bluuebunny/crossref_metadata_2025 huggingface dataset:
Endothelial Nuclear Factor-κB Translocation and Vascular Cell Adhesion Molecule-1 Induction by Complement
Authors: Collard Charles D. , Agah Azin , Reenstra Wende , Buras Jon , Stahl Gregory L.
Published: June 2011

Abstract—We have previously shown that reoxygenation of hypoxic human umbilical vein endothelial cells (HUVECs) leads to the activation and deposition of complement. In the present study, we investigated whether the terminal complement complex (C5b-9) influences HUVEC nuclear factor-κB (NF-κB) translocation and vascular cell adhesion molecule-1 (VCAM-1) protein expression after hypoxia/reoxygenation by decreasing endothelial cGMP. Additionally, we investigated the action of anti-human C5 therapy on endothelial cGMP, NF-κB translocation, and VCAM-1 protein expression. Reoxygenation (0.5 to 3 hours, 21% O2) of hypoxic (12 hours, 1% O2) HUVECs in human serum (HS) significantly increased C5b-9 deposition, VCAM-1 expression, and NF-κB translocation compared with hypoxic/reoxygenated HUVECs treated with the recombinant human C5 inhibitor h5G1.1-scFv. Acetylcholine (ACh)-induced cGMP synthesis was significantly higher in normoxic HUVECs compared with hypoxic HUVECs reoxygenated in HS but did not differ from hypoxic HUVECs reoxygenated in buffer or HS treated with h5G1.1-scFv. Treatment of hypoxic/reoxygenated HUVECs with h5G1.1-scFv or cGMP analogues significantly attenuated NF-κB translocation and VCAM-1 protein expression. Treatment with NO analogues, but not a cAMP analogue, cGMP antagonists, or an NO antagonist, also significantly attenuated VCAM-1 expression. We conclude that (1) C5b-9 deposition, NF-κB translocation, and VCAM-1 protein expression are increased in hypoxic HUVECs reoxygenated in HS; (2) reoxygenation of hypoxic HUVECs in HS, but not buffer alone, attenuates ACh-induced cGMP synthesis; and (3) treatment of hypoxic/reoxygenated HUVECs with h5G1.1-scFv attenuates C5b-9 deposition, NF-κB translocation, and VCAM-1 expression while preserving ACh-induced cGMP synthesis. C5b-9–induced VCAM-1 expression may thus involve an NO/cGMP-regulated NF-κB translocation mechanism.
Oxidized LDL Stimulates Matrix Metalloproteinase-1 Expression in Human Vascular Endothelial Cells
Authors: Huang Yan , Mironova Marina , Lopes-Virella Maria F.
Published: June 2011

Abstract—It has been well documented that acute myocardial infarction is triggered by disruption of atherosclerotic plaques. Immunocytochemistry studies have shown that matrix metalloproteinase-1 (MMP-1) is specifically expressed by cells present in atherosclerotic plaques, including luminal and neovascular endothelial cells. Since MMP-1 degrades type I collagen, a major type of collagen in atherosclerotic lesions, it is likely that MMP-1 is involved in promoting destabilization of plaques. To date, however, the stimulatory factors that induce MMP-1 expression in endothelial cells have not been well defined. In the present study, we found that oxidized low density lipoprotein (LDL) stimulated MMP-1 release from both human umbilical vein and aortic endothelial cells. We also found that oxidized LDL markedly stimulated MMP-1 expression in these cells and that the degree of LDL oxidation was positively correlated with the level of MMP-1 mRNA expression. Furthermore, our data showed that stimulated MMP-1 secretion was inhibited by actinomycin D and that the nascent MMP-1 mRNA synthesis was stimulated by oxidized LDL, indicating that oxidized LDL activated transcription of the MMP-1 gene. Finally, both zymography and activity assays demonstrated that collagenase activity in conditioned medium was stimulated by oxidized LDL. Taken together, these results have shown for the first time that oxidized LDL stimulates MMP-1 transcription and secretion by vascular endothelial cells, suggesting that oxidized LDL may be a potent stimulator for MMP-1 expression in atherosclerotic plaques, thus promoting plaque rupture.
Severe Hyperlipidemia in Apolipoprotein E2 Homozygotes Due to a Combined Effect of Hyperinsulinemia and an
Sst
I Polymorphism
Authors: Sijbrands Eric J. G. , Hoffer Mariëtte J. V. , Meinders A. Edo , Havekes Louis M. , Frants Rune R. , Smelt Augustinus H. M. , De Knijff Peter
Published: June 2011

Abstract
—More than 90% of patients with type III hyperlipoproteinemia are homozygous carriers of the apolipoprotein (apo) E*2 allele. The great majority of these apoE2(Arg158→Cys) homozygotes in the general population, however, are normolipidemic. Apparently, expression of the hyperlipidemic state requires additional genetic and/or environmental factors, suggesting a multifactorial etiology. To elucidate these additional risk factors, we analyzed normolipidemic and hyperlipidemic apoE2 homozygotes. Hyperinsulinemia was observed in 27 of 49 apoE2 homozygotes and associated with elevated lipid levels: hyperinsulinemic apoE2 homozygotes had type III hyperlipoproteinemia 6 times more often than apoE2 homozygotes with normal insulin levels (odds ratio 6.2,
P
=0.02). We screened the normolipidemic and hyperlipidemic apoE2 homozygotes for common variants in candidate genes involved in lipolysis—the APOA1-C3-A4 gene cluster, lipoprotein lipase, and hepatic lipase—and analyzed for associations with the expression of hyperlipidemia. In the hyperinsulinemic group, the 7 carriers of the
Sst
I polymorphism (S2) in the APOC3 gene displayed severely elevated VLDL cholesterol (
P

insulin by
Sst
I

<0.001) and VLDL triglyceride (
P

insulin by
Sst
I

<0.01) and low levels of HDL (
P

insulin by
Sst
I

<0.02). In the normoinsulinemic group, no such relation of the
Sst
I polymorphism with hyperlipidemia was observed. These data provide the first evidence for a combined effect of hyperinsulinemia and the
Sst
I polymorphism on the expression of hyperlipidemia in apoE2 homozygotes.
Menopause Is Associated With Reduced Protection From Postprandial Lipemia
Authors: van Beek André P. , de Ruijter-Heijstek Florianne C. , Erkelens D. Willem , de Bruin Tjerk W. A.
Published: June 2011

Abstract
—Deficiency of endogenous estrogens has been associated with a higher incidence of coronary heart disease (CHD) in women. We investigated whether natural menopause is associated with reduced protection from postprandial lipemia, which represents a risk indicator of CHD. Twenty-three postmenopausal women (mean age, 50±1 [SD] years; body mass index, 24.6±2.8 kg/m
2
) and 21 premenopausal women matched for age and body mass index (age, 49±1 years; body mass index, 24.1±2.6 kg/m
2
) underwent an oral vitamin A fat-loading test. Vitamin A is a marker of the metabolism of chylomicrons and chylomicron remnants. All women were normolipidemic, were in good health, were nonsmokers, and used no medication. Postprandial lipids and vitamin A were measured at hourly intervals up to 12 hours. In postmenopausal women, plasma total cholesterol and LDL cholesterol concentrations were significantly higher. Fasting plasma triglyceride (TG) concentrations were 1.14±0.57 mmol/L in postmenopausal women and 0.88±0.33 mmol/L in premenopausal women (
P
=NS). In the postprandial phase, postmenopausal women had higher plasma TG (13.0±6.1 versus 9.5±3.3 mmol · L
−1
· h
−1
;
P
=0.024) and vitamin A (54.1±22.9 versus 35.9±9.6 mg · L
−1
· h
−1
;
P
=0.001) responses. To correct for the possible confounding effect of fasting TG, 13 postmenopausal women were carefully matched with 19 premenopausal women. Although fasting TG levels were identical (0.72±0.20 versus 0.73±0.21 mmol/L), differences in postprandial vitamin A (45.3±14.5 versus 33.0±7.7 mg · L
−1
· h
−1
;
P
=0.006) and incremental TG (ie, after subtraction of baseline TG) (3.2±1.8 versus 2.3±1.0 mmol · L
−1
· h
−1
;
P
=0.023) persisted between postmenopausal and premenopausal women. Natural menopause is associated with aggravated postprandial lipemia in women matched for age and body mass index. Higher postprandial lipemia potentially explains the relation of TGs and CHD mortality risk in postmenopausal women.
Coronary Endothelial Function Is Preserved With Chronic Endothelin Receptor Antagonism in Experimental Hypercholesterolemia In Vitro
Authors: Best Patricia J. M. , Lerman Lilach O. , Romero Juan C. , Richardson Darcy , Holmes David R. , Lerman Amir
Published: June 2011

Abstract
—Hypercholesterolemia is associated with increased circulating and tissue endothelin-1 immunoreactivity, decreased nitric oxide (NO) activity, and altered endothelial function. We tested the hypothesis that chronic endothelin receptor antagonism preserves endothelial function and increases NO in experimental porcine hypercholesterolemia. Pigs were randomized to 3 groups: Group 1, a 2% high-cholesterol (HC) diet alone (n=7); group 2, RO-48-5695, a combined endothelin receptor antagonist, and an HC diet (n=8); and group 3, ABT-627, a selective endothelin-A receptor antagonist, and an HC diet (n=8). Coronary epicardial and arteriolar endothelial function was determined by a dose-response relaxation to bradykinin (10
−11
to 10
−6
mol/L)
,
in all groups and in pigs maintained on a normal diet. Plasma total oxidized products of NO (NO
x
) were determined by chemiluminescence at baseline and after 12 weeks. Bradykinin-stimulated coronary epicardial and arteriolar relaxation in group 1 was attenuated compared with normal-diet controls. This relaxation was normalized with endothelin receptor antagonism. Plasma NO
x
decreased after 12 weeks in group 1 (−74.8±5.5%). This decrease was attenuated in the endothelin receptor antagonist groups (group 2, −28.2±15.0%; group 3, −38.9±20.6%). Chronic endothelin receptor antagonism preserves coronary endothelial function and increases NO in hypercholesterolemia. This study supports a role of endothelin-1 in the regulation of NO activity and suggests a possible therapeutic role for endothelin receptor antagonists in hypercholesterolemia.
Exercise Training Increases Basal Nitric Oxide Production From the Forearm in Hypercholesterolemic Patients
Authors: Lewis Tamara V. , Dart Anthony M. , Chin-Dusting Jaye P. F. , Kingwell Bronwyn A.
Published: June 2011

Abstract
—The objective of this study was to investigate the effects of cycle training on basal nitric oxide (NO) production and endothelium-dependent dilator capacity in hypercholesterolemic patients in whom acetylcholine responsiveness is impaired. Nine sedentary hypercholesterolemic volunteers (total plasma cholesterol >6.0 mmol/L; 2 female) aged 44±3 years (mean±SEM) participated in the study. Subjects remained sedentary for 4 weeks and performed 4 weeks of home-based cycle training (3×30 minutes/week at 65% maximum oxygen consumption [VO
2
max]) in a randomized order. Arteriovenous nitrate/nitrite (NO
x
) gradient was assessed and plethysmography was used to measure the forearm blood flow responses to arterial infusions of acetylcholine, sodium nitroprusside, and N
G
mono methyl L-arginine. Training increased VO
2
max from 30.4±1.9 to 34.3±1.4 mL · kg
−1
· min
−1
(
P
=0.01). Intrabrachial diastolic blood pressure was reduced from 70±3 to 68±3 mm Hg (
P
=0.02) with training, whereas systolic pressure did not change. Plasma triglycerides and total, LDL, and HDL cholesterol were not different between interventions. In the sedentary state, there was a positive forearm arteriovenous difference in plasma NO
x
indicating net extraction (6.8±4.0 nmol · 100 mL
−1
· min
−1
), whereas in the trained state this difference was negative, indicating net production (−5.8±5.8 nmol · 100 mL
−1
· min
−1
;
P
=0.03). N
G
mono methyl L-arginine, at a dose of 4 μmol/min, caused a greater vasoconstriction after training (79.6±3.4% versus 69.9±6.8%;
P
=0.05). Acetylcholine and sodium nitroprusside induced dose-dependent elevations in forearm blood flow that were unaffected by training. These data suggest that basal release of endothelium-derived NO is increased with 4 weeks of home based training in hypercholesterolemic patients, independently of lipid profile modification. This may contribute to the cardiovascular protective effects of exercise training, including reduced blood pressure.
Stretch Induces Mitogen-Activated Protein Kinase Activation and Myogenic Tone Through 2 Distinct Pathways
Authors: Loufrani Laurent , Lehoux Stéphanie , Tedgui Alain , Lévy Bernard I. , Henrion Daniel
Published: June 2011

Abstract
—The aim of this study was to evaluate the involvement of the mitogen-activated protein kinase (ERK1/2) pathway in response to stretch in a blood vessel developing myogenic tone on stretch. Indeed, in resistance arteries and veins, the main effect of pressure is to induce a maintained vasoconstrictor (myogenic) tone. Isolated segments of rabbit facial vein were mounted in organ baths and submitted to isometric stretch. In this experimental model, myogenic tone was absent when the bath temperature was 33°C. ERK1/2 activity was determined in each isolated segment by an in-gel kinase assay. Wall tension and ERK1/2 activity were measured in the same samples in the presence (39°C) or in the absence of myogenic tone (33°C). At 39°C, a 5-mN wall tension induced myogenic tone (5.7±1.8 mN) and an increase in ERK1/2 activity (282±52% versus unstretched vessels,
P
<0.05). At 33°C, in the absence of myogenic tone, ERK1/2 activity was similarly increased by stretch (254±35% versus unstretched vessels). The calcium-dependent and -independent protein kinase C (PKC) blocker Ro-31-8220 (5×10
−7
mol/L), but not the calcium-dependent PKC blocker Go-6976 (10
−6
mol/L), inhibited myogenic tone. However, ERK1/2 activity was not affected by either PKC blocker. Genistein (10
−7
mol/L), a general tyrosine kinase inhibitor, but not herbimycin A (5×10
−7
mol/L), a cSrc-family tyrosine kinase inhibitor, suppressed stretch-induced ERK1/2 activation (
P
<0.05) without affecting myogenic tone. Nifedipine (10
−6
mol/L), a voltage-dependent calcium entry inhibitor, and ryanodine (10
−6
mol/L), which depletes calcium stores, both inhibited ERK1/2 activity (113±12% and 121±7%, respectively;
P
<0.05) without affecting myogenic tone. The mitogen-activated protein kinase kinase inhibitor PD 98059 (5×10
−6
mol/L) also inhibited ERK1/2 activation without affecting myogenic tone. The present results suggest that stretching the rabbit facial vein induced 2 distinct pathways, one leading to myogenic tone (via a non–calcium-dependent PKC activation) and one leading to ERK1/2 activation through a calcium-dependent pathway involving tyrosine kinase.
Simvastatin Inhibits Leukocyte–Endothelial Cell Interactions and Protects Against Inflammatory Processes in Normocholesterolemic Rats
Authors: Pruefer Diethard , Scalia Rosario , Lefer Allan M.
Published: June 2011

Abstract
—Simvastatin, a 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, has been shown to lower serum cholesterol levels and normalize endothelial cell function. Moreover, HMG-CoA reductase inhibitors exert beneficial effects in coronary artery and cerebrovascular diseases. We examined the effects of simvastatin on leukocyte–endothelial cell interaction in vivo by intravital microscopy. Simvastatin (12.5 or 25 μg per rat) was given 18 hours before study. Superfusion with the NO synthase inhibitor
N
G
-nitro-
l
-arginine methyl ester (L-NAME, 50 μmol/L) significantly increased leukocyte rolling from 12±2 to 60±8 leukocytes per minute, increased adherence to the mesenteric endothelium from 1.8±0.5 to 17±1.2 leukocytes per 100 μm of venular length, and raised leukocyte transmigration from 2.5±1.0 to 10±2 leukocytes per perivessel area (
P
<0.01). Similar results were obtained with thrombin (0.5 U/mL) superfusion of the mesentery. In contrast, pretreatment with simvastatin (25 μg per rat IP) significantly attenuated L-NAME–stimulated leukocyte rolling, to 12±2 (
P
<0.01); adherence, to 5±0.5 leukocytes per 100 μm (
P
<0.01); and leukocyte transmigration, to 3.5±1.5 leukocytes per perivessel area (
P
<0.01). Similar results were obtained in thrombin-superfused mesenteries. Moreover, immunohistochemical analysis demonstrated significantly increased P-selectin expression on the mesenteric venular endothelium after superfusion with either L-NAME (
P
<0.01) or thrombin (
P
<0.01), which was significantly attenuated by simvastatin. These results clearly demonstrate that simvastatin is a potent and effective endothelium-protective agent that reduces leukocyte–endothelial cell interactions independently of its well-known lipid-lowering effects. This effect was found to be at least partially mediated via downregulation of P-selectin expression on the microvascular endothelium. Thus, HMG-CoA reductase inhibitors like simvastatin have important anti-inflammatory effects besides their well-known lipid-lowering action.
25-Hydroxycholesterol Increases Eicosanoids and Alters Morphology in Cultured Pulmonary Artery Smooth Muscle and Endothelial Cells
Authors: Wohlfeil Eric R. , Campbell William B.
Published: June 2011

Abstract
—25-Hydroxycholesterol (25-OHC) is an oxidized derivative of cholesterol that has been implicated in the early development of arteriosclerosis. Changes in arterial smooth muscle cell (SMC) migration and proliferation have also been linked to the pathophysiology of arteriosclerosis. SMCs undergo “activation” in response to vascular injury by changing phenotypically and by increasing prostaglandin G/H synthase-2 (PGHS-2) protein levels and eicosanoid release. Activation is thought to be important in atheroma formation and arteriosclerosis progression. 25-OHC induces SMCs to change morphologically, increase PGHS-2, and increase eicosanoid release. Confluent monolayers were treated with 25-OHC (10 μg/mL) or the PGHS-2 inducer interleukin-1β (1 ng/mL) for 18 hours at 37°C. The 18-hour treatment resulted in morphological changes. After uptake of [
14
C]arachidonic acid, released radiolabeled arachidonic acid products were extracted and chromatographed by both normal and reverse-phase high-performance liquid chromatography systems. 25-OHC–treated cells increased their prostaglandin production, with the major component comigrating with a prostaglandin-E
2
standard. HETEs and epoxyeicosatrienoic acids were not affected. Immunoprecipitation analysis of treated and control cell lysates using anti–PGHS-1 and -2 and anti–α-actin primary antibodies indicated PGHS-2 induction over control and no change in contractile proteins. These changes are consistent with SMC activation, which occurs in vascular injury models. The notion that oxysterols can activate vascular SMCs may be important in ultimately understanding the pathophysiology of atheroma formation.
Enhanced Reduction of Fasting Total Homocysteine Levels With Supraphysiological Versus Standard Multivitamin Dose Folic Acid Supplementation in Renal Transplant Recipients
Authors: Beaulieu Andrew J. , Gohh Reginald Y. , Han Haewook , Hakas David , Jacques Paul F. , Selhub Jacob , Bostom Andrew G.
Published: June 2011

Abstract
—The mild fasting hyperhomocysteinemia commonly observed in chronic (ie, ≥6 months posttransplantation) renal transplant recipients (RTRs) can be effectively treated with combined B-vitamin supplementation featuring supraphysiological doses of folic acid. There are no controlled data evaluating the comparative efficacy of supraphysiological versus standard multivitamin dose folic acid supplementation in reducing fasting total homocysteine (tHcy) levels among RTRs. We block-randomized 60 chronic, stable RTRs on the basis of their screening fasting tHcy level to 3 groups of 20 subjects treated for 12 weeks with folic acid at either 2.4 (group 1), 0.4 (ie, standard multivitamin dose) (group 2), or 0.0 (group 3) mg/d. All 60 study participants also received 50 mg/d vitamin B
6
and 0.4 mg/d vitamin B
12
. The mean percent reductions (±SEM) in fasting tHcy were as follows: group 1, 32.3±2.4%; group 2, 23.4±2.3%; and group 3, 19.1±2.3%. ANCOVA accounting for the pretreatment matching and adjusted for pretreatment levels of fasting tHcy, folate, and albumin; change in creatinine during the study; and cyclosporine A use revealed significant overall group differences (
P
=0.005) and significant differences between groups 1 and 2 (
P
=0.038) and groups 1 and 3 (
P
=0.001), but not between groups 2 and 3 (
P
=0.153). Moreover, a χ
2
analysis of participants with pretreatment tHcy levels ≥15 μmol/L (n=29) indicated that a significantly greater proportion of those in group 1 achieved posttreatment levels <12 μmol/L: group 1, 5 of 10 (50%); group 2, 1 of 11 (9%); and group 3, 0 of 8 (0%) (
P
=0.016; test of trend
P
=0.007). We conclude that a supraphysiological dose of folic acid is superior to standard multivitamin dosing for the reduction of fasting tHcy levels in chronic RTRs.
Peroxisome Proliferator–Activated Receptor γ Gene Locus Is Related to Body Mass Index and Lipid Values in Healthy Nonobese Subjects
Authors: Knoblauch Hans , Busjahn Andreas , Müller-Myhsok Bertram , Faulhaber Hans-Dieter , Schuster Herbert , Uhlmann Regina , Luft Friedrich C.
Published: June 2011

Abstract
—The peroxisome proliferator–activated receptor γ (
PPARγ
) gene has been implicated in morbid obesity and is important to lipid and carbohydrate metabolism. However, the relevance of gene variations in healthy nonobese subjects has not been defined. We recruited monozygotic and dizygotic healthy nonobese twin subjects to test the hypothesis that the
PPARγ
gene is important to body mass index and lipid concentrations in healthy nonobese subjects. Both linkage and association strategies were used in the same dizygotic twins. The
PPARγ
gene locus was linked (
P
<0.01) to high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, and body mass index as quantitative traits. A biallelic variant in the
PPARγ
gene was associated with high-density lipoprotein cholesterol and body mass index (
P
<0.05). We also looked for linkage between the same variables and the retinoic X receptor gene locus. This locus was linked to total and low-density lipoprotein cholesterol as well as triglycerides. We conclude that the
PPARγ
gene is highly relevant to lipid metabolism and body mass index, not only in the morbidly obese but also in healthy nonobese subjects. The same appears to be true for its binding partner. Sequencing these genes in twins would serve to identify gene variations contributing to body mass index and lipid concentrations in healthy nonobese subjects.
Both Raloxifene and Estrogen Reduce Major Cardiovascular Risk Factors in Healthy Postmenopausal Women
Authors: de Valk-de Roo Gerdien W. , Stehouwer Coen D.A. , Meijer Piet , Mijatovic Velja , Kluft Cornelis , Kenemans Peter , Cohen Fredric , Watts Steven , Netelenbos Coen
Published: June 2011

Abstract
—Currently raloxifene, a selective estrogen receptor modulator, is being investigated as a potential alternative for postmenopausal hormone replacement to prevent osteoporosis and cardiovascular disease. We compared the 2-year effects of raloxifene on a wide range of cardiovascular risk factors with those of placebo and conjugated equine estrogens (CEEs). Analyses were based on 56 hysterectomized but otherwise healthy postmenopausal women aged 54.8±3.5 (mean±SD) years who entered this double-blind study and who were randomly assigned to raloxifene hydrochloride 60 mg/d (n=15) or 150 mg/d (n=13), placebo (n=13), or CEEs 0.625 mg/d (n=15). At baseline and after 6, 12, and 24 months of treatment, we assessed serum lipids, blood pressure, glucose metabolism, C-reactive protein, and various hemostatic parameters. Compared with placebo, both raloxifene and CEEs lowered the level of low density lipoprotein cholesterol by 0.53 to 0.79 mmol/L (all
P
<0.04) and lowered, at 24 months, the level of fibrinogen by 0.71 to 0.86 g/L (all
P
<0.05). The effects of raloxifene and CEEs did not differ significantly. In contrast to raloxifene, from 6 months on CEEs increased high density lipoprotein cholesterol by 0.25 to 0.29 mmol/L and reduced plasminogen activator inhibitor-1 antigen by 30.6 to 48.6 ng/mL (all
P
<0.02 versus both placebo and raloxifene). CEEs transiently increased C-reactive protein by 1.0 mg/L at 6 months (
P
<0.05 versus placebo) and prothrombin-derived fragment F1+2 by 0.79 nmol/L at 12 months (
P
<0.001 versus placebo). Finally, from 12 months on, CEEs increased triglycerides by 0.33 to 0.56 mmol/L (all
P
<0.05 versus both placebo and raloxifene). Our findings suggest that in healthy postmenopausal women, raloxifene and estrogen monotherapy have similar beneficial effects on low density lipoprotein cholesterol and fibrinogen levels. These treatments differ, however, in their effects on high density lipoprotein cholesterol, triglycerides, and plasminogen activator inhibitor-1 and possibly in their effects on prothrombin fragment F1+2 and C-reactive protein.
Age and Sex Differences in the Distribution and Ultrasound Morphology of Carotid Atherosclerosis
Authors: Joakimsen Oddmund , Bønaa Kaare H. , Stensland-Bugge Eva , Jacobsen Bjarne Koster
Published: June 2011

Abstract
—Atherosclerosis begins early in life and is the major underlying cause of cardiovascular morbidity and death. Yet, population-based information on age and sex differences in the extent and morphology of atherosclerosis throughout life is scarce. Carotid atherosclerosis can be visualized with B-mode ultrasound and is a marker of atherosclerosis elsewhere in the circulation. We assessed both the prevalence and the morphology of carotid atherosclerosis by B-mode ultrasound in 3016 men and 3404 women, 25 to 84 years old, who participated in a population health survey. The participation rate was 88%. Plaque morphology was graded according to whether a plaque was predominantly soft (echolucent) or hard (echogenic). Atherosclerotic plaques were found in 55.4% of the men and 45.8% of the women. In men, there was a linear increase with age in the prevalence of carotid atherosclerosis, whereas in women, there was a curvilinear age trend, with an inflection in the prevalence rate of women at ≈50 years of age. The male predominance in atherosclerosis declined after the age of 50 years, the plaque prevalence being similar in elderly men and women. Men had softer plaques than women; this sex difference in plaque morphology increased significantly (
P
=0.005) with age. The sex difference in the prevalence of atherosclerosis and the female age trend in atherosclerosis show significant changes at the age of ≈50 years, suggesting an adverse effect of menopause on atherosclerosis. The higher proportion of soft plaques in men compared with women increases with age and may partly account for the prevailing male excess risk of coronary heart disease in the elderly despite a similar prevalence of atherosclerosis in elderly men and women.
In Vivo Regulation of von Willebrand Factor Synthesis
Authors: Brouland Jean-Philippe , Egan Thomas , Roussi Jacqueline , Bonneau Michel , Pignaud Georges , Bal Claire , Vaiman Marcel , André Patrick , Hervé Philippe , Mazmanian Guy Michel , Drouet Ludovic
Published: June 2011

Abstract—To evaluate the regulation of plasma von Willebrand factor (vWF) and its in situ production by endothelial cells (ECs), 12 swine leukocyte antigen (SLA)-compatible left lung transplantations were performed. Normal lungs were transplanted into 10 pigs homozygous for von Willebrand disease and into 2 normal pigs. Additionally, 1 normal pig underwent pneumonectomy, and 1 SLA-incompatible lung transplantation between normal pigs was performed. None of the transplanted animals received immunosuppressive therapy. Plasma vWF level was evaluated by ELISA and multimeric pattern. EC vWF content was assessed by immunohistochemistry. Global hemostasis was assessed by standardized ear bleeding time. Six of 12 SLA-compatible lung transplantations and the incompatible transplantation were successful and were used for the study. The functions and the viability of ECs, reflected by their ability to produce vWF and normal multimeric plasma vWF pattern, were preserved in SLA-compatible and -incompatible lung transplantations. vWF production was preserved in ECs that initially synthesized it. EC constitutive and storage pathways are modulated differently according to transplantation compatibility and severity of rejection. In SLA-compatible lung transplantations without histological evidence of rejection, the production of vWF was preserved, whereas constitutive vWF secretion appeared to be altered in cases with minor histological signs of rejection. In pigs with von Willebrand disease that were transplanted with normal lungs without sign of rejection, plasma vWF was significantly increased in an amount expected from the estimated production of a normal lung. In the transplanted normal lung, there was no vWF overexpression by the ECs and no recruitment of ECs that initially did not express vWF. In SLA-incompatible transplantation, ECs were morphologically normal with increased and blurred vWF labeling, whereas plasma vWF levels remained normal, reflecting that EC activation is associated with an increased vWF production with probable diversion to storage pathway. This model depicts the changes of EC regulation of vWF secretion in pig lung transplants. However, this model cannot be directly extrapolated to human organ transplantation because animals did not receive any immunosuppressive therapy, which may be toxic to ECs.
Oleic Acid Inhibits Endothelial Activation
Authors: Carluccio Maria Annunziata , Massaro Marika , Bonfrate Cosima , Siculella Luisa , Maffia Michele , Nicolardi Giuseppe , Distante Alessandro , Storelli Carlo , Raffaele De Caterina
Published: June 2011

Abstract—Because oleic acid is implicated in the antiatherogenic effects attributed to the Mediterranean diet, we investigated whether this fatty acid can modulate endothelial activation, ie, the concerted expression of gene products involved in leukocyte recruitment and early atherogenesis. We incubated sodium oleate with human umbilical vein endothelial cells for 0 to 72 hours, followed by coincubation of oleate with human recombinant tumor necrosis factor, interleukin (IL)-1α, IL-1β, IL-4,Escherichia colilipopolysaccharide (LPS), or phorbol 12-myristate 13-acetate for a further 6 to 24 hours. The endothelial expression of vascular cell adhesion molecule-1 (VCAM-1), E-selectin, and intercellular adhesion molecule-1 was monitored by cell surface enzyme immunoassays or flow cytometry, and steady-state levels of VCAM-1 mRNA were assessed by Northern blot analysis. At 10 to 100 μmol/L for >24 hours, oleate inhibited the expression of all adhesion molecules tested. After a 72-hour incubation with oleate and a further 16-hour incubation with oleate plus 1 μg/mL LPS, VCAM-1 expression was reduced by >40% compared with control. Adhesion of monocytoid U937 cells to LPS-treated endothelial cells was reduced concomitantly. Oleate also produced a quantitatively similar reduction of VCAM-1 mRNA levels on Northern blot analysis and inhibited nuclear factor-κB activation on electrophoretic mobility shift assays. Incubation of endothelial cells with oleate for 72 hours decreased the relative proportions of saturated (palmitic and stearic) acids in total cell lipids and increased the proportions of oleate in total cell lipids without significantly changing the relative proportions of polyunsaturated fatty acids. Although less potent than polyunsaturated fatty acids in inhibiting endothelial activation, oleic acid may contribute to the prevention of atherogenesis through selective displacement of saturated fatty acids in cell membrane phospholipids and a consequent modulation of gene expression for molecules involved in monocyte recruitment.
Adventitial Angiogenesis Early After Coronary Angioplasty
Authors: Pels Klaus , Labinaz Marino , Hoffert Cyrla , O’Brien Edward R.
Published: June 2011

Abstract
—The spatial correlation between arterial wall microvessels and the accumulation of atherosclerotic plaque is well documented. The role of these microvessels in the development of primary and restenotic lesions is not known. To investigate the effect of interventional procedures on arterial wall microvessels, we studied the adventitial microvascularity of porcine coronary arteries subjected to angioplasty. Twenty-two juvenile domestic swine were subjected to single or repeated (double) balloon angioplasty of the coronary arteries, with the interval between the first and second injury being 14 days. The number, density, and size of adventitial microvessels were measured 1 hour as well as 3, 7, 14, and 28 days after injury. One hour after single balloon injury, there were very few intact adventitial microvessels. Adventitial microvessel number, microvessel area density, and microvessel size were maximal 3 days after single (SI) and double (DI) injury but subsequently underwent progressive regression. Adventitial endothelial cell replication, as assessed by the incorporation of bromodeoxyuridine, was very low for the majority of arteries. Maximal endothelial cell replication indices were observed 3 days after SI and DI (eg, 12.0±3.3%). Early after SI the central arterial lumen area transiently increased, then renarrowed. The lumen area did not change after DI. Arterial remodeling occurred, as the accumulation of intimal and medial mass was correlated with expansion of the external elastic lamina. Adventitial microvessel area density was correlated with central arterial luminal area (
R
=0.34,
P
=0.04). The adventitial microvessel area density and the microvessel size index were greater late after DI compared with SI. These data indicate that adventitial angiogenesis occurs within 3 days after balloon injury and that regression of adventitial microvessels after SI corresponds with arterial narrowing. Changes in the adventitial microvasculature may be a component of arterial remodeling after balloon angioplasty.
Urokinase Activates the Jak/Stat Signal Transduction Pathway in Human Vascular Endothelial Cells
Authors: Dumler Inna , Kopmann Angela , Weis Angelika , Mayboroda Oleg A. , Wagner Kai , Gulba Dietrich C. , Haller Hermann
Published: June 2011

Abstract—Endothelial cells demonstrate high urokinase expression and upregulation of urokinase receptors in response to vascular injury. Urokinase receptor binding facilitates endothelial cell migration into an arterial wound; however, the signaling cascade induced by the urokinase receptor in this cell type is incompletely understood. Because the Janus kinase (Jak)/signal transducer and activator of transcription (Stat) pathway seems to be important for vessel function, we investigated the hypothesis that urokinase receptor binding activates Jak/Stat signaling in human vascular endothelial cells. Incubation of endothelial cells with urokinase-type plasminogen activator (uPA,1 nmol/L) induced a rapid and pronounced increase in tyrosine phosphorylation of several proteins with a molecular weight between 80 to 90 and 130 to 140 kDa. The same pattern of tyrosine phosphorylation was found after treatment with 1 nmol/L ATF, the urokinase amino-terminal fragment, which is devoid of proteolytic activity but still binds to the urokinase receptor. Using coimmunoprecipitation techniques, we demonstrated that the activated urokinase receptor is associated with 2 cytoplasmic tyrosine kinases of the Jak family, viz, Jak1 and Tyk2. uPA and ATF induced a time-dependent activation of both kinases, as shown by immunoprecipitation and Western blot analysis. Using electrophoretic mobility shift and supershift assays, we then demonstrated that Stat1 is rapidly activated in endothelial cells in response to uPA and ATF. Furthermore, Stat1 specifically binds to the regulatory elements interferon-γ activation site/interferon-stimulated response element. The uPA-induced, time-dependent translocation of Stat1 to cell nuclei was confirmed by confocal microscopy study and immunoblotting of nuclear extracts with an anti-Stat1 antibody. This study provides evidence for a novel signaling pathway for uPA in human vascular endothelial cells. Direct activation of the Jak/Stat system via the uPA-receptor complex may be an important mechanism for endothelial cell migration and/or proliferation during angiogenesis and after vascular injury.
Tissue Factor Pathway Inhibitor Is Expressed by Human Monocyte–Derived Macrophages
Authors: Petit Laure , Lesnik Philippe , Dachet Christiane , Moreau Martine , Chapman M. John
Published: June 2011

Abstract—Lipid-laden macrophages express tissue factor (TF), which may activate the extrinsic coagulation pathway on rupture of the atherosclerotic plaque. Tissue factor pathway inhibitor (TFPI) is a major regulator of TF-induced coagulation. We evaluated the possibility that monocyte-derived macrophages express this protein, thereby contributing to regulation of TF activity (TFact). Equally, we investigated the effect of cholesterol and of oxidized LDL (Ox-LDL) on the expression of TFPI and TF by human monocyte–derived macrophages (HMDMs). Northern blot analysis of TFPI mRNA from cultured HMDMs revealed a single band at 4.2 kb with weak intensity; this finding was confirmed by reverse transcription–polymerase chain reaction. Gel filtration of HMDM supernatants showed the presence of an active 100-kDa form of TFPI, which was confirmed by sodium dodecyl sulfate–polyacrylamide gel electrophoresis under nonreducing conditions; under reducing conditions, however, the immunoblot revealed a 40-kDa form of TFPI. The TFPI in HMDM supernatants possessed heparin-binding affinity, suggesting potential interaction of TFPI with heparan sulfate proteoglycans. Stimulation of foam cell formation by incubation of macrophages for 48 hours with exogenous free cholesterol indicated that neither the biological activity nor the de novo synthesis of TFPI protein was affected. In contrast, cholesterol loading with exogenous free cholesterol induced significant upregulation of total TFact (2.6-fold: 25.0 versus 9.4 mU/mg cell protein, cholesterol-treated versus control cells;P<0.05); such induction was not correlated with an elevation in TF antigen (8.5 versus 7.8 ng/mg cell protein, cholesterol-treated versus control cells). Similarly, cholesterol-rich Ox-LDL induced an increase in TFact (1.9-fold: 18.9 versus 10.0 mU/mg cell protein, Ox-LDL–treated versus control cells;P<0.05); by contrast, the amount of TF antigen remained unchanged (7.1 versus 7.9 ng/mg cell protein, Ox-LDL–treated versus control cells). Our data indicate that enhancement of the procoagulant activity of TF in macrophage-derived foam cells is not counterbalanced by upregulation of TFPI activity, suggesting that lesion foam cells are in a procoagulant state; they may therefore contribute to thrombus generation on plaque rupture.
Compound Heterozygosity for an Apolipoprotein A1 Gene Promoter Mutation and a Structural Nonsense Mutation With Apolipoprotein A1 Deficiency
Authors: Matsunaga Akira , Sasaki Jun , Han Hua , Huang Wei , Kugi Mari , Koga Takafumi , Ichiki Sadanori , Shinkawa Tomoko , Arakawa Kikuo
Published: June 2011

Abstract
—Apolipoprotein (apo) A1 plays a central role in the metabolism of HDL. We describe a novel genetic variant of the apoA1 gene identified in a patient with low concentrations of plasma HDL cholesterol. The proband, a 12-year-old Japanese boy, exhibited markedly low levels of both plasma apoA1 and HDL cholesterol. Genomic DNA sequencing of apoA1 genes of the patient showed a compound heterozygosity for an A to C substitution at 27 bp upstream of the transcription start site of 1 apoA1 allele, and a C to T substitution in another allele at residue 84 resulting in aberrant termination. The point mutation at nucleotide position –27 changed ATAAATA of the putative TATA box signal sequence to ATACATA. In addition to this mutation, the patient was heterozygous for a G to A substitution at position –75. Immunoblotting of an isoelectric focusing electrophoresis gel of the proband’s plasma showed a trace amount of normal apoA1. No measurable plasma apoA1 and HDL cholesterol in a patient with homozygosity for nonsense mutation at residue 84 has been reported previously. To determine the effects of substitution either at position –27 or –75, plasmids containing the 5′-flanking region of the human apoA1 promoter fused to the CAT reporter gene were constructed and transfected in HepG2 cells. A construct with the A to C substitution at position –27 showed 41.8±4.2%, and G to A substitution at position –75 showed 72.8±15.2% (means±SD, n=3) of CAT activities, compared with the wild-type promoter sequence. A construct with the double substitutions at positions –27 and –75 showed only 22.8±1.3% (mean±SD, n=3) activity relative to the wild type. Our patient is the first case with a TATA box mutation etiologically related to lipoprotein disorders.
Relationships of Cerebral MRI Findings to Ultrasonographic Carotid Atherosclerosis in Older Adults
Authors: Manolio Teri A. , Burke Gregory L. , O’Leary Daniel H. , Evans Gregory , Beauchamp Norman , Knepper Laurie , Ward Beverly
Published: June 2011

Abstract
—Cerebral magnetic resonance imaging (MRI) has demonstrated a high prevalence of infarct-like lesions, white matter hyperintensities, and evidence of cerebral atrophy in older adults. While these findings are generally believed to be related to ischemia and atherosclerosis, their relationship to atherosclerosis in the carotid arteries remains to be explored. Study subjects were part of the multicenter Cardiovascular Health Study, a cross-sectional study of 3502 women and men ≥65 years of age undergoing cranial MRI and carotid ultrasonography. MRI infarcts were detected in 1068 participants (29.3%) and measurable carotid plaque in 2745 (75.3%). MRI infarcts, ventricular and sulcal widening, and white matter score were strongly associated with carotid intimal-medial thickness (IMT) and stenosis degree after adjustment for age and sex (all
P
<0.01). Associations with plaque characteristics were less strong and less consistent; MRI infarcts were weakly associated only with surface irregularity, and ventricular size was weakly associated only with lesion density (both
P
<0.04). In contrast, sulcal widening was strongly related to plaque characteristics, with scores being higher in those with heterogeneous and irregular plaque (both
P
<0.009). Adjustment for other risk factors, and for carotid IMT/stenosis, removed associations of MRI findings with plaque characteristics except for weak relationships remaining between MRI infarcts and surface irregularity and between sulcal score and heterogeneous plaque (both
P
<0.03). MRI abnormalities show strong and consistent relationships with increasing carotid IMT and stenosis degree but less strong associations with plaque characteristics, especially after adjusting for IMT and stenosis.
Focal Increases in Vascular Cell Adhesion Molecule-1 and Intimal Macrophages at Atherosclerosis-Susceptible Sites in the Rabbit Aorta After Short-Term Cholesterol Feeding
Authors: Truskey George A. , Herrmann Robert A. , Kait Jason , Barber Kevin M.
Published: June 2011

Abstract
—We tested the hypotheses that vascular cell adhesion molecule-1 (VCAM-1) expression on endothelium at lesion-prone sites in the rabbit aorta correlates with exposure to plasma cholesterol and that macrophage accumulation is associated with endothelial cells expressing VCAM-1. After rabbits were fed 0.25% cholesterol for 2 weeks, VCAM-1 expression was selectively increased at the distal and lateral portions of the major abdominal branches. In the arch and the celiac, superior mesenteric, and renal artery branches, VCAM-1 expression was positively correlated with the plasma cholesterol integrated over the duration of the experiments. After 2 weeks of cholesterol feeding, more macrophages were present around distal and lateral portions of the intercostal arteries and major abdominal branches relative to nonbranch regions. In the arch and around the intercostals and major abdominal branches, macrophage densities were positively correlated with the integrated plasma cholesterol. VCAM-1 and macrophage levels were correlated in lesion-prone regions. In normocholesterolemic rabbits, 23±4% (mean±SEM) of the macrophages were directly associated with VCAM-1–positive endothelium. After 2 weeks of 0.25% cholesterol feeding, the association increased to 37±4% (
P
<0.015). Associations were highest around the lateral and distal regions of the major abdominal branches. These results suggest that (1) VCAM-1 expression and intimal macrophage densities are influenced by plasma cholesterol and regional factors such as arterial fluid dynamics and (2) VCAM-1 plays a significant role in the localization of macrophages.
Analysis of Macrophage Scavenger Receptor (SR-A) Expression in Human Aortic Atherosclerotic Lesions
Authors: Gough Peter J. , Greaves David R. , Suzuki Hiroshi , Hakkinen Tomi , Hiltunen Mikko O. , Turunen Mikko , Herttuala Seppo Ylä , Kodama Tatsuhiko , Gordon Siamon
Published: June 2011

Abstract
—The class A scavenger receptors (SR-As) are trimeric, integral membrane glycoproteins that exhibit unusually broad ligand-binding properties. A number of studies have suggested that these receptors may play an important role in host defense and in many macrophage-associated pathological processes, including atherosclerosis and Alzheimer’s disease. The study of the expression and function of these receptors in human disease has been hampered by the lack of suitable antibodies recognizing human SR-A. This has generated questions regarding the nature of receptors responsible for scavenger receptor activity detected in a variety of cell types, including monocytes, macrophages, smooth muscle cells, and endothelial cells. To address these questions, we have produced high-titer antisera recognizing human SR-A by using mice deficient for SR-A (SR-A −/−). We show that SR-A −/− mice produce a significantly higher-titer immune response than do wild-type (SR-A +/+) littermates, with antisera of the former having a broad species reactivity and recognizing SR-A from humans, mice, and rabbits. The antisera recognize both type I and II SR-A in a wide range of immunological techniques. Using these antisera we show that the expression of SR-A protein is induced during monocyte to macrophage differentiation and that SR-A mediates 80% of the uptake of acetylated low density lipoprotein by human monocyte–derived macrophages. We also establish that human SR-A is expressed by tissue macrophages in liver and lung and by macrophage-derived foam cells within aortic atherosclerotic lesions, with little detectable expression by smooth muscle cells or aortic endothelium.
Ankle-Arm Index as a Predictor of Cardiovascular Disease and Mortality in the Cardiovascular Health Study
Authors: Newman Anne B. , Shemanski Lynn , Manolio Teri A. , Cushman Mary , Mittelmark Maurice , Polak Joseph F. , Powe Neil R. , Siscovick David
Published: June 2011

Abstract
—Peripheral arterial disease (PAD) in the legs, measured noninvasively by the ankle-arm index (AAI) is associated with clinically manifest cardiovascular disease (CVD) and its risk factors. To determine risk of total mortality, coronary heart disease, or stroke mortality and incident versus recurrent CVD associated with a low AAI, we examined the relationship of the AAI to subsequent CVD events in 5888 older adults with and without CVD. The AAI was measured in 5888 participants ≥65 years old at the baseline examination of the Cardiovascular Health Study. All participants had a detailed assessment of prevalent CVD and were contacted every 6 months for total mortality and CVD events (including CVD mortality, fatal and nonfatal myocardial infarction, congestive heart failure, angina, stroke, and hospitalized PAD). The crude mortality rate at 6 years was highest (32.3%) in those participants with prevalent CVD and a low AAI (
P
<0.9), and it was lowest in those with neither of these findings (8.7%,
P
<0.01). Similar patterns emerged from analysis of recurrent CVD and incident CVD. The risk for incident congestive heart failure (relative risk [RR]=1.61) and for total mortality (RR=1.62) in those without CVD at baseline but with a low AAI remained significantly elevated after adjustment for cardiovascular risk factors. Hospitalized PAD events occurred months to years after the AAI was measured, with an adjusted RR of 5.55 (95% CI, 3.08 to 9.98) in those at risk for incident events. A statistically significant decline in survival was seen at each 0.1 decrement in the AAI. An AAI of <0.9 is an independent risk factor for incident CVD, recurrent CVD, and mortality in this group of older adults in the Cardiovascular Health Study.