bluuebunny/crossref_metadata_2025 huggingface dataset:
The Role of Interleukin 12 in the Development of Atherosclerosis in ApoE-Deficient Mice
Authors: Lee Tzong-Shyuan , Yen Hwei-Chen , Pan Ching-Chien , Chau Lee-Young
Published: June 2011

Abstract—The cytokine profile of atherosclerotic aortas from apoE-deficient mice was assessed by reverse transcriptase-polymerase chain reaction. The results clearly showed that the expression of mRNA for IL-12p40 was evident in aortas from 3-month-old apoE-deficient mice. The mRNA for IL-10 was detected in aorta from these mice at the age of 6 months, indicating that expression of IL-12 is earlier than that of IL-10 in these animals. Concurrent with IL-12p40, the mRNA for the T-cell cytokine IFN-γ, but not IL-4, was detected in aortas of mice at young and old ages. Both in situ hybridization and immunostaining further demonstrated the localization of IL-12 in macrophages of atherosclerotic lesions. Immunohistochemistry also demonstrated the expression of costimulatory molecules B7–1 and B7–2 in macrophages, suggesting that activation of T lymphocytes by macrophages may occur via surface antigens in lesions. When the immunoglobulin isotype of the antioxidized LDL antibodies in sera of apoE-deficient mice was determined, it revealed that both IgM and IgG were present. Furthermore, IgG2a is predominant and comprises ≈50% of the antioxidized LDL IgG in sera from young mice (3 months), but decreased to lower levels (35%) in older mice (6 months). Daily administration of IL-12 led to an increase in serum levels of antioxidized LDL antibodies and accelerated atherosclerosis in young apoE-deficient mice compared with control mice injected with PBS alone. Taken together, these data suggest that IL-12 plays an active role in regulating the immune response during the early phase of atherosclerosis in apoE-deficient mice.
Characterization of a New Form of Inherited Hypercholesterolemia
Authors: Zuliani G. , Arca M. , Signore A. , Bader G. , Fazio S. , Chianelli M. , Bellosta S. , Campagna F. , Montali A. , Maioli M. , Pacifico A. , Ricci G. , Fellin R.
Published: June 2011

Abstract
—We previously described a Sardinian family in which the probands had a severe form of hypercholesterolemia, suggestive of familial hypercholesterolemia (FH). However, low density lipoprotein (LDL) receptor activity in fibroblasts from these subjects and LDL binding ability were normal. The characteristics of the pedigree were consistent with an autosomal recessive trait. Sitosterolemia and pseudohomozygous hyperlipidemia were ruled out. A second Sardinian kindred with similar characteristics was identified. Probands showed severe hypercholesterolemia, whereas their parents and grandparents were normolipidemic. FH, familial defective apoprotein (apo) B, sitosterolemia, and cholesteryl ester storage disease were excluded by in vitro studies. We addressed the metabolic basis of this inherited disorder by studying the in vivo metabolism of LDL in 3 probands from these 2 families.
125
I-LDL turnover studies disclosed a marked reduction in the fractional catabolic rate (0.19±0.01 versus 0.36±0.03 pools per day, respectively;
P
<0.001) and a significant increase in the production rate [20.7±4.4 versus 14.0±2.4 mg · kg
−1
· d
−1
, respectively;
P
<0.01] of LDL apoB in the probands compared with normolipidemic controls. We then studied the in vivo biodistribution and tissue uptake of
99m
technetium-labeled LDL in the probands and compared them with those in normal controls and 1 FH homozygote. The probands showed a significant reduction in hepatic LDL uptake, similar to that observed in the FH homozygote. A reduced uptake of LDL by the kidney and spleen was also observed in all patients. Our findings suggest that this recessive form of hypercholesterolemia is due to a marked reduction of in vivo LDL catabolism. This appears to be caused by a selective reduction in hepatic LDL uptake. We propose that in this new lipid disorder, a recessive defect causes a selective impairment of LDL receptor function in the liver.
Risk of Venous Thromboembolism and Clinical Manifestations in Carriers of Antithrombin, Protein C, Protein S Deficiency, or Activated Protein C Resistance
Authors: Bucciarelli Paolo , Rosendaal Frits R. , Tripodi Armando , Mannucci Pier Mannuccio , De Stefano Valerio , Palareti Gualtiero , Finazzi Guido , Baudo Francesco , Quintavalla Roberto
Published: June 2011

Abstract
—Deficiencies of antithrombin (AT), protein C (PC) or protein S (PS), and activated protein C resistance (APCR) are very well-established coagulation defects predisposing to venous thromboembolism (VTE). We performed a retrospective cohort family study to assess the risk for VTE in individuals with AT, PC, or PS deficiency, or APCR. Five hundred thirteen relatives from 9 Italian centers were selected from 233 families in which the proband had had at least 1 episode of VTE. We calculated the incidence of VTE in the whole cohort and in the subgroups after stratification by age, sex, and defect. The overall incidence of VTE (per 100 patient-years) in the group of relatives was 0.52. It was 1.07 for AT, 0.54 for PC, 0.50 for PS, 0.30 for APCR, and 0.67 in the group with a double defect. The incidence was associated with age, but not with sex. The mean age at onset was between 30 and 40 years for all the coagulation defects. Women had the peak of incidence in the age range of 21 to 40 years, earlier than men. The lifetime risk for VTE was 4.4 for AT versus APCR, 2.6 for AT versus PS, 2.2 for AT versus PC, 1.9 for PC versus APCR, and 1.6 for PS versus APCR. AT deficiency seems to have a higher risk for VTE than the other genetic defects. There is a relation between age and occurrence of thrombosis for both men and women. The latter had the peak of incidence earlier than the former.
NGF Activates Similar Intracellular Signaling Pathways in Vascular Smooth Muscle Cells as PDGF-BB But Elicits Different Biological Responses
Authors: Kraemer Rosemary , Nguyen Hiep , March Keith L. , Hempstead Barbara
Published: June 2011

Abstract
—The signaling pathways that regulate smooth muscle cell migration and proliferation are incompletely understood. Smooth muscle cells express at least 3 families of receptor tyrosine kinases that mediate cell migration: platelet-derived growth factor (PDGF) receptors, the trk family of neurotrophin receptors, and insulin-like growth factor 1 receptor. The neurotrophin, nerve growth factor (NGF), and insulin-like growth factor 1 induce the migration but not the proliferation of smooth muscle cells, whereas PDGF-BB stimulates both responses. To determine whether distinct signaling pathways downstream of receptor tyrosine kinases specifically mediate smooth muscle cell migration or proliferation, the ligand-induced activation of different signaling pathways in smooth muscle cells was examined. NGF induces prolonged activation of the Shc/MAP kinase pathway and phospholipase Cγ compared with PDGF-BB. The activation of phosphatidylinositol-3 kinase, however, was 10-fold greater in response to PDGF-BB compared with NGF. Insulin-like growth factor 1 activates only phosphatidylinositol-3 kinase. Pharmacological inhibitors of phosphatidylinositol-3 kinase, Wortmannin and LY294002, inhibit PDGF-BB and NGF-induced migration, whereas an inhibitor of MAP kinase kinase, PD98059, has no effect. Our results suggest that (1) different receptor tyrosine kinases use similar patterns of activation of signaling pathways to mediate distinct biological outcomes of cell migration and proliferation, (2) NGF activates signaling proteins in smooth muscle cells similar to those activated during NGF-induced neuronal differentiation, and (3) the combinatorial effects of different signaling pathways are important for the regulation of smooth muscle cell migration and proliferation. Further studies using mutant trk receptors will help to define the signal transduction pathways mediating NGF-induced smooth muscle cell migration.
Dose-Dependent Inverse Relationship Between Alcohol Consumption and Serum Lp(a) Levels in Black African Males
Authors: Fontana Pierre , Mooser Vincent , Bovet Pascal , Shamlaye Conrad , Burnand Bernard , Lenain Vincent , Marcovina Santica M. , Riesen Walter , Darioli Roger
Published: June 2011

Abstract
—Serum or plasma levels of Lp(a) vary widely between individuals and are higher in Africans and their descendants compared with white persons. In whites, high serum levels of Lp(a) are associated with the premature development of atherosclerosis. In both ethnic groups, serum Lp(a) levels are highly genetically determined and only a few environmental or physiological factors, like testosterone or estrogen, have been shown to lower serum Lp(a) levels. In whites, alcohol consumption is associated with lower serum Lp(a) levels. However, the mechanism underlying this association and whether it holds true for blacks is not known. To address these questions, we analyzed serum Lp(a) levels in 333 middle-aged males of African descent from the Seychelles Islands (Indian Ocean). In addition, we analyzed the size of the apo(a) isoforms and the serum levels of albumin and sex hormones in a subset of 279 subjects. Serum Lp(a) levels were similar in teetotalers (median, 32.5 mg/dL; n=42) and occasional drinkers (median, 34.1 mg/dL; n=112). In contrast, individuals consuming 10 to 80 g of ethanol/d (n=83) and heavy drinkers (>80 g of ethanol/d, n=96) had a 9% and 32% lower median Lp(a) level than teetotalers, respectively (
P
=0.01). The size distribution of the apo(a) isoforms and the mean serum levels of albumin, estradiol, and luteinizing hormone were similar in teetotalers and occasional drinkers compared with moderate and heavy drinkers. These latter 2 groups had lower serum levels of testosterone and sex hormone–binding globulin. These data indicate that alcohol intake is associated in a dose-dependent manner with lower serum Lp(a) levels in males of African descent and that this association is not related to the size of the apo(a) isoforms, to the synthetic function of the liver, or to sex hormone biochemical status.
Cholesterol and Lipoprotein Metabolism in Aging
Authors: Parini Paolo , Angelin Bo , Rudling Mats
Published: June 2011

Abstract
—Plasma cholesterol levels increase with age, as does the incidence of coronary heart disease. The mechanisms responsible for the age-related hypercholesterolemia are not well understood. An interesting hypothesis suggests that the relative deficiency in growth hormone (GH), which occurs with aging, contributes to the development of the age-related hypercholesterolemia, because GH has beneficial effects on cholesterol metabolism. In the present work, we tested this hypothesis by the administration of GH to normal rats of varying ages. Plasma lipids and hepatic cholesterol metabolism were characterized in 2-, 12-, and 18-month-old male Sprague-Dawley rats. In 2-month-old rats, GH specifically stimulated the hepatic low density lipoprotein (LDL) receptor expression in a dose-dependent way, both at the protein level and at the mRNA level. Concomitantly, plasma cholesterol increased by ≈30% within the large high density lipoprotein and LDL fractions. In 12-month-old animals, cholesterol 7α-hydroxylase (C7αOH) activity was reduced, whereas hepatic LDL receptors and plasma total cholesterol were unchanged. GH treatment (1 mg · kg
−1
· d
−1
) normalized the activity of C7αOH and had effects on plasma cholesterol and LDL receptors similar to those seen in 2-month-old animals. In 18-month-old rats, plasma cholesterol was increased 2-fold, whereas hepatic LDL receptor expression and C7αOH activity were similar to those of the 12-month-old animals. Infusion of GH to 18-month-old rats had similar effects on hepatic C7αOH and LDL receptors as seen in 12-month-old rats. However, GH treatment strongly reduced the hypercholesterolemia in 18-month-old animals. We conclude that the age-dependent increase of plasma cholesterol in rats can be reversed by the administration of GH, presumably through the pleiotropic effects of this hormone on lipoprotein metabolism.
Modified LDL–Mediated Increases in Endothelial Layer Permeability Are Attenuated With 17β-Estradiol
Authors: Gardner Gerrie , Banka Carole L. , Roberts Kim A. , Mullick Adam E. , Rutledge John C.
Published: June 2011

Abstract
—Current research suggests that estrogen may have primary effects on the artery wall. To investigate the mechanisms of female sex hormone actions in the artery wall, we used an isolated, perfused, rat carotid artery model to examine the effects of estradiol on the rates of accumulation of normal (N-LDL) and minimally modified (MM-LDL) low density lipoprotein in ovariectomized rats. N-LDL, MM-LDL, and oxidized LDL (OX-LDL) were fluorescently labeled and perfused into individual arteries. The rate of LDL accumulation was measured by quantitative fluorescence microscopy before and after treatment with estradiol (1 nmol/L, 272 pg/mL). Estradiol had no effect on the rate of N-LDL accumulation (45±12 versus 48±15 ng cholesterol per cm
2
per h). However, estradiol significantly decreased the rate of MM-LDL (240±48 versus 160±48 ng cholesterol per cm
2
per h;
P
<0.05) and OX-LDL (191±53 versus 112±36 ng cholesterol per cm
2
per h;
P
<0.05) accumulation. Further experiments showed that perfusion of unlabeled MM-LDL (100 μg/mL) increased endothelial layer permeability when the rate of accumulation of a water-soluble, fluorescently labeled, reference molecule (64 000–molecular weight dextran) was determined before and after perfusion of MM-LDL (319±96 versus 510±191 ng per cm
2
per h, n=6 arteries;
P
<0.05). Estradiol prevented the expected increase in the rate of dextran accumulation when perfused with MM-LDL (control, 415±49 ng per cm
2
per h and MM-LDL+estradiol, 415±160 ng per cm
2
per h). Our studies show that estradiol prevents compromise of the endothelial barrier mediated by MM-LDL and attenuates accumulation of MM-LDL in the artery wall.
AJvW-2, an Anti-vWF Monoclonal Antibody, Inhibits Enhanced Platelet Aggregation Induced by High Shear Stress in Platelet-Rich Plasma From Patients With Acute Coronary Syndromes
Authors: Eto Koji , Isshiki Takaaki , Yamamoto Hiroshi , Takeshita Satoshi , Ochiai Masahiko , Yokoyama Naoyuki , Yoshimoto Ryota , Ikeda Yasuo , Sato Tomohide
Published: June 2011

Abstract
—The platelet aggregation that is dependent on von Willebrand factor (vWF) is important in the thrombogenesis that occurs under conditions of high shear stress, eg, during acute coronary syndromes (ACSs). A monoclonal antibody, AJvW-2, directed against the A1 domain of human vWF specifically blocks the interaction between plasma vWF and platelet glycoprotein (GP) Ib. To evaluate the association between the vWF-GPIb interaction and the enhanced shear-induced platelet aggregation (SIPA) observed in ACSs, we tested the effect of this antibody on platelet aggregation. Platelet-rich plasma was prepared from the citrated blood of 12 patients with unstable angina (UAP) and 20 patients with acute myocardial infarction (AMI) who were admitted within 3 hours of the onset of cardiac symptoms and from 18 controls. We observed the following: (1) 1.7-fold higher plasma levels of vWF and ristocetin cofactor activity in UAP patients and (2) 2.8-fold higher levels in the AMI group than in controls. Using a cone-and-plate viscometer, we measured the mean value of SIPA under high-shear conditions (108 dyne/cm
2
) and found them to be 1.3-fold higher in the UAP group and 2.0-fold higher in the AMI group than in controls. The high SIPA in all groups was completely inhibited by 10 μg/mL AJvW-2. Under low-shear conditions (12 dyne/cm
2
), platelet aggregation was increased only in the AMI group, but this was unaffected by AJvW-2. We observed a significant correlation in both ACS groups between high SIPA and the plasma vWF level or vWF larger multimers. These findings suggest that the vWF-GPIb interaction is important in coronary occlusion and that inhibition of this interaction (with the use of AJvW-2) may prevent further events in the coronary arteries.
Individual Propensity for Arterial Thrombosis
Authors: Wysokinski Waldemar E. , McBane Robert D. , Owen Whyte G.
Published: June 2011

Abstract
—Arterial thrombophilia independent of vascular pathology has not been previously defined either experimentally or epidemiologically. To address the existence of an individual propensity to arterial thrombosis, we exploited a previously developed procedure entailing traumatic (crush) injury of paired porcine carotid arteries for generating platelet-rich thrombi. Porcine carotid arteries were injured bilaterally by serial hemostat crushes. Thrombus generation was monitored by local accumulation of autologous
111
In-labeled platelets and Doppler blood flow. Within this cohort of animals of similar age and size, the lowest to the highest responders in thrombus mass spanned a 7-fold range, showing no correlation with shear, platelet or leukocyte count, or plasma concentrations of fibrinogen or von Willebrand factor. However, there was strong intra-individual correlation (
r
2
=0.80;
P
<0.001) of thrombus deposition between carotid artery pairs. The wide variation in thrombotic response to a standardized stimulus, not accounted for by shear stress or typical hematological variables, appears to be an intrinsic propensity of the individual. The experimental system for thrombus generation is sufficiently quantitative for assessment of variables determining this propensity.
High-Density Lipoproteins Differentially Modulate Cytokine-Induced Expression of E-Selectin and Cyclooxygenase-2
Authors: Cockerill Gillian W. , Saklatvala Jeremy , Ridley Simon H. , Yarwood Helen , Miller Norman E. , Oral Barbaros , Nithyanathan Saro , Taylor Graham , Haskard Dorian O.
Published: June 2011

Abstract—Atherogenesis is a multifactorial chronic inflammatory disease in which low plasma levels of HDLs are a strong predictor of the condition. Although the mechanism of protection by HDLs is not precisely known, HDLs have been shown to influence many of the events involved in the development of atherosclerosis. Previously we have shown that HDLs inhibited the cytokine-induced expression of adhesion molecules (E-selectin, VCAM-1, and ICAM-1) by endothelial cells (ECs). As the complete transcriptional regulation of all 3 genes requires the NF-κB family of transcription factors, we examined the effect of HDLs on activation of NF-κB. We also investigated the effect of HDLs on 2 other cytokine-induced genes, granulocyte-macrophage colony-stimulating factor (GM-CSF) and cyclooxygenase (Cox-2; prostaglandin H2synthase, EC0.1.14.99.1). E-selectin expression in response to tumor necrosis factor-α (TNFα) was, as expected, inhibited in ECs that had been preincubated with HDLs. However, the level of secretion of GM-CSF in the same cultures was no different from control. In a similar manner, although HDLs had no effect on steady-state mRNA levels of GM-CSF, the levels of E-selectin were significantly inhibited by HDLs. In transient cotransfection experiments we found that HDLs inhibited the cytokine-induced expression of a reporter gene driven by the E-selectin proximal promoter (−383 to 80) but had no effect on the expression of a reporter gene driven under the control of the proximal promoter of GM-CSF (−627 to 28). As would be predicted from this differential response, HDLs did not influence the nuclear translocation or DNA binding of NF-κB, or alter the kinetics of degradation and resynthesis of the inhibitory protein IκBα. We found that HDLs synergized with cytokine to enhance the expression of Cox-2 and induce the synthesis of its main EC product, prostacyclin (PGI2), a potent inhibitor of platelet and leukocyte functions. In conclusion, HDL induces an antiinflammatory phenotype in cytokine-induced ECs, synergizing with cytokine to induce elevation of Cox-2 in addition to inhibiting adhesion molecule expression. Our studies show that these differential effects are mediated in a manner that is likely to be independent of NF-κB per se.
Increased Plasma 7β-Hydroxycholesterol Concentrations in a Population With a High Risk for Cardiovascular Disease
Authors: Ziedén Bo , Kaminskas Arvydas , Kristenson Margareta , Kucinskienê Zita , Vessby Bengt , Olsson Anders G. , Diczfalusy Ulf
Published: June 2011

Abstract
—The mortality in coronary heart disease among 50- to 54-year-old men is 4 times higher in Lithuania than in Sweden. It was recently suggested that traditional risk factors could not explain this mortality difference. LDL of Lithuanian men showed, however, a lower resistance to oxidation than that of Swedish men. In addition, the plasma concentration of γ-tocopherol, lycopene, and β-carotene were lower in Lithuanian men. In the present investigation, we determined plasma oxysterols in men from Lithuania and Sweden and found that the plasma concentration of 7β-hydroxycholesterol was higher in Lithuanian men, 12±5 versus 9±8 (SD) ng/mL (
P
=0.0011). This oxysterol is a cholesterol autoxidation product and there is no indication that it should have an enzymatic origin. Mean LDL oxidation lag time was shorter in Lithuanian men (75±14 versus 90±13 minutes,
P
<0.0001) and the concentration of LDL linoleic acid was lower (249±56 versus 292±54 μg/mg of LDL protein,
P
<0.0001). Lipid corrected γ-tocopherol was 0.07±0.02 mg/mL in Vilnius men and 0.12±0.04 mg/mL (
P
<0.0001) in Linköping men. There was a negative correlation between the concentration of 7β-hydroxycholesterol and lag time (
R
=−0.31,
P
=0.0023). It is suggested that the higher 7β-hydroxycholesterol concentration in Lithuanian men is an indication of an increased in vivo lipid peroxidation.
Retinoic Acid Regulates Arterial Smooth Muscle Cell Proliferation and Phenotypic Features In Vivo and In Vitro Through an RARα-Dependent Signaling Pathway
Authors: Neuville P. , Yan Z.-q , Gidlöf A. , Pepper M. S. , Hansson G. K. , Gabbiani G. , Sirsjö A.
Published: June 2011

Abstract
—We have recently shown that all-
trans
retinoic acid (tRA) modulates arterial smooth muscle cell (SMC) morphologic features and biochemical composition in vitro. It has been proposed that different SMC phenotypes coexist in arteries, which may be retrieved in culture: hence, a differential action of tRA on distinct SMC subsets is conceivable. We have examined the effect of tRA on SMC proliferation, migration, plasminogen activator activity, and α-smooth muscle actin expression in 2 phenotypically different rat SMC populations, cultured respectively from the normal aortic media and from the intimal thickening (IT) after endothelial injury. tRA inhibited proliferation and increased migration and tissue-type plasminogen activator activity in both SMC populations, but decreased α-smooth muscle actin only in SMC cultured from the IT. The action of tRA is mediated by 2 families of nuclear receptors, RAR and RXR, each containing 3 isoforms, α, β, and γ. RAR and RAR-α agonists, but not RXR agonists, inhibited SMC proliferation in both cell populations and α-smooth muscle actin expression only in IT SMC. When administered intraperitoneally to balloon-injured rats, tRA and RAR-α agonists reduced the intimal hyperplasia in the carotid artery. Our results show that tRA and synthetic retinoids can affect the proliferation, migration, and differentiation of SMC in vitro. Furthermore, retinoids are able to reduce the IT induced by endothelial injury in vivo.
The Natural Course of Atherosclerosis
Authors: Kiechl Stefan , Willeit Johann
Published: June 2011

Abstract
—Arterial remodelling is a potentially important component in atherogenesis aimed at delaying the development of significant lumen compromise. Current knowledge on this phenomenon is mainly restricted to experimental evaluations and a few postmortem studies. We used high-resolution duplex ultrasound to study 5-year changes (1990 to 1995) in vessel geometry in a large random sample of the general population (Bruneck Study). Carotid arteries free of atherosclerosis and wall thickening preserved a normal size to high ages. In contrast, common and internal carotid arteries with elevated intima-media thickness (≥50th percentile) experienced marked age-dependent dilation that started already in the 5th decade and continuously accelerated thereafter (structural ageing). Vessel diameters were subject to complex regulation involving morphometric characteristics, sex, wall thickness, hypertension, LDL cholesterol levels, and alcohol consumption. Vascular remodelling secondary to incident or slowly progressive (mural) atherosclerosis included local compensation and a generalised dilation response of vascular segments not primarily affected. Adaptive enlargement at the site of active atherogenesis effectively preserved a near-normal lumen in most instances. The current study identified a second main type of plaque growth, characterized by episodic marked increase in lesion volume probably on the basis of plaque thrombosis. In this setting, we did not observe maximum but insufficient compensation but instead usually observed no compensation at all. Failure of vascular remodelling and marked expansion in plaque size acted synergistically in producing significant lumen compromise. The current prospective survey describes fundamental principles and various facets of arterial remodelling and vascular biology in the general population (in vivo). Vessel geometry was subject to marked temporal changes and showed a correspondingly complex (multifactorial) and dynamic regulation. Vascular remodelling emerged as an important compensatory process in human atherogenesis, which crucially contributed to the determination of lumen obstruction. Efficacy and failure of compensation primarily depended on the type and pathomechanisms of underlying atherogenesis and only in second instance on plaque size and location.
Complete Processing of Type III Collagen in Atherosclerotic Plaques
Authors: Bode Michaela K. , Mosorin Martti , Satta Jari , Risteli Leila , Juvonen Tatu , Risteli Juha
Published: June 2011

Abstract
—The extent of processing of type III collagen is assessed, and the proportions of type I and III collagens are estimated in atherosclerotic plaques obtained from the carotid artery, common femoral artery, and aorta. The fraction of type III collagen that had retained its amino-terminal propeptide (pN-collagen) was 42% in the soluble extract but only 0.0081% in the insoluble residue. Taken together, only 0.011% of the type III collagen in whole plaques was in the form of type III pN-collagen. Together with the small amounts of the free propeptides of type I procollagen, this finding indicates a low rate of collagen turnover. The amounts of solubilized telopeptides of type I and III collagens were measured, after heat denaturation and trypsin digestion of the collagenous helix, by specific immunoassays for the corresponding trypsin-generated antigens. The mean proportion of type III collagen was 61% (95% confidence interval, 58% to 65%) in the carotid and femoral artery plaques and 56% (95% confidence interval, 44% to 68%) in the aortic specimens. The completely processed and cross-linked type III collagen seems to be the major collagen type in atherosclerotic plaques.
Mutations in thelipoprotein lipaseGene Associated With Ischemic Heart Disease in Men
Authors: Wittrup Hans H. , Tybjærg-Hansen Anne , Steffensen Rolf , Deeb Samir S. , Brunzell John D. , Jensen Gorm , Nordestgaard Børge G.
Published: June 2011

Abstract—The aim of this study was to test the hypothesis that the Asp9Asn substitution and theT(−93)→Gmutation in the promoter of thelipoprotein lipasegene affect plasma lipid levels and thereby the risk of ischemic heart disease (IHD). We genotyped 9033 men and women from a general population sample and 940 patients with IHD. The frequency of both theGallele and the Asn9 allele in the general population sample was ≈0.015 for both men and women. These 2 mutations appeared together in 95% of carriers. The average triglyceride-raising effect associated with double heterozygosity for theT(−93)→Gmutation and the Asp9Asn substitution was 0.28 mmol/L (P=0.004) and 0.16 mmol/L (P=0.10) in men and women, respectively. On logistic regression analysis allowing for age, the risk of IHD for double heterozygous men and women was increased 90% (95% confidence interval [CI], 20% to 200%) and 30% (95% CI, −40% to 170%), respectively, compared with noncarriers. When, in addition, other conventional cardiovascular risk factors were allowed for, the risk of IHD for double heterozygous men and women was increased 70% (95% CI, 0% to 190%) and 20% (95% CI, −50% to 180%), respectively. Of the overall risk of IHD in men in the general population, the fraction attributable to double heterozygosity was 3%, similar to the 5% attributable to diabetes mellitus. These results demonstrate that the Asp9Asn substitution is in linkage disequilibrium with theT(−93)→Gmutation and that the double-heterozygous carrier status is associated with elevated plasma triglycerides and an increased risk of IHD in men.
Beneficial Effects of Conversion From Cyclosporine to Azathioprine on Fibrinolysis in Renal Transplant Recipients
Authors: van den Dorpel Marinus A. , Veld Arie J. Man in’t , Levi Marcel , ten Cate Jan Wouter , Weimar Willem
Published: June 2011

Abstract
—Cyclosporin A (CsA) has been implicated as one of the factors contributing to the high cardiovascular morbidity and mortality after renal transplantation. This may be mediated by either a high prevalence of conventional risk factors for atherosclerosis, such as hypertension, hypercholesterolemia, and diabetes mellitus, or by impairment of the fibrinolytic activity evoked by CsA, possibly through interference with prostanoid metabolism. We therefore assessed the impact of conversion of CsA to azathioprine immunosuppressive treatment on parameters of fibrinolytic activity and plasma concentration of the prostanoids prostaglandin E
2
and thromboxane B
2
in 18 stable renal transplant recipients. During CsA, mean arterial pressure and serum creatinine were significantly higher than during azathioprine (116±15 mm Hg versus 106±13 mm Hg,
P
=0.0003; and 147±34 μmol/L versus 127±35 μmol/L,
P
=0.002; mean±SD). On conversion, the plasma tissue plasminogen activator activity increased from 1.2 (1.1 to 1.7; median, 95% CI) to 1.8 (1.6 to 2.0) IU/mL (
P
=0.011), without a significant change of the plasminogen activator antigen concentration. This was associated with a substantial decrease in plasminogen activator inhibitor-1 activity from 10.4 (8.5 to 16.7) to 6.4 (5.6 to 9.2) IU/mL (
P
=0.009). Furthermore, plasma levels of prostaglandin E
2
and thromboxane B
2
markedly decreased (from 9.7 [7.4 to 12.9] to 4.6 [4.3 to 8.1] pg/mL,
P
=0.0006; and from 106.1 [91.7 to 214.2] to 70.2 [50.3 to 85.6] pg/mL,
P
=0.002, respectively). During CsA, but not azathioprine, plasma tissue plasminogen activator antigen and plasminogen activator inhibitor-1 levels correlated significantly with prostaglandin E
2
(
r
=0.53,
P
=0.02; and
r
=0.60,
P
=0.008, respectively), and thromboxane B
2
(
r
=0.75,
P
=0.0001; and
r
=0.77,
P
=0.0001, respectively) levels. In conclusion, CsA induced substantial impairment of fibrinolytic activity, which recovered after conversion to azathioprine. The impaired fibrinolysis observed during CsA treatment may be caused by modulation of eicosanoid production or metabolism in vascular endothelial cells and possibly contributes to the high incidence of cardiovascular disease after kidney transplantation.
Effect of Strenuous, Acute Exercise on α2-Adrenergic Agonist–Potentiated Platelet Activation
Authors: Wang Jong-Shyan , Cheng Lee-Ju
Published: June 2011

Abstract—Vigorous exercise transiently increases the risk of primary cardiac arrest. Strenuous, acute exercise can also increase the release of plasma epinephrine. Previous investigations have indicated that epinephrine can potentiate platelet activation by activating platelet α2-adrenoceptors. This study investigated how strenuous, acute exercise affects α2-adrenergic agonist–potentiated platelet activation by closely examining 15 sedentary men who exercised strenuously on a bicycle ergometer. Before and immediately after exercise, platelet adhesiveness on fibrinogen-coated surfaces, [Ca2+]iin platelets, the number and affinity of α2-adrenergic sites on the platelet surface, and plasma catecholamine levels were determined. The results of this study can be summarized as follows: (1) The affinity of α2-adrenergic receptors on platelets decreases while the maximal binding number significantly increases after strenuous exercise, thereby correlating with the rise in plasma catecholamine levels. (2) Basal, clonidine-treated, ADP-treated, and clonidine plus ADP–treated adhesiveness and [Ca2+]iin platelets increased after strenuous exercise. (3) Strenuous exercise is associated with higher percentages of ADP- and clonidine plus ADP–enhanced platelet adhesiveness and [Ca2+]ithan at rest. (4) The synergistic effects of clonidine on ADP-enhanced platelet adhesiveness and [Ca2+]iafter strenuous exercise are much greater than those at rest. Therefore, we conclude that strenuous, acute exercise enhances platelet activation, possibly by altering the performance of platelet α2-adrenergic receptors, facilitating the ability of ADP-activated fibrinogen receptors, and enhancing fibrinogen binding to platelet fibrinogen receptors.
Coexistence of Oxidized Lipids and α-Tocopherol in All Lipoprotein Density Fractions Isolated From Advanced Human Atherosclerotic Plaques
Authors: Niu Xianwa , Zammit Vivienne , Upston Joanne M. , Dean Roger T. , Stocker Roland
Published: June 2011

Abstract
—After investigation of the contents and redox status of antioxidants and lipids in homogenates of both normal artery and atherosclerotic plaque, we now investigated them in the density fractions (very low, low, high, and protein fractions) of atherosclerotic plaque freshly obtained from carotid endarterectomy. By using the optimum extraction method (homogenization in carbonate buffer) and after density gradient ultracentrifugation, we isolated and characterized density fractions of plaque for apolipoproteins, size and contents of α-tocopherol (α-TOH), unesterified cholesterol, cholesteryl linoleate (Ch18:2), and hydroxides and hydroperoxides of Ch18:2, ie, Ch18:2-O(O)H. The distribution of apolipoproteins was more heterogeneous than that in the corresponding lipoproteins isolated from blood, and the majority of material in all plaque density fractions was present in large particles eluting in the void volume of gel-filtration columns. The content of unesterified cholesterol per unit of protein in low- and high-density fractions was 10-fold that in corresponding plasma lipoproteins. Low- and very-low-density fractions contained most of the lesion lipids and α-TOH. Two to five percent of lesion Ch18:2 was present as Ch18:2-O(O)H and distributed more or less equally among all density fractions, yet the content of α-TOH per unit of Ch18:2 was higher than that in corresponding plasma lipoproteins. These results demonstrate that α-TOH and oxidized lipids coexist in all lesion density fractions, further supporting the notion that large proportions of lipids in lipoproteins of advanced stages of atherosclerosis are oxidized. However, although not ruling it out, our results do not support the suggestion that advanced stages of atherosclerosis are associated with gross deficiencies in the lipoproteins’ vitamin E content.
Association of Polymorphisms at the SR-BI Gene Locus With Plasma Lipid Levels and Body Mass Index in a White Population
Authors: Acton Susan , Osgood Doreen , Donoghue Mary , Corella Dolores , Pocovi Miguel , Cenarro Ana , Mozas Pilar , Keilty John , Squazzo Sharon , Woolf Elizabeth A. , Ordovas Jose M.
Published: June 2011

Abstract
—The scavenger receptor class B type I (SR-BI) is a lipoprotein receptor that has been shown to be important in high density lipoprotein cholesterol (HDL-C) metabolism in mice. To determine its role in humans, we have characterized the human SR-BI gene and investigated its genetic variation in 489 white men and women. Five variants were demonstrated: 2 in introns (3 and 5) and 3 in exons (1, 8, and 11). Three variants at exons 1 and 8 and intron 5 with allele frequencies >0.1 were used to examine associations with lipid or anthropometric variables. The exon 1 variant was significantly (
P
<0.05) associated with increased HDL-C and lower low density lipoprotein cholesterol (LDL-C) values in men, but no associations were observed in women. The exon 8 variant was associated in women with lower LDL-C concentrations (3.05±0.98 mmol/L and 3.00±0.93 mmol/L for heterozygotes and homozygotes, respectively) compared with women homozygous for the common allele (3.39±1.09 mmol/L,
P
=0.043). No associations for this variant were observed in men. Women carriers of the intron 5 variant showed a higher body mass index (23.8±3.8 kg/m
2
,
P
=0.031) than those women homozygous for the common allele (22.4±3.4 kg/m
2
). Similar results were observed after haplotype analysis. Multiple regression analysis using HDL-C, LDL-C, and body mass index as dependent variables and age, sex, and each of the genetic variants as predictors also provided similar results. The associations found with both LDL-C and HDL-C suggest that SR-BI may play a role in the metabolism of both lipoprotein classes in humans.
Large Amounts of Vascular Endothelial Growth Factor at the Site of Hemostatic Plug Formation In Vivo
Authors: Weltermann Ansgar , Wolzt Michael , Petersmann Kai , Czerni Christine , Graselli Ursula , Lechner Klaus , Kyrle Paul A.
Published: June 2011

Abstract
—Vascular endothelial growth factor (VEGF) is important for the proliferation, differentiation, and survival of microvascular endothelial cells. It is a potent angiogenic factor and a specific endothelial cell mitogen that increases fenestration and extravasation of plasma macromolecules. Recently, large quantities of VEGF were detected in human megakaryocytes. Incubation of human platelets with thrombin in vitro resulted in the release of large amounts of VEGF. To investigate whether VEGF is released from platelets during coagulation activation in vivo, we measured in human subjects VEGF at the site of plug formation, ie, in blood emerging from a standardized injury made to determine bleeding time (shed blood). VEGF was also determined in the same volunteers after treatment with the specific thrombin inhibitor recombinant hirudin (r-hirudin). In a double-blind, randomized, crossover study, 17 healthy male volunteers (aged 20 to 35 years) were investigated. VEGF concentrations were measured in venous blood and in shed blood by the use of an immunoassay 10 minutes after intravenous administration of r-hirudin (0.35 mg/kg of body weight) or physiological saline. Prothrombin fragment f1.2 (f1.2) and β-thromboglobulin (β-TG) were determined as indicators of coagulation and platelet activation, respectively. Concentrations of VEGF, f1.2, and β-TG in shed blood 4 minutes after injury were significantly higher than in venous blood (VEGF, 55.8±9.2 versus <20 pg/mL,
P
<0.001; f1.2, 71.3±10.4 versus 0.78±0.03 nmol/L,
P
<0.001; β-TG, 2290±170 versus 53.2±14.0 ng/mL,
P
<0.001). Administration of r-hirudin caused a >50% inhibition of the β-TG and f1.2 levels in shed blood. In a similar manner, much lower amounts of VEGF were detectable at the site of plug formation after r-hirudin treatment (69.0±9.5 versus 37.8±2.6 pg/mL per minute;
P
=0.0015). Our data indicate that substantial quantities of VEGF are released from platelets during the interaction with the injured vessel wall in vivo. This finding may be relevant with respect to wound healing and tissue repair, tumor vascularization, or arterial thrombus formation.
Low Folate Levels and Thermolabile Methylenetetrahydrofolate Reductase as Primary Determinant of Mild Hyperhomocystinemia in Normal and Thromboembolic Subjects
Authors: Gemmati Donato , Previati Maurizio , Serino Maria L. , Moratelli Stefano , Guerra Severino , Capitani Silvano , Forini Elena , Ballerini Giorgio , Scapoli Gian L.
Published: June 2011

Abstract
—Several studies have indicated that mild to moderate hyperhomocystinemia is a common cause of arterial occlusive disease. Whether hyperhomocystinemia per se is an independent risk factor for vein thromboembolism (VTE) is still somewhat controversial. Both genetic and nutritional factors influence plasma homocysteine levels. Therefore, we evaluated plasma total homocysteine (tHcy), folate, and vitamin B
12
levels and established, by polymerase chain reaction, the presence of the C677T mutation (A223V) in the methylenetetrahydrofolate reductase (
MTHFR
) gene in 220 cases with VTE without well-established prothrombotic defects. As a control group, 220 healthy subjects from the same geographic area as the cases were investigated. Hyperhomocystinemia was defined as a plasma tHcy level above the 95th percentile in the controls (18.05 μmol/L). Hyperhomocystinemia was found in 16% of cases (odds ratio=3.59;
P
<0.001); deficiencies of folate (<2.47 ng/mL) or vitamin B
12
(<165 pg/mL), defined as values below the 5th percentile in controls, were found in 17.7% (
P
<0.001) and 12.3% (
P
=0.015) of cases, respectively. The homozygous condition for the
MTHFR
mutation (VV) was present in 28.2% of cases and 17.7% of controls (odds ratio=1.82;
P
=0.013). Comparing only the idiopathic forms of VTE (n=80/220; 36.3%) with normal controls, individuals with hyperhomocystinemia, or individuals homozygous for
MTHFR
mutation increased the odds ratios to 4.03 (
P
=0.005) and 2.11 (
P
=0.018), respectively. No statistically significant difference was observed in the
MTHFR
genotype distribution of cases and controls with hyperhomocystinemia (
P
=0.386); however, the normal
MTHFR
genotype (AA) appeared in control subjects only when tHcy levels were below the 80th percentile (10.57 μmol/L) of the distribution, whereas in case patients, it was present at the highest tHcy levels. A strong association between mutated homozygosity (VV), low folate levels, and hyperhomocystinemia was found in both groups. We conclude that in patients with VTE who do not have coexisting prothrombotic defects, hyperhomocystinemia increases the risk of developing idiopathic and venous thrombosis; the homozygous condition for the
MTHFR
mutation confers a moderate risk but, together with low folate levels, it is the main determinant of mild hyperhomocystinemia in normal and thromboembolic populations.
Endothelin-1 Enhances Plasminogen Activator Inhibitor-1 Production by Human Brain Endothelial Cells via Protein Kinase C-Dependent Pathway
Authors: Zidovetzki Raphael , Wang Jin-Lin , Kim Jeong A. , Chen Peijia , Fisher Mark , Hofman Florence M.
Published: June 2011

Abstract—The effects of endothelin-1 (ET-1) on the production of plasminogen activator inhibitor 1 (PAI-1) and tissue plasminogen activator (t-PA) by human brain-derived endothelial cells in culture were studied. At 100 nmol/L, ET-1 increased PAI-1 production by 88±6% within 72 hours, and increased PAI-1 mRNA expression within 1 hour of stimulation; there was no significant effect on t-PA production. PAI-1 activity was also examined and found to increase with ET-1 treatment. Suboptimal concentrations of ET-1 and tumor necrosis factor-α (TNF-α) acted synergistically to increase PAI-1 production. ET-1 activated protein kinase C and cAMP-dependent protein kinase pathways within 3 to 5 minutes of treatment, with the peak at 10 minutes. Activation of protein kinase C by phorbol-12-myristate-13-acetate (PMA) resulted in increased PAI-1 production, whereas activation of the cAMP-dependent protein kinase by forskolin or dibutyryl cAMP (dBu-cAMP) significantly decreased PAI-1 production. However, simultaneous activation of protein kinase C by PMA and cAMP-dependent protein kinase by dBu-cAMP only slightly attenuated PMA-induced PAI-1 increase. Inhibition of protein kinase C by GF-109213X abolished the effects of ET-1. These results demonstrate that ET-1 and TNF-α function synergistically to induce procoagulant activity of brain endothelial cells in a process that involves a protein kinase C-dependent pathway.
Comparison of the Inhibitory Effects of ApoB100 and Tissue Factor Pathway Inhibitor on Tissue Factor and the Influence of Lipoprotein Oxidation
Authors: Ettelaie Camille , Wilbourn Barry R. , Adam Jacqueline M. , James Nicola J. , Bruckdorfer K. Richard
Published: June 2011

Abstract
—The procoagulant activity of tissue factor is regulated by circulating inhibitors such as tissue factor pathway inhibitor (TFPI) and LDL. These 2 inhibitors also readily associate making the distinction between their activities difficult. We have examined the relative contributions of intact and C-terminal truncated TFPI and ApoB100. By following the inhibitory potential of the preparations, over a period of 120 minutes, it was demonstrated that TFPI and LDL-resembling particles inhibited tissue factor at different rates. TFPI was found to be a short, fast-acting inhibitor, whereas the action of LDL-resembling particles was more prolonged but slower. The oxidation of LDL has been closely associated with the development of cardiovascular disease, including atherosclerosis and thrombosis. Positively charged amino acids, particularly lysine residues, are prone to alterations via the formation of adducts by lipid peroxidation products. These residues are important in the inhibition of tissue factor activity by ApoB100. They also play an important role in the inhibitory Kunitz domains of TFPI. We have shown that the decline in the ability of LDL to inhibit tissue factor was as a result of modifications in LDL arising from oxidation. By examining the effects of oxidation on full-length and C-terminal truncated TFPI bound to LDL-resembling particles, we found that TFPI is only affected when in close association with ApoB100. C-terminal truncated TFPI was not affected significantly by oxidation. Finally, chemical modification of lysine and arginine residues reduced the overall inhibition of tissue factor by TFPI. We propose that TFPI and LDL act separately to inhibit tissue factor in vivo. However, the oxidation of LDL can alter both the endogenous activity of ApoB100 and reduce that of closely associated TFPI, compromising normal hemostasis.
Vascular Endothelial Cells Respond to Spatial Gradients in Fluid Shear Stress by Enhanced Activation of Transcription Factors
Authors: Nagel Tobi , Resnick Nitzan , Dewey C. Forbes , Gimbrone Michael A.
Published: June 2011

Abstract
—The vascular endothelium is exposed to a spectrum of fluid mechanical forces generated by blood flow; some of these, such as fluid shear stress, can directly modulate endothelial gene expression. Previous work by others and in our laboratory, using an in vitro uniform laminar shear stress model, has identified various shear stress response elements (SSREs) within the promoters of certain endothelial genes that regulate their expression by interacting with various transcription factors, including nuclear factor-κB (NF-κB), early growth response-1 (Egr-1), and activator protein-1 (AP-1, composed of c-Jun/c-Jun and c-Jun/c-Fos protein dimers). In the current study, we have examined the topographical patterns of NF-κB, Egr-1, c-Jun, and c-Fos activation in a specially designed in vitro disturbed laminar shear stress model, which incorporates regions of significant spatial shear stress gradients similar to those found in atherosclerosis-prone arterial geometries in vivo (eg, arterial bifurcations, curvatures, ostial openings). Using newly developed quantitative image analysis techniques, we demonstrate that endothelial cells subjected to disturbed laminar shear stress exhibit increased levels of nuclear localized NF-κB, Egr-1, c-Jun, and c-Fos, compared with cells exposed to uniform laminar shear stress or maintained under static conditions. In addition, individual cells display a heterogeneity in responsiveness to disturbed flow, as measured by the amount of NF-κB, Egr-1, c-Jun, and c-Fos in their nuclei. This differential regulation of transcription factor expression by disturbed versus uniform laminar shear stress indicates that regional differences in blood flow patterns in vivo–in particular, the occurrence of spatial shear stress gradients–may represent important local modulators of endothelial gene expression at anatomic sites predisposed for atherosclerotic development.