瑞舒伐他汀对载脂蛋白E基因敲除小鼠动脉粥样硬化的影响

目的 探讨瑞舒伐他汀对载脂蛋白E基因敲除(apoE-/-)小鼠主动脉粥样硬化的影响.方法 取apoE-/-小鼠18只建立动脉粥样硬化模型,C57BL/6小鼠12只为对照组.apoE-/-小鼠予高脂饲料,C57BL/6小鼠予普通饲料.12周后,随机抽取C57BL/6小鼠和apoE-/-小鼠各6只,判断是否成模.将成模后余下的12只apoE-/-小鼠随机分为模型组和瑞舒伐他汀治疗组(瑞舒伐他汀10 mg·kg-1·d-1灌胃),每组6只.余下的6只C57BL/6小鼠作为对照组.再过12周后处死小鼠,行血脂及主动脉HE、Masson、油红O染色观察动脉斑块,免疫组织化学方法检测主动脉组织平滑肌肌动蛋白(α-SMA)、转化生长因子β1(TGF-β1)、巨噬细胞表面分子-3(Mac-3)表达.结果 模型组小鼠胆固醇、低密度脂蛋白水平均高于对照组(P均＜0.01),甘油三酯水平与对照组比较差异无统计学意义.模型组小鼠主动脉组织内可见明显动脉粥样硬化斑块形成,α-SMA、TGF-β1和Mac-3表达均较高于对照组(P均＜0.01).瑞舒伐他汀治疗组小鼠胆固醇、低密度脂蛋白、甘油三酯水平与模型组比较差异无统计学意义,但斑块内脂肪含量少于模型组,胶原含量多于模型组.治疗组α-SMA表达与模型组比较差异无统计学意义,治疗组TGF-β1、Mac-3表达均低于较模型组(P均＜0.01).结论 瑞舒伐他汀可以减轻apoE-/-小鼠动脉粥样硬化模型中的脂质沉积和炎症反应,可以增加其胶原含量,利于斑块的稳定,具有抗动脉粥样硬化的作用,对血脂无影响.

Abstract：

Objective To investigate the effect of rosuvastatin on atherosclerosis in apoE-knockout ( apoE - / - ) mice. Methods Eighteen 6-week-old apoE - / - mice fed with high fat diet were used as atherosclerosis models, twelve 6-week-old C57BL/6 mice fed with normal diet were used as control. After twelve weeks, six apoE -/ - mice were used to observe the formation of atherosclerosis. Another 12 apoE -/- mice were divided into placebo treated group (n =6) and rosuvastatin group (n =6,10 mg· kg-1 ·d -1 per gavage) and treated for 12 weeks. Then, blood was collected for measuring lipid, aorta was prepared for morphologic study (HE, Oil red O, Masson) and immunohistochemical analysis (α-smooth activor protein, transforming growth factor β1, macrophage surface molecule-3 ). Results Serum cholesterol and low density lipoprotein levels were significantly higher in apoE -/- mice fed with high fat diet than in C57/ BL6 mice( all P ＜0. 01 )while triglyceride level was similar between the two groups, these were not affected by rosuvastatin. Similarly, atherosclerotic lesion area in apoE -/ - mice fed with high fat diet was also not significantly reduced by rosuvastatin, while lipid deposition could be significantly reduced and collagen deposition could be significantly increased in the aortic atherosclerotic lesions by treatment with rosuvastatin.Upregulated TGF-β1 and Mac-3 expression in the aortic atherosclerotic lesions in apoE -/- mice fed with high fat diet could also be significantly reduced by rosuvastatin (all P ＜ 0. 01 ), suggesting reduce inflammatory responses in the atherosclerotic lesion and stable atherosclerotic plaque post rosuvastatin treatment. Conclusion Reducing inflammatory responses and stabilizing plaque properties might contribute to the anti-atherosclerosis effects of rosuvastatin in mice high fat diet fed apoE -/- mice.     

Increased atherosclerotic lesion area in apoE deficient mice overexpressing bovine growth hormone

Human growth hormone (GH) excess is linked to increased cardiovascular morbidity and mortality. However, little is known about the effect of GH excess on atherosclerosis. We developed a new mouse model to assess the hypothesis that GH overexpression accelerates atherosclerotic lesion formation. apoE(-/-) mice were crossed with bovine GH (bGH) transgenic mice to yield apoE(-/-) mice overexpressing bGH (apoE(-/-)/bGH). The mice were fed either standard or Western diet. At 22 weeks, atherosclerotic lesion area of thoracic aorta was larger in apoE(-/-)/bGH mice compared with littermate apoE(-/-) mice fed either diet (standard: +161+/-50%, Western: +430+/-134%). Aortic sinus lesions were more severe in apoE(-/-)/bGH mice fed standard diet compared with littermate apoE(-/-) mice. apoE(-/-)/bGH mice had lower (VLDL+LDL)/HDL ratios compared with littermate apoE(-/-) mice, while systolic blood pressure was higher in apoE(-/-)/bGH mice, irrespective of diet. The levels of serum amyloid A and hepatic CRP mRNA were higher in apoE(-/-)/bGH mice than in littermate apoE(-/-) mice. In conclusion, this study shows that excess GH augments the development of atherosclerosis in apoE(-/-) mice. The mechanisms could be direct effects of GH on cellular processes in the vessel wall or the result of concomitant processes such as hypertension or a general inflammatory state.     

Leptin deficiency suppresses progression of atherosclerosis in apoE-deficient mice

Both experimental and epidemiological studies suggest that leptin is one of the molecules responsible for accelerated atherosclerosis in obese humans. To confirm the notion, we studied whether leptin accelerates atherosclerosis in apoE(-/-) mice. Leptin deficient hyperlipidemic mice (ob/ob;apoE(-/-) mice) developed significantly less atherosclerosis than apoE(-/-) mice, when fed an atherogenic diet for 16 weeks from 8 weeks of age. Histological analysis revealed that most of the atherosclerotic lesions in ob/ob;apoE(-/-) mice remained as fatty streaks, while those in apoE(-/-) mice were mainly fibrous plaques. The decrease in atherosclerosis was not due to changes in the serum levels of cholesterol, TNF-alpha, or adiponectin. Exogenous leptin significantly increased atherosclerotic areas in apoE(-/-) mice, even though it decreased food intake and body weight. Our findings support the notion that leptin accelerates atherosclerosis.     

Cardiovascular protective effects of synthetic isoflavone derivatives in apolipoprotein e-deficient mice

Dietary flavonoids are thought to protect against cardiovascular disease. We have studied the effects of bioactive isoflavone metabolites on hyperlipidemia, endothelial dysfunction and the development of atherosclerotic lesions in apolipoprotein E-deficient (apoE(0)) mice fed a Western high-fat diet. Supplementation with dihydrodaidzein (DiD), dehydroequol (DeE) (both 25 mg kg(-1) x day(-1)) and their combination (D/D; 12.5 mg kg(-1) x day(-1) for each) for 24 weeks reduced plasma high-density lipoprotein (HDL) and non-HDL cholesterol. D/D also reduced the triglyceride level. In the abdominal aorta of apoE(0) mice, these compounds significantly increased endothelial nitric oxide (NO)-mediated vasorelaxations induced by acetylcholine, but had a minor effect on relaxations induced by the NO donor S-nitroso-N-acetylpenicillamine. Isoflavone treatment for 24 weeks had no effect on the total area of atherosclerotic plaques in the whole aorta, but DeE reduced the plaque thickness in the aortic arch by 29%, although this did not reach statistical significance. The endothelial dysfunction in apoE(0) mice is associated with hyperlipidemia and increased vascular oxidative stress measured as increased superoxide production. Both isoflavones have superoxide-scavenging activities in vitro. We suggest that chronic supplementation with bioactive isoflavone metabolites may protect endothelial NO function in apoE(0) mice, through both lipid-lowering and antioxidant actions.     

The TNF alpha converting enzyme (TACE/ADAM17) is expressed in the atherosclerotic lesions of apolipoprotein E-deficient mice: possible contribution to elevated plasma levels of soluble TNF alpha receptors

TNF alpha converting enzyme (TACE) critically regulates the inflammatory processes as it releases from the cell surface several transmembrane proteins, including TNFalpha (TNF) and its receptors TNFR1 and TNFR2. We investigated the expression of TACE in atherosclerotic lesions of apolipoproteinE-deficient (apoE (-/-)) mice. Five-week-old apoE(-/-) male mice were fed a high-fat diet and examined at 5, 10, 15 and 25 weeks of age. A group of wild-type C57BL/6 mice (WT) fed the high-fat diet for 25 weeks was included. In apoE(-/-) mice, lesions progressed with time in both aortic sinus and arch, in which TACE immunostaining also increased particularly between 5 and 15 weeks. TACE expression was also observed in human atherosclerotic plaques. The plasma levels of soluble TNFR1 and TNFR2 rose with atherosclerosis. In the 25-week-old WT mice, no lesions were observed and the plasma levels of TNFRs were 17% of those of age-matched apoE(-/-) mice. Incubated aortas of 25-week-old apoE(-/-) mice released much higher amounts of sTNF and sTNFRs than did aortas of 5-week-old apoE(-/-) mice or 25-week-old WT mice. Active TACE was expressed at the surface of macrophages isolated from apoE(-/-) mice. In conclusion, TACE expression is associated with lesions in atherosclerosis-prone sites. Our data suggest that atherosclerotic lesions-expressing TACE may contribute to the elevated levels of circulating sTNFRs.     

Low carbohydrate, high protein diet promotes atherosclerosis in apolipoprotein E/low-density lipoprotein receptor double knockout mice (apoE/LDLR(-/-))

Although in apoE/LDLR(-/-) mice atherosclerotic plaques develop spontaneously, various atherogenic diets (e.g. Western diet) are frequently used to accelerate the disease in this model. The objective of this study was to compare the effects on atherosclerosis of Western diet and other types of high-fat, high cholesterol, hypertriglyceridemic diets with the effects of the low carbohydrate, high protein (LCHP) diet. 16-18 week old mice with pre-established atherosclerosis were assigned to experimental groups and fed for the next 10 weeks with control diet, margarine diet (margarine 7%), hypertrigliceridemic diet (fructose 62%), high-fat diet (Western diet), high cholesterol diet (egg yolk diet) or with LCHP diet. No differences in body weight were observed among experimental groups. Plasma cholesterol concentration was significantly increased in egg yolk diet- and LCHP diet-fed apoE/LDLR(-/-) mice as compared to other types of diets. Plasma concentration of triacylglycerols was significantly elevated in egg yolk diet- and LCHP diet-fed apoE/LDLR(-/-) mice. The area of atherosclerotic plaques in the aortic root was substantially increased in LCHP diet-fed mice as compared to other types of diets. Furthermore, in brachiocephalic arteries of LCHP diet-fed mice there was evidence of plaque rupture. In conclusion, the LCHP diet promoted atherosclerosis in apoE/LDLR(-/-) mice more intensively than classical Western diet and favored the development of unstable lesions.     

C3H apoE(-/-) mice have less atherosclerosis than C57BL apoE(-/-) mice despite having a more atherogenic serum lipid profile

Wild-type C57BL mice are known to be susceptible to diet-induced atherosclerosis, whilst C3H mice are resistant. We investigated the effect of these background strains on the hyperlipidaemia and atherosclerosis that develops in mice deficient in apolipoprotein E (apoE(-/-)). Male and female apoE(-/-) mice on C3H/HeNHsd (C3H) and C57BL/6J (C57) backgrounds were fed atherogenic Western diet for 12 weeks. Serum cholesterol and triglyceride concentrations were measured and atherosclerosis quantified in the aortic sinus. C3H apoE(-/-) mice fed normal diet had 1.5 2 fold higher serum cholesterol levels than C57 apoE(-/-) mice and 4-5 fold higher serum triglyceride concentrations. Feeding Western diet caused a 4-5 fold increase in serum cholesterol in all mice, but levels of triglyceride were either attenuated or were unaffected in C3H apoE(-/-) and C57 apoE(-/-) mice, respectively. C3H apoE(-/-) mice had approximately 2 fold higher serum cholesterol and 4 fold higher triglyceride concentrations than the C57 apoE(-/-) mice throughout the study. Serum triglyceride concentrations were 35-108% higher in male C3H apoE(-/-) than female C3H apoE(-/-) mice. Most of the lipids were present in the very low density lipoprotein (VLDL)/chylomicron fraction in both strains of mice whether they were fed normal or Western diet. Notwithstanding the lower plasma lipid concentrations, atherosclerotic lesion areas were more than 2-fold larger in C57 apoE(-/-) than in C3H apoE(-/-) mice (males 68 +/- 11 x 10(3) vs 30 +/- 6 x 10(3) females 102 +/- 12 x 10(3) vs 41 +/- 8 x 10(3) microm2. mean +/- SEM).     

Hydrogen-rich Saline Reduces Atherogenesis in Apolipoprotein E Knockout Mice Fed a High-fat Diet by Inhibiting the Non-HDL-mediated Arterial Inflammation and Promoting the Expression of Genes Involving Reverse Cholesterol Transport

Aim Hydrogen(dihydrogen,H2) is an effective antioxidant to reduce oxidative stress and oxidative stress is implicated in atherogene-sis.In this study we examined whether hydrogen-saturated saline can prevent atherosclerosis in apolipoprotein E knockout(apoE-/-) mice fed either chowdiet or high-fat diet,and characterized the underlying molecular mechanisms.Methods and Results The atherosclerotic lesion formation displayed by oil red O staining positive area was reduced significantly in either aortic root section or aortic arch en face in hydrogen administrated apoE-/-mice fed either chowdiet or high-fat diet,compared to the control.Plasma analysis by enzymatic method showed that total cholesterol(TC) and non-high-density lipoprotein cholesterol(non-HDL-C)were remarkably decreased by treatment with hydrogen.Western blot analysis revealed a significant decrease of both plasma apoli-poprotein B(apoB) level and hepatic expression of apoB after hydrogen treatment,suggesting hydrogen could downregulate the expression of the major protein constituent of non-HDL.In addition,spectrophotometric measurement showed that plasma levels of malondi-aldehyde(MDA) and serum amyloid Awas decreased and paraoxonase-1 activity was increased in mice treated with hydrogen,suggesting plasma lipid oxidation and peroxidation was impaired by hydrogen treatment.Besides,the MDA content of the non-HDL,whichseparated by ultracentrifugation from the plasma of mice treated with and without hydrogen,was reduced by hydrogen,suggesting the oxidation of non-HDL was impaired by hydrogen.Moreover,we found hydrogen treatment significantly suppressed the production of tumor necrosis factor-α(TNF-α) and interleukin-6 in RAW264.7 macrophages after stimulation with the isolated non-HDL,suggesting hydrogen reduces atherogenesis by inhibiting non-high-density lipoprotein(HDL)-mediated inflammation.Furthermore,immunohis-tochemistry of aortic valve sections revealed that hydrogen attenuated lesion formation by suppressing the expression of several proin-flammatory factors and decreasing vessel wall infiltration of macrophages,indicating hydrogen-treatment reduces arterial inflammation.Besides,real-time PCR and western blot analysis disclosed that the expression of several transporter genes involved in the process ofreverse cholesterol transport,including hepatic scavenger receptor class B type I(SR-BI),ATP-binding cassette(ABC) transporters ABCG8,ABCB4,ABCB11,and macrophage SR-BI,were all induced by hydrogen treatment.Conclusion These results re-vealed that administration of hydrogen-rich saline reduces atherogenesis in apoE-/-mice fed a high-fat diet by inhibiting the non-HDL-mediated arterial inflammation and promoting the expression of genes involving reverse cholesterol transport.     

Atherosclerosis in C3H/HeJ mice reconstituted with apolipoprotein E-null bone marrow

Previous studies showed that reconstitution of atherosclerosis-susceptible C57BL/6 (B6) female mice with apolipoprotein E (apoE)-deficient (apoE(-/-)) bone marrow resulted in markedly increased atherosclerosis, despite the fact that plasma lipid levels were unchanged. To determine whether apoE(-/-) bone marrow would increase atherosclerosis in an atherosclerosis-resistant strain, female C3H/HeJ (C3H) mice were lethally irradiated and reconstituted with bone marrow from either C3H.apoE(-/-) mice or wild-type C3H mice. Four weeks after transplantation, the mice were fed an atherogenic diet for 12 weeks. We found that reconstitution of C3H mice with apoE(-/-) bone marrow resulted in a slight reduction in plasma apoE levels and a dramatic reduction in apoE and apolipoprotein B (apoB) in the aortic wall. Plasma apoB and cholesterol levels were unchanged, as were atherosclerotic lesions at the aortic root. These data indicate that reconstitution of C3H mice with apoE(-/-) bone marrow has no effect on atherosclerosis susceptibility and that apoE promotes accumulation of apoB in the vessel wall.     

Accelerated atherosclerosis in apolipoprotein E-deficient mice fed Western diets containing palm oil compared with extra virgin olive oils: a role for small, dense high-density lipoproteins

To test the hypothesis that extra virgin olive oils from different cultivars added to Western diets might behave differently than palm oil in the development of atherosclerosis, apoE-deficient mice were fed diets containing different cultivars of olive oil for 10 weeks. Female mice were assigned randomly to one of the following five groups: (1-4) fed chow diets supplemented with 0.15% (w/w) cholesterol and 20% (w/w) extra virgin olive oil from the Arbequina, Picual, Cornicabra, or Empeltre cultivars, and (5) fed a chow diet supplemented with 0.15% cholesterol and 20% palm oil. Compared to diets containing palm oil, a Western diet supplemented with one of several varieties of extra virgin olive oil decreased atherosclerosis lesions, reduced plaque size, and decreased macrophage recruitment. Unexpectedly, total plasma paraoxonase activity, apoA-I, plasma triglycerides, and cholesterol played minor roles in the regulation of differential aortic lesion development. Extra virgin olive oil induced a cholesterol-poor, apoA-IV-enriched lipoparticle that has enhanced arylesterase and antioxidant activities, which is closely associated with reductions in atherosclerotic lesions. Given the anti-atherogenic properties of extra virgin olive oil evident in animal models fed a Western diet, clinical trials are needed to establish whether these oils are a safe and effective means of treating atherosclerosis.     

小鼠腹主动脉斑块模型建立的两种方法及超高场强磁共振成像的应用价值

目的 探讨建立腹主动脉粥样硬化斑块模型的方法及超高场强磁共振成像(MRI)在活体监测及量化小鼠腹主动脉粥样硬化斑块中的应用价值.方法 高脂饮食法小鼠腹主动脉粥样硬化斑块模型的建立及磁共振检测(高脂饮食组):选择3批10～12月龄apoE-/-小鼠13只、WT鼠3只高脂饮食喂养,分别于喂养前、喂养3个月、喂养6个月3个时期进行小鼠腹主动脉7.0 T磁共振活体扫描.血管紧张素Ⅱ(AngⅡ)灌注法小鼠腹主动脉粥样硬化斑块模型的建立及磁共振检测(AngⅡ灌注组):选用10只6月龄apoE-/-小鼠,分为AngⅡ1000 ng·kg-1·min-1组3只、AngⅡ500 ng·kg-1·min-1组3只(以上两组均在背部埋置AngⅡ缓释泵14 d)和对照组为4只(埋置生理盐水缓释泵),分别于灌注前后行磁共振扫描,选用FLASH T1WI黑血及MSME-T2WI-PDWI双回波序列.高脂饮食组依次在各时期扫描后分别处死3、5、5只小鼠,AngⅡ灌注组于装泵后14 d行磁共振扫描,然后处死本组小鼠,取处死小鼠的肾动脉段腹主动脉制作病理切片,进行苏木素-伊红(HE)染色、Masson胶原纤维(CME)染色.每只小鼠选5～7层肾动脉段腹主动脉病理图像及多对比MRI图像,分别测量外腔面积(VOA)、内腔面积(LA),计算管肇面积(VWA)并进行MRI与病理测量结果的相关性分析.结果 两种方法建立的小鼠模型,其腹主动脉MRI与病理切片均可见不稳定斑块形成.高脂饮食组随着高脂饮食时间的延长,斑块进展,VWA不断增加,3个时期VWA方差分析F=29.94(P<0.05),斑块信号于PDWI、T2WI逐渐增加,且不均匀.高脂饮食组MRI测量的斑块面积与病理测量的斑块面积有较高的-致性(高脂喂养前、喂养3和6个月3个时期r值分别为0.84、0.95、0.90).病理切片中斑块成分与磁共振显示信号一致,均表现为脂质成分增加,纤维成分减少.AngⅡ1000 ng·kg-1·min-1组AngⅡ灌注后斑块面积与灌注前比较差异有统计学意义(P=0.017),分别为(2.65±0.48)mm2和(1.21±0.21)mm2,部分小鼠可见夹层动脉瘤形成.AngⅡ500 ng·kg-1·min-1组灌注后斑块面积也比灌注前有所进展,面积分别为(1.01±0.17)mm2和(0.85±0.11)mm2,MRI测量的斑块面积与病理测量的斑块面积有较高的一致性(r值为0.93).结论 AngⅡ灌注显著加快动脉粥样硬化进展并促进腹主动脉夹层动脉瘤形成,长期高脂饮食亦可形成晚期斑块.超高场强MRI多种序列黑血技术的综合应用能显示小鼠腹主动脉粥样硬化斑块的进展,其检测分析斑块大小结果与病理表现基本一致,对斑块成分的判定亦有一定价值.     

Contrasting effect of fish oil supplementation on the development of atherosclerosis in murine models

OBJECTIVE: Increased fish oil intake is associated with protection against coronary heart disease and sudden death, while effects on atherosclerosis are controversial. We explored the effects of supplementing fish oil (rich in n-3 polyunsaturated fatty acids, PUFA) or corn oil (rich in n-6 PUFA) in two different models of atherosclerosis. METHODS AND RESULTS: Sixty-three low density lipoprotein receptor-deficient (LDLR(-/-)) mice and sixty-nine apolipoprotein E-deficient (apoE(-/-)) mice were fed diets without supplementations or supplemented with either 1% fish oil or 1% corn oil. In apoE(-/-) mice, neither fish oil nor corn oil had any major impact on plasma lipids or atherosclerosis. In LDLR(-/-) mice, conversely, the fish oil and the corn oil group had lower levels of LDL-cholesterol and triglycerides and had lesser atherosclerosis in the aortic root and in the entire aorta (p < 0.01 versus unsupplemented group). Atherosclerosis was significantly less in the fish oil group compared with the corn oil group when evaluated en face in the aortic arch (area positive to lipid staining: 32% with fish oil versus 38% with corn oil; 48% with unsupplemented diet). CONCLUSIONS: n-3 and n-6 PUFA supplementation retarded the development of atherosclerosis in LDLR(-/-) mice, with a stronger effect seen with n-3 PUFA. There was an important strain-dependence of the effect, with no protection against atherosclerosis in apoE(-/-) mice.     

Contribution of Polyunsaturated Fatty Acids to Intestinal Repair in Protein-Energy Malnutrition

The aim of this study was to assess the effect of polyunsaturated fatty acids supplied in the diet on intestinal mucosa repair in a rat model of protein-energy malnutrition. Rats were fed either a standard semipurified diet or the same diet containing lactose as the only source of carbohydrate to cause protein-energy malnutrition. Diarrhea was induced within 24 h and was maintained for 2 weeks, after which both groups of rats were fed for 1 week either the standard diet or the standard diet supplemented with different sources of fatty acids, such as olive oil (OO), fish oil (FO), and purified phospholipids from pig brain (BPL). The lactose-enriched diet caused loss of enterocyte microvilli, lymphocyte infiltration, supranuclear cytoplasmic vesiculation, decreased number of goblet cells, low-density enlarged mitochondria, and less cristae. The FO diet improved the pathology score according to the histological and ultrastructural analysis, with an increased number of goblet cells, ratio of microvilli length to crypt depth, and percentage of intraepithelial lymphocytes compared to those found in rats with protein-energy malnutrition. We previously reported that chronic diarrhea depletes the antioxidant defense in rat intestine; we now show that both, the FO and the BPL diets, increase GSH levels in colon and that some antioxidant enzyme activities vary according to the source of fatty acids, with higher catalase and superoxide dismutase by the FO diet in jejunum, increased catalase by the BPL diet in jejunum, and elevated glutathione peroxidase by the OO diet in colon. The fatty acid profile of intestinal mucosa reflects the source of fat in the diet, with the lowest ratio of n-6/n-3 for rats fed the FO diet. These results suggest that dietary polyunsaturated fatty acids, particularly those in the n-3 series, may play an important role in intestinal repair in chronic diarrhea due to protein-energy malnutrition. This work was supported by Grant CI1-CT91-0078 from the European Union.     

Dietary cholesterol suppresses the ability of olive oil to delay the development of atherosclerotic lesions in apolipoprotein E knockout mice

To test the hypothesis that cholesterol might suppress the beneficial effect of olive oil in atherosclerosis, we fed apoE KO mice diets containing extra virgin olive oil, either with or without cholesterol, for 10 weeks and assessed the development of atherosclerosis. Within each sex, mice were assigned randomly to one of the following four experimental groups: (1) a standard chow diet, (2) a chow diet supplemented with 0.1% cholesterol (w/w) cholesterol, (3) a chow diet enriched with 20% (w/w) extra virgin olive oil and (4) a chow diet containing 0.1% cholesterol and 20% extra virgin olive oil. On the standard chow diet, average plasma cholesterol levels were higher in males than in females. Olive oil- and cholesterol-enriched diets, separately or in combination, induced hypercholesterolemia in both sexes, and abolished the difference between the sexes in plasma cholesterol levels. The addition of cholesterol to chow or olive oil diets decreased apolipoprotein A-I significantly in females and serum paraoxonase activities in males. The latter activity was higher in females than in males. In both sexes, the size of aortic atherosclerotic lesions was similar in olive oil- and chow-fed animals and smaller than in cholesterol-supplemented groups. Size of aortic lesions were positively correlated with circulating paraoxonase activity, particularly in males, and the relationship remained after adjusting for apolipoprotein A-I and HDL cholesterol levels. Our results demonstrate that the nutritional regulation of paraoxonase is an important determinant of atherosclerotic lesions dependent on sex. They also suggest that the mere inclusion of olive oil in Western diets is insufficient and the adoption of Mediterranean diet would be more effective in retarding the development of atherosclerotic lesions.     

Regression of atherosclerosis in monkeys reduces vascular superoxide levels

Superoxide (O2*-) in arteries may contribute to atherosclerosis in part by inactivation of nitric oxide. We hypothesized that regression of atherosclerosis in nonhuman primates is associated with a decrease in vascular NAD(P)H oxidase, decreased O2*- levels, and improved endothelium-dependent relaxation. Cynomolgus monkeys (n=28) were fed an atherogenic diet for 47+/-10 (mean+/-SE) months. In carotid arteries (containing advanced lesions), femoral arteries (moderate lesions), and saphena arteries (minimal lesions), we examined O2*- levels and vasomotor function. Compared with vessels from normal monkeys (n=8), O2*- levels (measured by lucigenin-enhanced chemiluminescence) were 3.3-fold higher in carotid, 1.7-fold higher in femoral, and not different in saphena arteries from atherosclerotic monkeys. Dihydroethidium staining also demonstrated increased O2*- levels throughout the vessel wall in femoral and carotid arteries from atherosclerotic monkeys. Components of the NAD(P)H oxidase (p22(phox) and p47(phox)) were increased in atherosclerotic arteries, and immunohistochemistry demonstrated colocalization primarily to areas of macrophage infiltration. Relaxation to acetylcholine was impaired in carotid and femoral, but not saphena, arteries from atherosclerotic monkeys. After 8 months of regression diet (n=9), serum cholesterol decreased to normal, and O2*- levels (basal and NAD(P)H-stimulated), as well as expression of NAD(P)H oxidase, returned toward normal. Relaxation to acetylcholine improved in femoral arteries, but not in the more diseased carotid arteries. We conclude that, in a primate model of moderately severe atherosclerosis and regression of atherosclerosis, changes in endothelial function are inversely related to O2*- and NAD(P)H oxidase levels. Reduction in vascular O2*- during regression of atherosclerosis may contribute to improvement in vasomotor function.     

Activation of human aortic endothelial cells by LDL from Type 1 diabetic patients: an in vitro study

Hyperlipidaemia may accelerate the development of atherosclerosis by enhancing the expression of chemokines by cells within the arterial wall. Chemokines of the CC subfamily are clearly implicated in atherogenesis; however, recent reports suggest that CXC chemokines may play a hitherto unrecognised role in monocyte recruitment into atheromatous lesions expressing these molecules. Here, we examine whether circulating levels of CXC chemokines may reflect the pathogenic changes occurring during early atherogenesis. ApoE*3 Leiden mice developed marked hypercholesterolaemia, and early Type I 'fatty streak' lesions, following consumption of an atherogenic diet high in saturated fat and cholesterol, and containing sodium cholate, for up to 4 weeks. By contrast, their non-transgenic littermates (C57BL/6J) exhibited a much less pronounced hypercholesterolaemia and did not develop fatty streak lesions, when fed the same diet. Under these conditions, serum concentrations of CXC chemokines, KC and Macrophage Inflammatory Protein-2 (MIP-2) were significantly (P相似文献   

非洛地平对载脂蛋白E基因敲除小鼠动脉粥样硬化斑块及烟酰胺腺嘌呤二核苷酸氧化酶的影响

目的:探讨钙通道阻断剂(CCB)非洛地平对载脂蛋白E基因敲除(apoEKO)小鼠动脉粥样硬化斑块及烟酰胺腺嘌呤二核苷酸[NAD(P)H]氧化酶的影响。方法:apoEKO小鼠随机分为普通饮食组、高胆固醇饮食组、高胆固醇饮食加非洛地平组。无创血压系统测小鼠血压;内眦动脉取血检测血清TC和TG水平;冷冻切片光镜下定位主动脉根部,油红O染色评估斑块大小;实时定量PCR和Western blot方法检测主动脉中NAD(P)H氧化酶亚基p47phox和Rac-1表达。结果:高胆固醇饮食组小鼠血压没有明显变化,血脂明显升高(P<0.01),且斑块面积明显高于普食组(P<0.01);非洛地平可以明显减小斑块面积(P<0.01),同时还可以降低NAD(P)H氧化酶亚基p47phox和Rac-1表达(P<0.01)。结论:非洛地平可能通过阻断氧化应激反应抑制动脉粥样硬化发生发展。     

Short-term hyperglycemia increases arterial superoxide production and iron dysregulation in atherosclerotic monkeys

The incidence and severity of atherosclerotic vascular disease are increased in diabetic patients, in part because of increased production of reactive oxygen species (ROS). Previously, we found both increased atherosclerosis and arterial protein oxidation 6 months after streptozotocin-induced diabetes in monkeys fed an atherogenic diet, the pattern of which was indicative of redox-active transition metal involvement. The goal of this study was to determine if short-term (1 month) hyperglycemia increases oxidative stress and dysregulates iron metabolism before differences in atherosclerosis. Cynomolgus monkeys with preexisting atherosclerosis were stratified by dietary history and plasma lipids and received either streptozotocin (STZ-DM; n = 10) or vehicle (control; n = 10). One month after diabetes induction, blood and artery samples were collected. There were no differences in plasma lipoprotein cholesterol, arterial cholesterol, and atherosclerosis between control and STZ-DM. However, plasma lipid peroxides were elevated 137% (P < .01); arterial superoxide was increased 47% (P < .05); plasma ferritin, an indicator of whole-body iron stores, was 46% higher (P < .05); and iron deposition within aortic atherosclerotic lesions was more prevalent in STZ-DM compared with controls. Arterial levels of the antioxidant enzymes, superoxide dismutase, catalase, and heme oxygenase-1 were not higher in STZ-DM, although superoxide was higher, suggesting impaired antioxidant response. The increase in ROS before differences in atherosclerosis supports ROS as an initiating event in diabetic vascular disease. Further studies are needed to determine if increases in iron stores and arterial iron deposition promote hydroxyl radical formation from superoxide and accelerate diabetic vascular damage.