bluuebunny/crossref_metadata_2025 huggingface dataset:
Different Vasculoprotective Roles of NO Synthase Isoforms in Vascular Lesion Formation in Mice
Authors: Yogo Kenji , Shimokawa Hiroaki , Funakoshi Hajime , Kandabashi Tadashi , Miyata Kenji , Okamoto Shinichiro , Egashira Kensuke , Huang Paul , Akaike Takaaki , Takeshita Akira
Published: June 2011

Abstract
—NO is known to have several important vasculoprotective actions. Although NO is synthesized by 3 different NO synthase (NOS) isoforms, the vasculoprotective action of individual NOS isoforms remains to be clarified. Permanent ligation of the left common carotid artery was performed in control, endothelial NOS (eNOS) knockout (eNOS-KO), and inducible NOS (iNOS) knockout (iNOS-KO) mice. Four weeks after the procedure, neointimal formation and reduction of cross-sectional vascular area (constrictive remodeling) were noted in the left carotid artery. In the eNOS-KO mice, the extent of neointimal formation was significantly larger than in the control or iNOS-KO mice, whereas the extent of vascular remodeling was the highest in the iNOS-KO mice compared with other 2 strains. Antiplatelet therapy with aspirin or antihypertensive treatment with bunazosin failed to inhibit the accelerated neointimal formation in the eNOS-KO mice. These results indicate that eNOS and iNOS have different vasculoprotective actions against the vascular lesion formation caused by blood flow disruption in vivo: NO derived from eNOS inhibits neointimal formation, whereas NO derived from iNOS suppresses the development of constrictive remodeling.
Thrombin Activatable Fibrinolysis Inhibitor and an Antifibrinolytic Pathway
Authors: Bajzar Laszlo
Published: June 2011

Abstract
—Coagulation and fibrinolysis are processes that form and dissolve fibrin, respectively. These processes are exquisitely regulated and protect the organism from excessive blood loss or excessive fibrin deposition. Regulation of these cascades is accomplished by a variety of mechanisms involving cellular responses, flow, and protein-protein interactions. With respect to regulation mediated by protein-protein interaction, the coagulation cascade appears to be more complex than the fibrinolytic cascade because it has more components. Yet each cascade is regulated by initiators, cofactors, feedback reactions, and inhibitors. Coagulation is also controlled by an anticoagulant pathway composed of (minimally) thrombin, thrombomodulin, and protein C.

1

Protein C is converted by the thrombin/thrombomodulin complex to activated protein C (APC), which catalyzes the proteolytic inactivation of the essential cofactors required for thrombin formation, factors Va and VIIIa. An analogous antifibrinolytic pathway has been identified recently. This pathway provides an apparent symmetry between coagulation and fibrinolysis and is also composed of thrombin, thrombomodulin, and a zymogen that is activated to an enzyme. The enzyme proteolytically inactivates a cofactor to attenuate fibrinolysis. However, unlike APC, which is a serine protease, the antifibrinolytic enzyme is a metalloprotease that exhibits carboxypeptidase B–like activity. Within a few years of each other, 5 groups independently described a molecule that accounts for this antifibrinolytic activity. We refer to this molecule as thrombin activatable fibrinolysis inhibitor (TAFI), a name that is based on functional properties by which it was identified, assayed, and purified. (Because of the preferences of some journals “activatable” is occasionally referred to as “activable.”) This review will encompass a historical account of efforts to isolate TAFI and characterize it with respect to its activation, activity, regulation, and potential function in vivo.
Mitogen-Activated Protein Kinases Mediate Matrix Metalloproteinase-9 Expression in Vascular Smooth Muscle Cells
Authors: Cho Aesim , Graves Jonathan , Reidy Michael A.
Published: June 2011

Abstract—Expression of matrix metalloproteinase (MMP)-9 has been linked to the progression of plaque rupture and intimal formation in arterial lesions. In this study, we determined which factors and signaling pathways are involved in regulating the MMP-9 gene. Rat carotid arterial smooth muscle cells treated with tumor necrosis factor (TNF)-α showed a marked increase in MMP-9 activity and mRNA level, whereas platelet-derived growth factor (PDGF) showed a slight induction of the MMP-9 mRNA level. TNF-α treatment caused an increase in c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38 MAPK), and extracellular signal–regulated kinase (ERK) activities, whereas PDGF treatment caused an increase in ERKs and p38 MAPK activities without any effect on JNK activity. Treatment with either SB203580 (inhibitor of p38 MAPK) or U0126 (inhibitor of the ERK pathway) downregulated the TNF-α–induced MMP-9 expression in a dose-dependent manner. Treatment of cells with TNF-α and PDGF together stimulated the MMP-9 expression at a level higher than that observed with either factor alone, suggesting that TNF-α and PDGF have a synergistic effect on MMP-9 expression in arterial smooth muscle cells. Furthermore, suboptimal inhibitory concentrations of SB203580 and U0126 together almost completely inhibited the MMP-9 expression. These results suggest that p38 MAPK and ERK pathways contribute to the transcriptional regulation of MMP-9 in arterial smooth muscle cells.
In Vivo Activation of Rat Aortic Platelet-Derived Growth Factor and Epidermal Growth Factor Receptors by Angiotensin II and Hypertension
Authors: Kim Shokei , Zhan Yumei , Izumi Yasukatsu , Yasumoto Hideo , Yano Masahiko , Iwao Hiroshi
Published: June 2011

Abstract
—It is unclear whether the previous in vitro evidence of a link between angiotensin II (Ang II) and growth factor receptors can apply to the in vivo situation. In this study, we examined vascular platelet-derived growth factor (PDGF) and epidermal growth factor (EGF) receptor activation in stroke-prone spontaneously hypertensive rats (SHRSP) and the role of Ang II. Tyrosyl phosphorylation of the growth factor receptors was determined by Western blot analysis coupled with immunoprecipitation. Tyrosyl phosphorylation of the aortic PDGF β-receptor, but not the EGF receptor, was chronically increased in SHRSP with hypertension, compared with normotensive rats, being accompanied by increased extracellular signal–regulated kinase (ERK) activity. Treatment of SHRSP with ACE inhibitors (perindopril or enalapril) significantly reduced aortic PDGF β-receptor tyrosyl phosphorylation and ERK activity, whereas treatment with hydralazine failed to reduce these activities. Therefore, these aortic changes in SHRSP were mediated by Ang II in response to vascular ACE. Ang II was infused into rats to examine the effects on aortic growth factor receptors. Chronic Ang II infusion, via the angiotensin type 1 receptor, significantly increased activation of the aortic PDGF β-receptor but not the EGF receptor. Thus, the aortic PDGF β-receptor, activated by ACE-mediated Ang II, seems to be responsible for vascular remodeling in hypertensive rats.
Platelet-Leukocyte Cross Talk in Whole Blood
Authors: Li Nailin , Hu Hu , Lindqvist Malin , Wikström-Jonsson Eva , Goodall Alison H. , Hjemdahl Paul
Published: June 2011

Abstract
—Thrombosis and inflammation involve complex platelet-leukocyte interaction, the details of which are not fully elucidated. Therefore, we investigated cross talk between platelets and leukocytes in whole blood, under the following physiological conditions: at 37°C, with normal calcium concentrations, and with shear force. Platelet P-selectin and leukocyte CD11b expression were used to monitor platelet and leukocyte activation, respectively, and platelet-leukocyte aggregation (PLA) was analyzed. The leukocyte-specific agonist
N
-formyl-methionyl-leucyl-phenylalanine (10
−6
mol/L) increased P-selectin–positive platelets from 2.5±0.1% to 5.1±0.6% (
P
<0.05). The increase was inhibited by either the platelet-activating factor (PAF) antagonist SR27417, the superoxide anion scavenger superoxide dismutase, the 5-lipoxygenase inhibitor Zileuton, or the 5-lipoxygenase–activating protein inhibitor MK-886, suggesting the involvement of PAF, superoxide anion, and 5-lipoxygenase products in leukocyte-induced platelet activation. The platelet-specific agonist collagen (1 μg/mL) increased leukocyte CD11b expression from 2.94±0.52 to 3.81±0.58 (
P
<0.05); this was not inhibited by the thromboxane A
2
receptor antagonist ICI 192.605 or the PAF antagonist SR27417. Platelet P-selectin expression induced by
N
-formyl-methionyl-leucyl-phenylalanine and leukocyte CD11b expression induced by collagen could be suppressed by glycoprotein IIb/IIIa blockade or P-selectin blockade. This study documents platelet-leukocyte cross talk under conditions that mimic a physiological state and suggests that this involves multiple mediators and mechanisms. Furthermore, new evidence of integrin and selectin involvement in intracellular and intercellular signaling during platelet-leukocyte cross talk is provided.
Macrophage Scavenger Receptor Class A
Authors: de Winther Menno P. J. , van Dijk Ko Willems , Havekes Louis M. , Hofker Marten H.
Published: June 2011

Abstract
—In atherogenesis, elevated plasma levels of low density lipoprotein (LDL) lead to the chronic presence of LDL in the arterial wall. There, LDL is modified (eg, oxidized), and these modified lipoproteins activate endothelial cells, which attract circulating monocytes. These monocytes enter the vessel wall, differentiate into macrophages, and subject the modified lipoproteins to endocytosis through scavenger receptor pathways. This unrestricted uptake, which is not limited by intracellular cholesterol levels, eventually leads to the formation of lipid-filled foam cells, the initial step in atherosclerosis. Macrophage scavenger receptor class A (SRA) is thought to be one of the main receptors involved in foam cell formation, mediating the influx of lipids into the macrophages. In addition to this role in modified lipoprotein uptake by macrophages, the SRA has been shown to be important in the inflammatory response in host defense, cellular activation, adhesion, and cell-cell interaction. Given the importance of these processes in atherogenesis, these latter functions may prove to make the SRA a multifunctional player in the atherosclerotic process.
Lesions in Ryanodine Channels in Smooth Muscle Cells Exposed to Oxidized Low Density Lipoprotein
Authors: Massaeli Hamid , Austria J. Alejandro , Pierce Grant N.
Published: June 2011

Abstract
—The purpose of the present investigation was to investigate the subcellular basis responsible for the loss of vasoreactivity in atherosclerotic vessels. We have chosen to focus on the potential of oxidized low density lipoprotein (oxLDL), an important atherogenic agent, to alter sarcoplasmic reticulum (SR) structure and function. Vascular smooth muscle cells (VSMCs) were exposed for 1 to 6 days to low concentrations of minimally oxidized LDL. ATP was used to probe SR function in VSMCs. ATP can increase [Ca
2+
]
i
in control VSMCs because of a release of Ca
2+
from the SR. However, after chronic exposure to oxLDL, cells lose their ability to increase [Ca
2+
]
i
in response to ATP. These cells also exhibit a depressed rise in [Ca
2+
]
i
after exposure to ryanodine. These effects were associated with a decreased immunoreactivity for the ryanodine-sensitive Ca
2+
-release channels in the SR of oxLDL-treated cells. Immunohistochemical analysis of aortic sections obtained from rabbits fed a cholesterol-supplemented diet revealed a significant decrease in the immunoreactivity for ryanodine channels in the plaque and in the medial layer underlying the plaque. In summary, our data identify oxLDL as a component within the atherosclerotic milieu capable of inducing a decrease in smooth muscle ryanodine channel density. This alteration is associated with a significant defect in the ability of the SR within the smooth muscle cell to regulate Ca
2+
. These lesions may contribute to the altered vasoreactivity exhibited by atherosclerotic vessels.
Mouse Model of Femoral Artery Denudation Injury Associated With the Rapid Accumulation of Adhesion Molecules on the Luminal Surface and Recruitment of Neutrophils
Authors: Roque Merce , Fallon John T. , Badimon Juan J. , Zhang Wen X. , Taubman Mark B. , Reis Ernane D.
Published: June 2011

Abstract
—Techniques of arterial injury commonly used in animals to mimic endovascular procedures are not suitable for small mouse arteries. This has limited examination of the response to arterial injury in genetically modified mice. We therefore sought to develop a model of transluminal injury to the mouse femoral artery that would be reproducible and result in substantial levels of intimal hyperplasia. Mice of the C57BL/6 strain underwent bilateral femoral artery denudation by passage of an angioplasty guidewire. Intimal hyperplasia was observed in 10% of injured arteries at 1 week, in 88% at 2 weeks, and in 90% at 4 weeks. The mean intimal-to-medial area ratio reached 1.1±0.1 at 4 weeks. No intimal proliferation was found in control sham-operated arteries. One hour after injury, the denuded surface was covered with platelets and leukocytes, predominantly neutrophils. This was associated with the accumulation of P-selectin, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1. Expression of these adhesion molecules was not seen in the underlying medial smooth muscle cells. At 24 hours, few neutrophils remained on the denuded surface. At 1 week, macrophages and platelets were present in the vessel wall, partially covered by regenerated endothelium. Transluminal wire injury to the mouse femoral artery induces abundant intimal hyperplasia formation by 2 and 4 weeks and elicits the rapid accumulation of leukocytes and adhesion molecules on the denuded luminal surface. This model will be a valuable tool to study arterial injury in genetically modified mouse models.
Insulin-Like Growth Factor Binding Protein-4 Expression Is Decreased by Angiotensin II and Thrombin in Rat Aortic Vascular Smooth Muscle Cells
Authors: Anwar A. , Zahid A. A. , Phillips L. , Delafontaine P.
Published: June 2011

Abstract
—Insulin-like growth factor-I (IGF-I) is a ubiquitous peptide that regulates cellular growth and differentiation and is involved in vascular proliferative responses. The effects of IGF-I are modulated by several IGF-I binding proteins (IGFBPs), including IGFBP-4, the main IGFBP produced by vascular smooth muscle cells (VSMCs). We have previously shown that angiotensin II (Ang II)–induced and thrombin-induced mitogenesis in VSMCs is dependent on autocrine IGF-I. In addition, we have demonstrated that IGF-I and IGFBP-4 mRNA levels are upregulated in the hypertensive aorta of abdominally coarcted rats, a high-renin hypertension model. To obtain further insight into the IGF-I system and to specifically study changes in IGFBP-4, a known inhibitor of IGF-I action, VSMCs were incubated with Ang II or thrombin. Compared with control, Ang II induced an 87±2% downregulation of IGFBP-4 mRNA levels at 24 hours, with a 61±6% decrease of IGFBP-4 levels, as determined by Western ligand blot analysis. Thrombin had the same depressor effects (87±2% for the mRNA levels and 61±3% for the protein levels). Ang II and thrombin coincubation with
125
I-IGFBP-4 in the conditioned media failed to reveal any increase in fragmentation, indicating that proteolytic cleavage of IGFBP-4 was not involved in the observed effects. Exogenous recombinant human IGFBP-4 decreased thrombin-induced DNA synthesis of human aortic VSMCs by 64%, whereas anti–IGFBP-4 antibody potentiated thrombin-induced DNA synthesis. These data suggest that downregulation of IGFBP-4 expression in VSMCs may play a critical role in vascular growth response to Ang II and thrombin in normal and diseased states, by increasing the bioavailability of IGF-I for its cell-surface receptor.
Convergence of Redox-Sensitive and Mitogen-Activated Protein Kinase Signaling Pathways in Tumor Necrosis Factor-α–Mediated Monocyte Chemoattractant Protein-1 Induction in Vascular Smooth Muscle Cells
Authors: De Keulenaer Gilles W. , Ushio-Fukai Masuko , Yin QiQin , Chung Andrew B. , Lyons P. Reid , Ishizaka Nobukazu , Rengarajan Kalpana , Taylor W. Robert , Alexander R. Wayne , Griendling Kathy K.
Published: June 2011

Abstract
—Monocyte chemoattractant protein-1 (MCP-1) is an important component of the inflammatory response of the vessel wall and has been shown to be regulated by cytokines, such as tumor necrosis factor-α (TNF-α). However, the precise signaling pathways leading to MCP-1 induction have not been fully elucidated in vascular smooth muscle cells (VSMCs). Cytokine signal transduction involves protein kinases as well as reactive oxygen species (ROS). The relation between these 2 factors is not clear. In this study, we show that TNF-α induces a parallel phosphorylation of extracellular signal–regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (p38MAPK) and increases MCP-1 mRNA expression in cultured VSMCs. Inhibition of ERK1/2 but not p38MAPK caused a partial attenuation of MCP-1 induction (43±10% inhibition). Incubation of VSMCs with multiple antioxidants (diphenylene iodonium, liposomal superoxide dismutase, catalase,
n
-acetylcysteine, dimethylthiourea, and pyrrolidine dithiocarbamate) had no effect on TNF-α–mediated MCP-1 upregulation. However, simultaneous blockade of the ERK1/2 and ROS pathways by using PD098059 combined with diphenylene iodonium or
n
-acetylcysteine potently enhanced the ability of MAPK kinase inhibitors to abrogate MCP-1 mRNA expression (100±2% inhibition). Thus, parallel ROS-dependent and ERK1/2-dependent pathways converge to regulate TNF-α–induced MCP-1 gene expression in VSMCs. These data unmask a complex but organized integration of ROS and protein kinases that mediates cytokine-induced vascular inflammatory gene expression.
In Vivo Evidence of the Importance of Cardiac Angiotensin-Converting Enzyme in the Pathogenesis of Cardiac Hypertrophy
Authors: Higaki Jitsuo , Aoki Motokuni , Morishita Ryuichi , Kida Iwao , Taniyama Yoshiaki , Tomita Naruya , Yamamoto Kei , Moriguchi Atsushi , Kaneda Yasufumi , Ogihara Toshio
Published: June 2011

Abstract
—Cardiac angiotensin-converting enzyme (ACE) may play an important role in regulating cardiac hypertrophy. Angiotensin II (Ang II) stimulates cardiac hypertrophy as well as the production of extracellular matrix. However, it is still unclear whether Ang II exerts a direct effect on cardiac hypertrophy independent of its effect on blood pressure or the circulating renin-angiotensin system. Although ACE inhibitors and/or Ang II receptor antagonists have regressed cardiac hypertrophy, classic pharmacological experiments cannot exclude the contribution of hemodynamics and the circulating renin-angiotensin system. In vivo gene transfer provides the opportunity of assessing the effects of increased cardiac angiotensin in the intact animal without circulating angiotensin or blood pressure. Therefore, we used a “gain of function” approach to obtain local overexpression of cardiac ACE. Transfection of the human ACE vector into rat myocardium resulted in a significant increase in cardiac ACE activity (
P
<0.01). More interestingly, morphometry at 2 weeks after transfection revealed a significant increase in the thickness and areas of cardiac myocytes in hearts transfected with the ACE vector (
P
<0.01). In addition, transfection of the ACE vector also resulted in a significant increase in collagen content (
P
<0.01). This increase in cardiac hypertrophy was abolished by the administration of perindopril. Local transfection of the ACE vector into the heart did not result in systemic effects such as increased blood pressure, heart rate, or serum ACE activity. In summary, we have demonstrated that increased autocrine/paracrine angiotensin can directly cause cardiac hypertrophy independent of systemic factors and hemodynamic effects. This approach has important potentials for defining the role of autocrine/paracrine substances in cardiovascular disease.
Interaction of Anti-Phospholipid Antibodies With Late Endosomes of Human Endothelial Cells
Authors: Rochemonteix Béatrix Galve-de , Kobayashi Toshihide , Rosnoblet Corinne , Lindsay Margaret , Parton Robert G. , Reber Guido , de Maistre Emmanuel , Wahl Denis , Kruithof Egbert K. O. , Gruenberg Jean , de Moerloose Philippe
Published: June 2011

Abstract
—Anti-phospholipid antibodies (APLAs) are associated with thrombosis and/or recurrent pregnancy loss. APLAs bind to anionic phospholipids directly or indirectly via a cofactor such as β
2
-glycoprotein 1 (β
2
GPI). The lipid target of APLA is not yet established. Recently, we observed that APLAs in vitro can bind lysobisphosphatidic acid (LBPA). The internal membranes of late endosomes are enriched in this phospholipid. The current study was undertaken to determine to what extent binding of APLA to LBPA is correlated with binding to cardiolipin and to β
2
GPI and to determine whether patient antibodies interact with late endosomes of human umbilical vein endothelial cells (HUVECs) and thus modify the intracellular trafficking of proteins. Binding of patient immunoglobulin G (n=37) to LBPA was correlated significantly with binding to cardiolipin. Although LBPA binding was correlated to a lesser extent with β
2
GPI binding, we observed that β
2
GPI binds with high affinity to LBPA. Immunofluorescence studies showed that late endosomes of HUVECs contain LBPA. Patient but not control antibodies recognized late endosomes, but not cardiolipin-rich mitochondria, even when we used antibodies that were immunopurified on cardiolipin. Incubation of HUVECs with patient plasma samples immunoreactive toward LBPA resulted in an accumulation of the antibodies in late endosomes and led to a redistribution of the insulinlike growth factor 2/mannose-6-phosphate receptor from the Golgi apparatus to late endosomes. Our results suggest that LBPA is an important lipid target of APLA in HUVECs. These antibodies are internalized by the cells and accumulate in late endosomes. By modifying the intracellular trafficking of proteins, APLA could contribute to several of the proposed pathogenic mechanisms leading to the antiphospholipid syndrome.
Effect of Individual Plasma Lipoprotein(a) Variations In Vivo on Its Competition With Plasminogen for Fibrin and Cell Binding
Authors: Soulat Thierry , Loyau Stéphane , Baudouin Véronique , Maisonneuve Lydia , Hurtaud-Roux Marie-France , Schlegel Nicole , Loirat Chantal , Anglés-Cano Eduardo
Published: June 2011

Abstract
—Simultaneous natural changes in lipoprotein(a) [Lp(a)] and plasminogen occur in the nephrotic syndrome and offer a unique opportunity to investigate their effects on plasminogen activation under conditions fashioned in vivo. Plasminogen, Lp(a), and apolipoprotein(a) in plasma were characterized, and their competitive binding to carboxy-terminal lysine residues of fibrin and cell membrane proteins was determined in nephrotic children during a flare-up of the disease (61 cases) and after 6 weeks (33 cases) and 6 months (42 cases) of remission. Low plasminogen concentrations (median 1.34 μmol/L, range 0.39 to 1.96 μmol/L) and high Lp(a) levels (median 0.27 g/L, range 0.07 to 2.57 g/L) were detected at flare-up. These changes were associated with an increased Lp(a) binding ratio onto fibrin (3.13±0.48) and cells (1.53±0.24) compared with binding ratios of control children (1.31±0.19 and 1.05±0.07, respectively) with normal plasminogen and low Lp(a) (median 0.071 g/L). After 6 weeks and 6 months of remission, the values for net decrease in Lp(a) binding to fibrin were 1.7±0.22 (after 6 weeks) and 1.88±0.38 (after 6 months) and were correlated with low Lp(a) concentrations (median 0.2 g/L, range 0.07 to 0.8 g/L; and median 0.12 g/L, range 0.07 to 1.34 g/L) and inversely associated with increased plasminogen levels (median 1.82 μmol/L, range 1.4 to 2.1 μmol/L; and median 1.58 μmol/L, range 1.1 to 2.1 μmol/L). These studies provide the first quantitative evidence that binding of Lp(a) to lysine residues of fibrin and cell surfaces is directly related to circulating levels of both plasminogen and Lp(a) and that these glycoproteins may interact as competitive ligands for these biological surfaces in vivo. This mechanism may be of relevance to the atherothrombotic role of Lp(a), particularly in nephrotic patients.
Antibodies Against Oxidized LDL and Carotid Artery Intima-Media Thickness in a Healthy Population
Authors: Fukumoto Mariko , Shoji Tetsuo , Emoto Masanori , Kawagishi Takahiko , Okuno Yasuhisa , Nishizawa Yoshiki
Published: June 2011

Abstract
—Oxidation of LDLs plays an important role in atherosclerosis, and immune response to oxidized LDL (oxLDL) may modulate atherogenesis. Although immunization with oxLDL is shown to suppress atherogenesis in animal models, the role of the immune response to oxLDL is not well established in humans. We investigated the relationship between the titer of anti-oxLDL antibody (oxLDL Ab) and arterial wall thickness in a healthy population with no clinical signs of atherosclerosis. Intima-media thickness of the carotid arteries (CA-IMT) was measured by high-resolution B-mode ultrasonography in 446 healthy subjects. The titer of IgG-class oxLDL Ab was measured by a solid-phase ELISA. In univariate analysis, CA-IMT correlated positively with age, systolic blood pressure, total cholesterol, triglyceride, LDL cholesterol, body mass index, and waist-to-hip ratio, whereas it correlated negatively with HDL cholesterol and oxLDL Ab titer. The inverse association between oxLDL Ab titer and CA-IMT remained significant in multiple regression analysis, which took other confounding variables into account. These results indicate an independent inverse relationship between oxLDL Ab titer and CA-IMT in healthy subjects, supporting the hypothesis that immune response to oxLDL may have a protective role at an early stage of human atherosclerosis.
Identification of Soluble Forms of Lectin-Like Oxidized LDL Receptor-1
Authors: Murase Takatoshi , Kume Noriaki , Kataoka Hiroharu , Minami Manabu , Sawamura Tatsuya , Masaki Tomoh , Kita Toru
Published: June 2011

Abstract
—Lectin-like oxidized LDL receptor-1 (LOX-1) is a type II membrane protein belonging to the C-type lectin family molecules, which can act as a cell-surface endocytosis receptor for atherogenic oxidized LDL. In this study, we show that soluble forms of LOX-1 are present in conditioned media of cultured bovine aortic endothelial cells (BAECs) and CHO-K1 cells stably transfected with LOX-1 cDNA. Immunoblot analysis of conditioned media from TNF-α–activated BAECs and CHO-K1 cells stably expressing LOX-1 revealed that soluble LOX-1 has an approximate molecular mass of 35 kDa. In TNF-α–activated BAECs, cell-surface expression of LOX-1 precedes soluble LOX-1 production. Cell-surface biotinylation followed by immunoprecipitation and immunoblotting showed that soluble LOX-1 in cell-conditioned media is derived from LOX-1 expressed on the cell surface. Production of soluble LOX-1 was inhibited by PMSF, suggesting that PMSF-sensitive proteases may be involved in this process. Purification of soluble LOX-1 by high-performance liquid chromatography and N-terminal amino acid sequencing of soluble LOX-1 identified the 2 cleavage sites between Arg
86
-Ser
87
and Lys
89
-Ser
90
, which were located in the membrane proximal extracellular domain of LOX-1. The data demonstrate that cell-surface LOX-1 can be cleaved at 2 different sites and transformed into soluble forms. Further studies may explore therapeutic and diagnostic applications of soluble LOX-1 in atherosclerotic diseases.
Hypercholesterolemia Increases Coronary Endothelial Dysfunction, Lipid Content, and Accelerated Atherosclerosis After Heart Transplantation
Authors: Perrault Louis P. , Mahlberg Florence , Breugnot Christine , Bidouard Jean-Pierre , Villeneuve Nicole , Vilaine Jean-Paul , Vanhoutte Paul M.
Published: June 2011

Abstract
—Hyperlipidemia may increase endothelial damage and promote accelerated atherogenesis in graft coronary vasculopathy. To study the effects of hypercholesterolemia on coronary endothelial dysfunction, intimal hyperplasia, and lipid content, a porcine model of heterotopic heart transplantation, allowing nonacute rejection without immunosuppressive drugs, was used. A high cholesterol diet was fed to donor and recipient swine 1 month before and after transplantation. The endothelial function of coronary arteries of native and transplanted hearts from cholesterol-fed animals was studied in organ chambers 30 days after implantation and compared with endothelial function in arteries from animals fed a normal diet. The total serum cholesterol increased 3-fold in donors and recipients. Endothelium-dependent relaxations to serotonin, to the α
2
-adrenergic agonist UK14,304, and to the direct G-protein activator sodium fluoride were decreased significantly in allografted hearts compared with native hearts from both groups. Relaxations to the calcium ionophore A23187 and bradykinin were decreased significantly in allografts from animals fed the high cholesterol diet. The prevalence of intimal hyperplasia was significantly increased in coronary arteries from hypercholesterolemic swine. There was a significant increase in the lipid content of allograft arteries of hypercholesterolemic recipients. Hypercholesterolemia causes a general coronary endothelial dysfunction, increases the prevalence of intimal hyperplasia, and augments the incorporation of lipids in the vascular wall after heart transplantation. Hyperlipidemia accelerates graft coronary atherosclerosis through its effects on the endothelium.
Expression of Secretory Group IIA Phospholipase A
2
in Relation to the Presence of Microbial Agents, Macrophage Infiltrates, and Transcripts of Proinflammatory Cytokines in Human Aortic Tissues
Authors: Menschikowski Mario , Rosner-Schiering Andrea , Eckey Rolf , Mueller Erich , Koch Rainer , Jaross Werner
Published: June 2011

Abstract
—Recent seroepidemiological and immunohistochemical studies have demonstrated an association between microbial infections and atherosclerosis. However, the mechanisms underlying this association are widely unknown. In the present study, arterial specimens obtained at autopsy after sudden death were analyzed concerning (1) the presence of
Chlamydia pneumoniae
, cytomegalovirus, herpes simplex virus, and
Helicobacter pylori
; (2) the expression of secretory group IIA phospholipase A
2
(sPLA
2
-IIA) and of proinflammatory cytokines; and (3) the stage of atherosclerosis. Genomic DNA of microbial pathogens was determined by the polymerase chain reaction technique. The expression of sPLA
2
-IIA was studied immunohistochemically by using monoclonal antibodies against human sPLA
2
-IIA. Transcripts specific for sPLA
2
-IIA, interleukin-1β, tumor necrosis factor-α, and interferon-γ were identified by reverse transcription–polymerase chain reaction. In 18 of 102 analyzed specimens, DNA of microbial pathogens was found. Thirteen sections were positive for
C pneumoniae
, whereas 2 specimens were positive either for cytomegalovirus or for herpes simplex virus. One section contained genomic DNA of all 3 pathogens simultaneously. None of the analyzed tissues exhibited nucleic acids specific for
H pylori
. In addition to macrophage infiltrates, the presence of microbial DNA was closely associated with the occurrence of transcripts specific for proinflammatory cytokines and sPLA
2
-IIA. Pathogens as well as sPLA
2
-IIA and cytokines were found to be present not only in advanced but also in early stages of atherosclerosis. In tissues negative for sPLA
2
-IIA and cytokine expression, none of the pathogens could be identified. Because macrophages exposed to phospholipase A
2
–treated lipoproteins are transformed into foam cells in vitro, the results of this study suggest an alternative mechanism by which microbial infections may act in a proatherogenic fashion in vessel walls.
Dehydroepiandrosterone Retards Atherosclerosis Formation Through Its Conversion to Estrogen
Authors: Hayashi Toshio , Esaki Teiji , Muto Emiko , Kano Hatsuyo , Asai Yukako , Thakur Navin Kumar , Sumi Daigo , Jayachandran Muthuvel , Iguchi Akihisa
Published: June 2011

Abstract
—Dehydroepiandrosterone (DHEA) is speculated to have an antiatherosclerotic effect, although the mechanism of action remains unclear. The objective of the current study was to determine whether the antiatherosclerotic effect of DHEA is related to its conversion to estrogen and to define the role of nitric oxide (NO) in the antiatherosclerotic effect of DHEA. Forty-eight oophorectomized rabbits were divided into 5 groups and fed the following diets for 10 weeks: group 1, a regular rabbit diet plus 1% cholesterol (a high-cholesterol diet [HCD]); group 2, an HCD plus 0.3% DHEA; group 3, an HCD plus 0.3% DHEA and fadrozole (2.0 mg · kg
−1
· d
−1
), a specific aromatase inhibitor; group 4, an HCD plus 17β-estradiol (20 μg · kg
−1
· d
−1
); and group 5, a regular diet. Atherosclerotic lesions, lipid deposition in aortic vessels, and basal and stimulated NO release were measured in the aforementioned groups of rabbits. NO release was measured by using an NO-selective electrode as well as by measuring vascular responses and the plasma NO metabolites nitrite and nitrate. The plasma total cholesterol level was increased, but there were no significant differences in lipid profile in the 4 groups of rabbits that were fed the HCD. The area occupied by atherosclerosis in the thoracic aorta was diminished by ≈60% in the DHEA-treated rabbits (group 2) compared with the HCD group of rabbits (group 1); there was a corresponding 80% decrease in the estradiol group (group 4) but only a 30% decrease in the DHEA plus fadrozole group (group 3). In the aortas of rabbits from groups 1 and 3, the acetylcholine-induced and tone-related basal NO-mediated relaxations were diminished compared with those of the controls (group 5). However, these relaxations were restored in the aortas of group 2 and 4 rabbits, and an increase in NO release was observed in groups 2 and 4 compared with groups 1 and 3, as measured by an NO-selective electrode. Injection of neither solvent (20% ethanol/distilled water) nor fadrozole significantly affected the atherosclerotic area or the NO-related responses described above. We conclude that ≈50% of the total antiatherosclerotic effect of DHEA was achieved through the conversion of DHEA to estrogen. NO may also play a role in the antiatherosclerotic effect of DHEA and 17β-estradiol.
Segregation Analysis of Apolipoproteins A-1 and B-100 Measured Before and After an Exercise Training Program
Authors: An Ping , Rice Treva , Gagnon Jacques , Borecki Ingrid B. , Bergeron Jean , Després Jean-Pierre , Leon Arthur S. , Skinner James S. , Wilmore Jack H. , Bouchard Claude , Rao D. C.
Published: June 2011

Abstract
—Complex segregation analyses of apolipoproteins (apo) A-1 and B-100 were performed in a sample of 520 individuals from 99 white families who participated in the HERITAGE Family Study. In these sedentary families, plasma apo A-1 and B-100 concentrations were measured before and after a 20-week endurance exercise training program. Baseline apo A-1 and B-100 were adjusted for the effects of age (age-adjusted baseline apo A-1 and B-100) and for the effects of age and BMI (age-BMI–adjusted baseline apo A-1 and B-100). The change in response to training was computed as a simple Δ (posttraining minus baseline) and was adjusted for age and the baseline (age-baseline–adjusted apo A-1 and B-100 responses to training). In the present study, a major gene could not be inferred for baseline apo A-1. Rather, we found a major effect along with a multifactorial effect accounting for 8% to 9% and 51% to 56% of the variance, respectively. In addition, no clear evidence supported a major-gene effect for its response to training, whereas the transmission of a major effect from parents to offspring was ambiguous, ie, genetic in nature or familial environmental in origin. The major effect accounted for 15% of the variance, with an additional 21% and 58% of the variance being accounted for by a multifactorial effect in parents and offspring, respectively. It is interesting to have obtained evidence of a putative recessive major locus for baseline apo B-100, which accounted for 50% to 56% of the variance, with an additional 25% to 29% of the variance due to a multifactorial effect. In contrast, no major effect for its response to training was identified, although a multifactorial effect was found that accounted for 27% of the variance. The novel findings arising from the present study are summarized as follows. Baseline apo A-1 and its response to training were influenced by a major effect and a multifactorial effect. Baseline apo B-100 was influenced by a putative major recessive gene with a multifactorial component, but its response to training was influenced solely by a multifactorial component in these sedentary families.
Polymorphisms in the 5′ Regulatory Region of the Tissue Factor Gene and the Risk of Myocardial Infarction and Venous Thromboembolism
Authors: Arnaud Emmanuel , Barbalat Véronique , Nicaud Viviane , Cambien François , Evans Alun , Morrison Caroline , Arveiler Dominique , Luc Gérald , Ruidavets Jean-Bernard , Emmerich Joseph , Fiessinger Jean-Noël , Aiach Martine
Published: June 2011

Abstract
—Tissue factor (TF) is a transmembrane protein considered to be responsible for the initiation of coagulation. TF gene expression may be induced in monocytes and endothelial cells and is present in atherosclerotic plaque to initiate thrombus formation. To investigate whether individual differences in TF gene expression could predispose subjects to thrombosis, we sequenced the 5′ domain of the gene up to nucleotide 2732 and found 6 different polymorphisms: 4 of them were completely concordant and defined 2 haplotypes with similar frequencies, designated as 1208 D and 1208 I. Genotyping of patients with myocardial infarction in a case-control study involving 2354 subjects showed no association between the polymorphisms and nonfatal coronary thrombosis. In another study involving 255 patients with venous thromboembolism and 1204 controls, allele
D
was less common in the cases (
P
=0.022). The odds ratio associated with the presence of at least 1
D
allele was 0.72 (
P
=0.031). Comparison of subgroups of control subjects who were homozygous for the
D
or
I
allele demonstrated a lower plasma TF concentration in
DD
homozygotes. These results indicate that the TF gene promoter exists in 2 major forms differing at 4 sites. The 1208 D haplotype is not associated with coronary thrombosis but is associated with reduced plasma TF levels and a lower risk of venous thrombosis.
Sphingosine-1-Phosphate and Lysophosphatidic Acid Stimulate Endothelial Cell Migration
Authors: Panetti Tracee Scalise , Nowlen Julie , Mosher Deane F.
Published: June 2011

Abstract
—Endothelial cell migration is necessary for the formation of new blood vessels. We investigated the effects of 2 lysophospholipid mediators, sphingosine-1-phosphate (S1P) and lysophosphatidic acid (LPA), on endothelial cell migration. S1P and LPA stimulated migration of fetal bovine heart endothelial cells (FBHEs) in a 3D-modified Boyden chamber assay with concentrations as low as 15 nmol/L stimulating a 2-fold change and concentrations in the 1- to 2-μmol/L range stimulating 14- to 20-fold changes. S1P specifically stimulated the migration of several endothelial cell strains but did not stimulate the migration of tumor cells or smooth muscle cells. LPA stimulated some endothelial and nonendothelial cell types to migrate. For FBHEs, S1P and LPA were mostly chemokinetic in checkerboard assays. S1P and LPA stimulated extracellular signal–regulated kinase 1/2 phosphorylation and enhanced paxillin localization to focal contacts, with no discernible change in the actin cytoskeleton in FBHEs. To characterize responsible receptor-dependent signaling pathways, we investigated the involvement of G
i
, Rho, and phosphoinositide 3-OH kinase in S1P- and LPA-stimulated migration. Although perturbation of all 3 signaling molecules resulted in decreased migration, the mechanisms underlying the decreased migration were different. Pertussis toxin treatment, to target G
i
, caused endothelial cells to develop dense bundles of F-actin and distribute paxillin staining to the cell periphery in response to S1P or LPA. Modification of Rho with C3 toxin disrupted the actin cytoskeleton. Inhibition of phosphoinositide 3-OH kinase decreased S1P- or LPA-induced endothelial cell migration with only minor disruption of the actin cytoskeleton. Inhibition of extracellular signal-regulated kinase kinase with PD98059 caused a loss of phosphorylation of extracellular signal–regulated kinase 1/2, similar to pertussis toxin, but only a minimal decrease in migration. These results indicate that S1P and, for some cells, LPA stimulate migration of endothelial cells through a mechanism that likely requires a balance between G
i
and Rho signaling to achieve the cytoskeletal remodeling necessary for cell migration.
C-Reactive Protein Is a Strong but Nonspecific Risk Factor of Fatal Stroke in Elderly Persons
Authors: Gussekloo Jacobijn , Schaap Marianne C. L. , Frölich Marijke , Blauw Gerard J. , Westendorp Rudi G. J.
Published: June 2011

Abstract
—An elevated level of C-reactive protein is a strong predictor of cardiovascular events in elderly persons. Whether C-reactive protein has direct adverse vascular effects or is a marker of aspecific systemic inflammation remains to be determined. The aim of this study was to investigate the relation between C-reactive protein and the occurrence of fatal strokes in elderly persons. In the Leiden 85-Plus Study, a population-based prospective follow-up study, we studied the levels of C-reactive protein in 80 participants who died from stroke within the first 5 years of follow-up. Levels of C-reactive protein were determined in serum samples at baseline. Levels of C-reactive protein were also determined in 82 control subjects who survived for the first 5 years of follow-up and in 83 participants who died from noncardiovascular causes. Mortality risks were estimated with logistic regression and adjusted for differences in age, sex, smoking, medication, total cholesterol, history of diabetes or hypertension, and previous cardiovascular events. Levels of C-reactive protein at baseline were 2-fold higher in subjects who died from stroke than in control subjects (median 5.7 versus 2.7 mg/L,
P
<0.005). The levels of C-reactive protein in subjects who died from stroke or from noncardiovascular causes were similar (median 5.7 versus 4.9 mg/L,
P
=0.7). The risk of death from stroke as well as from noncardiovascular causes increased linearly up to 10-fold in subjects with the highest levels of C-reactive protein at baseline (
P
<0.001). The levels of C-reactive protein were lower when more time had elapsed between blood sampling and time of death during follow-up (
P
=0.01). C-reactive protein is a strong but nonspecific risk factor of fatal stroke in old persons. The data do not support the idea that C-reactive protein has direct vascular effects that underlie fatal cerebrovascular disease.
Serum C-Reactive Protein and Self-Reported Stroke
Authors: Ford Earl S. , Giles Wayne H.
Published: June 2011

Abstract
—C-reactive protein may predict the risk of coronary heart disease, but its association with stroke has not been well studied. We used data from the Third National Health and Nutrition Examination Survey, conducted from 1988 to 1994, to examine the association between serum C-reactive protein concentrations and self-reported past history of stroke among 8850 US men and women aged ≥40 years. The unadjusted geometric mean of C-reactive protein concentration was higher among participants with stroke than those without stroke (0.45±0.02 versus 0.32±0.01,
P
<0.001). After adjusting for age, sex, race or ethnicity, education, smoking status, systolic blood pressure, serum cholesterol, high density lipoprotein cholesterol, history of diabetes mellitus, body mass index, and physical activity, the odds ratio for stroke among participants with C-reactive protein concentrations ≥0.55 mg/dL compared with participants with concentrations ≤0.21 mg/dL was 1.71 (95% CI 1.11 to 2.64 [odds ratio per mg/dL 1.19, 95% CI 1.05 to 1.34]). These cross-sectional data support findings from other studies suggesting that C-reactive protein concentration may be a risk factor or marker for stroke in the US population.
Increased LDL Cholesterol and Atherosclerosis in LDL Receptor–Deficient Mice With Attenuated Expression of Scavenger Receptor B1
Authors: Huszar Dennis , Varban Mariet Lee , Rinninger Franz , Feeley Roslyn , Arai Takeshi , Fairchild-Huntress Victoria , Donovan Michael J. , Tall Alan R.
Published: June 2011

Abstract
—Scavenger receptor BI (SR-BI) is a multiligand cell-surface receptor that plays a central role in high density lipoprotein homeostasis in rodents. To investigate a role for SR-BI in atherosclerosis, mice with attenuated SR-BI expression were crossed with low density lipoprotein (LDL) receptor–deficient mice. Compound-homozygous mutants showed increased plasma cholesterol, surprisingly due primarily to increased LDL cholesterol and apolipoprotein B levels. LDL turnover studies showed that this resulted from increased LDL cholesterol production rather than decreased LDL catabolism. Atherosclerotic lesion size was significantly increased in male compound-mutant mice relative to LDL receptor–deficient controls (93 427±16 079 versus 34 448±5 331 μm
2
, respectively;
P
=0.003). The proatherogenic effect of attenuated SR-BI expression may in part be due to increased LDL cholesterol levels. These findings suggest that upregulation of the receptor could have therapeutic potential for the treatment of atherosclerosis.
Human Plasma Phospholipid Transfer Protein Increases the Antiatherogenic Potential of High Density Lipoproteins in Transgenic Mice
Authors: van Haperen Rien , van Tol Arie , Vermeulen Pieter , Jauhiainen Matti , van Gent Teus , van den Berg Paul , Ehnholm Sonja , Grosveld Frank , van der Kamp Arthur , Rini de Crom
Published: June 2011

Abstract
—Plasma phospholipid transfer protein (PLTP) transfers phospholipids between lipoprotein particles and alters high density lipoprotein (HDL) subfraction patterns in vitro, but its physiological function is poorly understood. Transgenic mice that overexpress human PLTP were generated. Compared with wild-type mice, these mice show a 2.5- to 4.5-fold increase in PLTP activity in plasma. This results in a 30% to 40% decrease of plasma levels of HDL cholesterol. Incubation of plasma from transgenic animals at 37°C reveals a 2- to 3-fold increase in the formation of pre-β-HDL compared with plasma from wild-type mice. Although pre-β-HDL is normally a minor subfraction of HDL, it is known to be a very efficient acceptor of peripheral cell cholesterol and a key mediator in reverse cholesterol transport. Further experiments show that plasma from transgenic animals is much more efficient in preventing the accumulation of intracellular cholesterol in macrophages than plasma from wild-type mice, despite lower total HDL concentrations. It is concluded that PLTP can act as an antiatherogenic factor preventing cellular cholesterol overload by generation of pre-β-HDL.