO dependent response stimulated by acetylcholine and calcium ionophore A23187 and tone related basal NO response induced by N(G)-monomethyl-l-arginine acetate were all improved by pitavastatin treatment. Pitavastatin treatment increased the level of cyclic GMP in the aorta of cholesterol fed rabbits. In the aorta, the expression of eNOS mRNA was significantly up regulated and O(2)(-) production was slightly reduced in Gp.NK animals. Atherosclerotic area was significantly decreased in aortic arch and thoracic aorta from Gp.NK compared with those from Gp.HCD ( 15.1 +/- 5.3 versus 41.9 +/- 10.2%, 3.1 +/- 1.1versus 7.9 +/- 1.2% in Gp.NK and Gp.HCD aortic arch and thoracic aorta). Anti-macrophage staining area, the MMP1 or 2 and the nitrotyrosine positive area were decreased in Gp.NK. CONCLUSION: Pitavastatin retards the progression of atherosclerosis formation and it improves NO bioavailability by eNOS up-regulation and decrease of O(2)(-).     

Progression of induced aortic atherosclerosis in rabbits fed a high cholesterol diet

In this study we have evaluated the progression of atheromatosis in aortic arch, thoracic aorta and abdominal aorta in rabbits fed a high cholesterol diet. The aortic atheromatosis decreased progressively from the aortic arch to the abdominal aorta after 6 months of high cholesterol diet. It is possible that a different segmental resistance to hypercholesterolemic damage may be involved in the cranio caudal progression of atheromatosis in rabbits.     

Hypocholesterolemic and antiatherosclerotic effect of flax lignan complex isolated from flaxseed

Hypercholesterolemia, low HDL-C and oxygen radicals have been implicated in the development of atherosclerosis. Lignan complex isolated from flaxseed contains secoisolariciresinol diglucoside (SDG), 3-hydroxy-3methylglutaric acid (HMGA) and cinnamic acids. SDG and cinnamic acids are antioxidants, and HMGA is a hypocholesterolemic agent. Antioxidants are known to reduce hypercholesterolemic atherosclerosis. The objectives of this study were to determine if lignan complex reduces (i) serum cholesterol, (ii) oxidative stress, and (iii) atherosclerosis in hypercholesterolemic rabbits. Rabbits were assigned to four groups: Group I, control; Group II, lignan complex control (lignan complex, 40 mg/kg body weight daily orally); Group III, 0.5% cholesterol; Group IV, 0.5% cholesterol diet+lignan complex, (40 mg/kg body weight daily orally). Blood samples were collected before (time 0) and after 1 and 2 months of experimental diets for measurement of serum triglycerides (TG), total cholesterol (TC), LDL-C, HDL-C and serum malondialdehyde (MDA), a lipid peroxidation product. At the end of the protocol, the aorta was removed for measurement of atherosclerotic plaques, MDA and aortic tissue chemiluminescence (Aortic CL), a marker of antioxidant reserve. Rabbits in Group III developed atherosclerosis (50.84+/-6.23% of the intimal surface of the aorta was covered with atherosclerotic changes) which was associated with an increase in the serum TG, TC, LDL-C, HDL-C, MDA and aortic MDA and antioxidant reserve. Lignan complex reduced the development of atherosclerosis by 34.37% and this was associated with a decrease in serum TC by 20%, LDL-C by 14%, TC/HDL-C by 34%, serum MDA by 35% and aortic MDA by 58%. Serum HDL-C was elevated by 30% in hypercholesterolemic rabbits and by 25% in normocholesterolemic rabbits with lignan complex. Lignan complex did not affect the TC and LDL-C and serum MDA in the normocholesterolemic rabbits. However, it increased the aortic MDA in the normocholesterolemic rabbits. These results suggest that lignan complex isolated from flaxseed reduced the extent of hypercholesterolemic atherosclerosis and this effect was associated with marked decreases in oxidative stress, serum total cholesterol, LDL-C and risk ratio, and elevation of serum HDL-C. Lignan complex may, therefore, be beneficial in preventing atherosclerosis, and reducing risk factors for coronary artery disease and stroke.     

Effect of 17-beta estradiol on pre-existing atherosclerotic lesions: role of the endothelium.

The atheroprotective effects of estrogen during the process of atherogenesis is well documented, whereas limited information is available about the effect of estrogen on pre-existing atherosclerotic lesions. After bilateral ovariectomy, 24 New Zealand White rabbits were randomized into three groups of eight animals each and subsequently fed a 0.5% cholesterol diet. In group I, the vessels were excised at day 84, whereas in group II, the cholesterol diet was continued for a total of 168 days. In group III, the animals were first fed with a cholesterol diet for 84 days; in the second phase of the experiment, the cholesterol diet was continued for a further 84 days with a combined estrogen treatment (1 mg estradiol valerate per kg body weight per week intramuscularly). At the end of the experiment, the proximal aortic arch, right carotid artery, thoracical aorta and abdominal aorta of each animal were excised and prepared for histological and immunohistological examination. By day 168, morphometrical analysis displayed a significantly lower plaque development under estrogen therapy in the carotid artery (0.08+/-0.18 mm(2) vs. 0.60+/-0.39 mm(2)), the thoracic aorta (0.56+/-0.94 mm(2) vs. 3.63+/-2.06 mm(2)), and in the abdominal aorta (0.55+/-0.70 mm(2) vs. 1.71+/-1.05 mm(2)) in comparison with the corresponding 168 day control group. However, estrogen treatment has failed to reduce further atherosclerotic plaque development in the aortic arch (9.42+/-1.79 mm(2) vs. 11. 64+/-3.29 mm(2)). Immunohistological detection of the 'anti-human factor VIII related antigen', i.e. the 'von Willebrand factor' (vWF), showed a significantly lower number of luminal cells positive for vWF in the aortic arch in the 84-day cholesterol group, compared with the corresponding controls of normocholesterolemic rabbits (65. 9+/-12.4% vs. 83.1+/-6.2%; P<0.05). Estradiol was able to inhibit the further progression of atherosclerosis when moderate vessel wall alterations were present, whereas pre-existing severe atherosclerosis was associated with a failure of the anti-atherosclerotic estrogen action. As suggested by the in situ detection of vWF as a morphological marker for endothelial cells, an intact endothelial layer might play an important role in mediating the beneficial effect of estrogen in the process of atherosclerosis.     

雌激素对高胆固醇兔一氧化氮及内皮素生成的影响

观察雌激素对去卵巢高胆固醇兔动物模型主动脉一氧化氮及血浆内皮素-1的影响,以阐明雌激素抗动脉粥样硬化的部分机理.将21只纯种新西兰雌兔随机分入3组(各7只):A组(去卵巢未补充激素兔),B组(去卵巢补充雌激素兔),C组(未切除卵巢兔),都给予1%胆固醇饮食,8周后取主动脉应用Greiss试剂测定不同浓度的乙酰胆碱作用下主动脉NO代谢物NO_2~-及血浆内皮素的量.10~(-5)乙酰胆碱作用下,B、C组主动脉NO_2~-产量明显低于A组P<0.05),B、C组兔主动脉反应性高,10~(-5)乙酰胆碱作用下NO_2~-产量明显高于同组无乙酰胆碱时NO_2~-产量,A组即使在10~(-5)乙酰胆碱作用下,NO_2~-无明显增高.A组内皮素明显高于B组(P<0.05)及C组(P<0.01).雌激素在整体水平促进NO产生,并抑制内皮素-1生成,可能为抑制动脉粥样硬化形成的重要机理.     

Protective role of heparin on in vitro functional aortic response in Watanabe heritable hyperlipidemic rabbits.

The effects of prolonged in vivo heparin treatment upon vasomotor responses and content of cholesterol and energy related compounds were studied in isolated thoracic and abdominal aortas from Watanabe heritable hyperlipidemic (WHHL) rabbits. Unfractionated heparin was administered subcutaneously (2 mg/kg twice a day) to 3-month-old WHHL rabbits for a period of 6 months. A group of WHHL rabbits was treated with saline solution and considered as control. Aortic cholesterol infiltration and serum cholesterol were not significantly decreased by the prolonged heparin treatment. In heparin-treated WHHL rabbits, the in vitro aortic endothelium-dependent relaxation produced by acetylcholine or calcimycin (A 23187) was greater than in saline-treated WHHL group. ATP-induced aorta relaxation (endothelium-dependent and endothelium-independent) did not vary significantly in the two groups of WHHL rabbits, even after mechanical removal of endothelium. Also the noradrenaline-induced aorta contraction did not vary between the two groups of WHHL rabbits. No significant variation in energy-related compounds (except for ADP) was found in the aortic arch. These results suggest that heparin produces a protective effect on aortic tissue by acting mainly at endothelial level.     

Effect of HMG-CoA reductase inhibition on endothelial dysfunction-inducing protein in hypercholesterolemic rabbits

INTRODUCTION AND OBJECTIVES: In our laboratory, we recently obtained evidence that cultured bovine endothelial cells contain cytosolic proteins that form complexes with the 3'-unstranslated region of endothelial nitric oxide synthase (eNOS) mRNA and are associated with its destabilization. The aim of this study was to determine the presence of such proteins and the level of eNOS expression in hypercholesterolemic rabbits as an in vivo model of endothelial dysfunction. METHODS AND RESULTS: Endothelium-dependent relaxation in response to acetylcholine was reduced in aortic segments from hypercholesterolemic rabbits compared with controls. Treatment of hypercholesterolemic rabbits with simvastatin (25 mg/kg body weight/day) restored endothelium-dependent relaxation. Aortic eNOS expression was reduced in hypercholesterolemic rabbits and was accompanied by enhanced binding activity of a 60-KDa cytosolic protein and reduced stability of eNOS mRNA. Simvastatin treatment upregulated eNOS expression and reduced the interaction of cytosolic protein with the 3'-untranslated region of eNOS mRNA. CONCLUSIONS: These results demonstrate the presence of a 60-KDa protein that binds to eNOS mRNA and reduces eNOS expression in the vascular wall.     

老年大鼠主动脉对胰岛素敏感性的改变及其机制

目的观察大鼠主动脉对胰岛素敏感性的增龄改变并分析其可能机制。方法老年组采用老年（18月龄）SD大鼠20只,随机选取成年（15周龄）SD大鼠20只为对照组。采用离体血管灌流技术,观察胸主动脉对胰岛素反应性的变化,并同时测定两组主动脉血管一氧化氮（NO）释放量及血管一氧化氮合酶（eNOS）活性;免疫组织化学法及Western Blot法检测老年组及成年组胸主动脉eNOS的蛋白表达变化。结果胰岛素可以浓度依赖性舒张成年大鼠胸主动脉,舒血管作用具有内皮依赖性。与之相比,老年大鼠主动脉对胰岛素的舒张反应显著下降（P<0.05）。同时发现,与成年组相比,老年组NO释放量以及eNOS活性显著降低（P<0.05）,免疫组织化学染色显示老年组主动脉eNOS的蛋白表达显著降低;而Western Blot检测发现老年组血管eNOS磷酸化水平显著降低（P<0.05）。结论血管组织内源性eNOS-NO系统活性下降可能是衰老导致的血管胰岛素敏感性下降的重要机制。     

Effects of estradiol and progesterone on the increased synthesis of collagen in atherosclerotic rabbit aortas

The effects of sex hormones on atherosclerosis and collagen synthesis in blood vessels were studied in rabbits fed an atherogenic diet. Ovariectomy without hormone replacement resulted in significantly greater degree of atherosclerosis and collagen synthesis in the aortic arch when compared with the intact state. The administration of estradiol to ovariectomized rabbits resulted in a degree of atherosclerosis and collagen synthesis similar to that of intact rabbits. Ovariectomized rabbits administered progesterone on the other hand resembled the ovariectomized rabbits without hormone replacement. Specific activity of hydroxyproline in individual specimens of the arch correlated strongly with degree of atherosclerosis. An analysis of regional variation demonstrated declining values for collagen synthesis in the caudal direction along the aorta. Collagen synthesis was much higher in the portal vein as compared with the inferior vena cava. Plasma cholesterol values were higher in the ovariectomized rabbits than in the intact rabbits. Tissue deposition of cholesterol paralleled the degree of atherosclerosis.     

Hydrophobic surfactant effects on aortic cholesterol accumulation and atherosclerosis in hypercholesterolemic rabbits

Studies were performed in hypercholesterolemic rabbits to determine whether the hydrophobic surfactant, Poloxalene 2930 (Pol), is of benefit under these conditions. Lipoprotein analyses plus chemical and morphologic studies of the aorta were performed to evaluate the results. In one study, rabbits were made hypercholesterolemic by dietary means and then divided into two groups and given a cholesterol-free diet with one group additionally given Pol with treatment continued for 10 weeks. Pol treatment resulted in less atherosclerosis but the mechanism for this effect was not apparent from lipoprotein analysis. In the other study 3 groups of rabbits were given a cholesterol-rich diet for 16 weeks. Two groups received Pol supplement with one of these groups receiving a dose that was too small to prevent hypercholesterolemia. In this group plus the group on diet alone comparable degrees of hypercholesterolemia were maintained throughout the study. Lipoprotein abnormalities were similar in these two groups except that those on Pol had a more normal cholesterol to apolipoprotein B ratio. The amount of atherosclerosis in both groups was mild but aortic cholesterol content was much less for the Pol group. It is concluded that Pol limits cholesterol accumulation in the aortic wall of hypercholesterolemic rabbits and can retard the development of atherosclerosis.     

L-ascorbic acid: effects on aortic glycosaminoglycan 35S incorporation in rabbit-induced atherogenesis.

The effects of L-ascorbic acid on 35S-incorporation into thoracic aorta glycosaminoglycans and upon aorta cholesterol levels were determined in hypercholesterolemic rabbits. No significant difference was observed in serum free or esterified cholesterol levels between animals receiving L-ascorbic acid supplementation or saline while maintained on a cholesterol diet (0.5%). A 15-fold higher serum cholesterol was observed in animals on the cholesterol diet to those animals which received a normal rabbit (Purina) diet. L-Ascorbic acid increased sulfated glycosaminoglycans concentrations in hypercholesterolemic rabbits which paralleled lower tissue free and esterified cholesterol levels. The 35S-specific activity of glycosaminoglycans in hypercholesterolemic animals receiving saline was much greater than in those animals receiving L-ascorbic acid. This suggests that L-ascorbic acid plays a role in the maintenance of adequate levels of aortic sulfated glycosaminoglycans. This then is a suggested biochemical mechanism of L-ascorbic acids interaction in the atherogenic process.     

Nitric oxide production and endothelium-dependent vasorelaxation ameliorated by N1-methylnicotinamide in human blood vessels

N(1)-methylnicotinamide (MNA(+)) has until recently been thought to be a biologically inactive product of nicotinamide metabolism in the pyridine nucleotides pathway. However, the latest observations imply that MNA(+) may exert antithrombotic and anti-inflammatory effects through direct action on the endothelium. We examined both in vivo and in vitro whether the compound might induce vasorelaxation in human blood vessels through the improvement of nitric oxide (NO) bioavailability and a reduction of oxidative stress mediated by endothelial NO synthase (eNOS) function. MNA(+) treatment (100 mg/m(2) orally) in healthy normocholesterolemic and hypercholesterolemic subjects increased the l-arginine (l-NMMA)-inhibitable flow-mediated dilation (FMD) of brachial artery responses that also positively correlated with MNA(+) plasma concentrations (r=0.73 for normocholesterolemics and r=0.78 for hypercholesterolemics; P<0.0001). MNA(+) increased FMD at the same concentration range at which it enhanced NO release from cultured human endothelial cells after stimulation with either the receptor-dependent (acetylcholine) or the receptor-independent endothelial NO synthase agonists (calcium ionophore A23187). MNA(+) restored the endothelial NO synthase agonist-stimulated NO release after the exposure of the cells to oxidized low-density lipoprotein. This effect was also associated with the normalization of the [NO]/[superoxide] balance in the endothelial cells. Taken together, the increased NO bioavailability in the endothelium contributes to the vasorelaxating properties of MNA(+). Targeting eNOS with MNA(+) might be therapeutically relevant for functional disorders of the endothelium, such as hypercholesterolemia and atherosclerosis.     

The effect of renal hypertension on the regional deposition of cholesterol and phospholipid in the aorta of normally- and cholesterol-fed rabbits.

The effect of renal hypertension on dry defatted tissue mass and lipid accumulation in different segments of the aortic intima was studied in both normally-fed and cholesterol-fed rabbits. In normally-fed rabbits hypertension caused an increase in intimal dry weight in the aorta. The increase was greatest in the lower thoracic intimal segment but was not significant in the aortic arch. The increase in tissue mass was not influenced by the addition of cholesterol to the diet and no regression of the increased tissue mass occurred when a 4-week period of hypertension was followed by a 4-week period of normotension. Hypertension did not increase the intimal cholesterol or phospholipid concentrations in normally-fed rabbits, suggesting that an observed increase in lipid content represented the cellular component of the intimal hypertrophy. Hypertension in cholesterol-fed animals caused preferential lipid accumulation in the lower thoracic segment, an effect that was independent of the total intimal cholesterol level. Intimal cholesterol, cholesterol ester and phospholipid were all increased. When a 4-week period of normotension and cholesterol feeding was preceded by a 4-week period of hypertension with normal feeding the amount of cholesterol deposited did not exceed that of the normotensive control, suggesting either that hypertension increased intimal permeability to lipid only in the presence of hypercholesterolaemia, or that healing of damaged intima had occurred before hypercholesterolaemia was fully established.     

CGP 43371 paradoxically inhibits development of rabbit atherosclerotic lesions while inducing extra-arterial foam cell formation

Ansamycins are hypolipidemic compounds which, when administered to various animal species, dramatically lower high density lipoprotein (HDL) cholesterol levels, in addition to reducing the levels of other lipoprotein classes. The current study tested one of these ansamycins (CGP 43371) for its hypolipidemic and anti-atherosclerotic activity in cholesterol-fed rabbits. Rabbits were fed a 0.25% cholesterol-enriched diet with or without admixed CGP 43371, equivalent to 30 mg/kg per day for 16 weeks. Compared with control rabbits, CGP 43371 treatment lowered total cholesterol levels (46%, P<0.05) and lipoprotein cholesterol levels (HDL, 58%; VLDL, 49% [both P<0.05]; LDL, 28% [not significant]). Despite the dramatic lowering of HDL cholesterol levels, aortic atherosclerosis, assessed by grossly visible sudanophilia, was significantly inhibited versus controls (total aorta=38%; aortic arch=32%; thoracic aorta=60%). Of particular note in CGP 43371-treated rabbits was a striking splenomegaly, which correlated with the presence of massive accumulations of macrophage foam cells in the splenic red pulp. We speculate that CGP 43371 inhibits the development of atheroselerotic lesions in rabbits by both a hypolipidemic mechanism, and by a mechanism(s) in which macrophage foam cells accumulate in the spleen.     

Intrauterine undernutrition: expression and activity of the endothelial nitric oxide synthase in male and female adult offspring

OBJECTIVE: Epidemiological studies suggest that intrauterine undernutrition plays an important role in the development of arterial hypertension in adulthood. In an attempt to define the mechanisms whereby blood pressure may be raised, we have hypothesized that arteries from offspring of nutritionally restricted dams exhibit abnormalities in the endothelial function and in nitric oxide synthesis. In order to investigate the existence of potential gender differences on the effects of intrauterine undernutrition, both male and female offspring of pregnant Wistar rats on normal and restricted diets were studied in adulthood. METHODS: Female pregnant Wistar rats were fed either normal or 50% of the normal intake diets, during the whole gestational period. At 14 weeks of age, the rats were used for the study of vascular reactivity, eNOS and iNOS gene expression, eNOS activity and, in the case of females, estrogen levels. RESULTS: Intrauterine undernutrition induced hypertension in both male and female offspring, but hypertension was more severe in male rats. Endothelium-intact aortic rings from male and female rats in the restricted diet group exhibited increased responses to norepinephrine, decreased vasodilation to acetylcholine and unaltered responses to sodium nitroprusside in comparison to aortic rings from control rats. No gender-related differences were observed in the vascular reactivity studies. Intrauterine undernutrition promoted decreased gene expression for eNOS in aorta isolated from male, but not female, offspring, reduction in eNOS activity in both male and female offspring and impairment in synthesis of estrogen in female offspring. CONCLUSION: Our data show that intrauterine undernutrition: (1) induces hypertension both in the male and female offspring, hypertension being more severe in male than in female rats; (2) alters endothelium-dependent responses in aortas from the resulting offspring. The endothelial dysfunction is associated with a decrease in activity/expression of eNOS in aortas from male offspring. The mechanism involved in altered response to ACh in female offspring might be a consequence of reduction in estrogen levels leading to reduced eNOS activity.     

HDL does not promote cholesterol efflux from macrophages of hypercholesterolemic rabbit: efflux differences between species.

The purpose of this study was to investigate the difference between species (mouse and rabbit) and the effect of hypercholesterolemia on the ability of their peritoneal macrophages to release unesterified cholesterol to an exogenous acceptor. The macrophages from mouse, normocholesterolemic rabbits and hypercholesterolemic rabbits (Mm, Mnr, Mhr) were loaded with cholesterol esters by incubation with oxidatively modified low density lipoprotein or plasma lipoprotein (beta-migrating very low density lipoproteins). When human HDL was used as cholesterol acceptor for 24 h of incubation, the Mm Cells, the Mnr cells and Mhr cells retained about 40%, 70%, and more than 90% of their cholesterol esters. The difference between species of lipoproteins were not effective for the ability to release cholesterol esters. ACAT (acylcoenzyme A: cholesterol acyltransferase) inhibitor 58-035 increased these capacities. These data suggest that the limited capacity of macrophages from hypercholesterolemic rabbits to release cholesterol may be related to the progression and resistance to regression of atherosclerosis, and that ACAT activity might influence this capacity.     

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.     

Effects of anticalcifying and antifibrobrotic drugs on pre-established atherosclerosis in the rabbit.

The effects of the anticalcifying drug, ethane-hydroxydiphosphonate (EHDP) and the inhibitors of collagen biosynthesis, colchicine, penicillamine and azetidine were studied in the rabbit with pre-established atherosclerosis. The drugs were administered with a cholesterol-free diet (regression diet) for 8 weeks following the induction of atherosclerosis by feeding a hypercholesterolemic diet containing 2% cholesterol and 8% peanut oil for 8 weeks. The extent and severity of aortic atherosclerosis, as revealed by the morphological and biochemical findings, increased significantly during the regression period. In rabbits treated with EHDP (5 mg/kg/day) the aorta had fewer gross lesions and contained significantly less cholesterol, collagen and elastin than did the aorta of the rabbits fed the regression diet alone. These changes were associated with a significant reduction in aortic calcium caused by EHDP. The aortic content of cholesterol, collagen and elastin in the EHDP-treated rabbits, although less than that of the rabbits fed the regression diet alone, was about the same as that of the rabbits fed a high cholesterol diet for 8 weeks. Both colchicine (0.2 mg/kg/day) and penicillamine (100 mg/kg/day) had a selective action on the induced plaques in that they suppressed the fibrous proliferation in the lesions without preventing lipid and calcium accumulation in the lesions. Neither colchicine nor penicillamine reduced the extent of aortic atherosclerosis as determined by gross examination of the vessel. Azetidine had no significant effect on the pre-established atherosclerotic lesions. The lipid, fibrous protein and calcium content of the aorta of the azetidine-treated animals was not significantly different from that of the untreated animals. The biochemical findings in the aorta were consistent with the microscopic changes.     

Effect of nisoldipine on atherosclerosis in the cholesterol fed rabbit: endothelium dependent relaxation and aortic cholesterol content

STUDY OBJECTIVE--The aim was to determine the effect of the calcium channel blocker nisoldipine on the loss of endothelium dependent relaxation and the accumulation of cholesterol in the aorta produced by feeding a diet enriched with cholesterol. DESIGN--12 week old New Zealand white rabbits were assigned randomly to four groups with the following dietary and drug regimens: group A--standard diet + 2.5% cholesterol (n = 45); group B--standard diet + nisoldipine (n = 9); group C--standard diet + nisoldipine + 2.5% cholesterol (n = 9); group D--standard diet (n = 9). After 3 weeks the cholesterol supplements were stopped and all animals were given the standard rabbit diet. The animals in groups B and C were given nisoldipine (1 mg.kg-1.d-1) by mouth for the entire 7 week period. EXPERIMENTAL MATERIAL--Aortic tissue was removed for measurement of cholesterol content, endothelium dependent relaxation to acetylcholine, contractile responses to noradrenaline, relaxant responses to sodium nitrite, and sudan staining. Serum was obtained for measurement of cholesterol and triglyceride concentration. MEASUREMENTS AND MAIN RESULTS--At 7 weeks, endothelium dependent relaxation to acetylcholine was impaired in group A compared to group D, while that in group C was not. Aortic tissue cholesterol content in group A was significantly greater than in groups B, C, and D. At 15 weeks, ie, 12 weeks after reversal of the diet, endothelium dependent relaxation had recovered in the animals in group A. There was a significant reduction in the aortic cholesterol content at this stage. In two subgroups of A (groups A2 and A4) which were given nisoldipine immediately after and 4 weeks after cessation of cholesterol feeding respectively, the drug was found to have no influence upon restoration of endothelium dependent relaxation. However, the drug appeared to promote the retention of cholesterol within the aorta after cessation of cholesterol feeding. CONCLUSIONS--Nisoldipine protects against the accumulation of cholesterol and loss of endothelium dependent relaxation in the aorta of rabbits fed a diet supplemented with 2.5% cholesterol for three weeks. Administration of the drug after the lesions are established in the aorta also appears to retard the removal of cholesterol from the aorta.