bluuebunny/crossref_metadata_2025 huggingface dataset:
Cyclooxygenase-2 Is Widely Expressed in Atherosclerotic Lesions Affecting Native and Transplanted Human Coronary Arteries and Colocalizes With Inducible Nitric Oxide Synthase and Nitrotyrosine Particularly in Macrophages
Authors: Baker Christopher S. R. , Hall Roger J. C. , Evans Thomas J. , Pomerance Ariela , Maclouf Jacques , Creminon Christophe , Yacoub Magdi H. , Polak Julia M.
Published: June 2011

Abstract
—Inflammation appears to have a major role in the development of atherosclerotic lesions affecting native and transplanted coronary arteries. The subsequent risk of plaque rupture and acute ischemic events correlates with the degree of inflammation and may be modified by aspirin, an anti-inflammatory cyclooxygenase inhibitor. Cyclooxygenase-2 (Cox-2) and inducible nitric oxide synthase (iNOS) are involved in the inflammatory response via the rapid and exaggerated production of prostanoids and nitric oxide, both of which may have proatherosclerotic effects. These effects may be mediated by the formation of peroxynitrite in the case of nitric oxide and involve “cross talk” between the two enzyme systems. This study aimed to investigate native and transplant atherosclerosis for the presence and distribution of Cox-2 and iNOS. Immunocytochemical studies were performed on atherosclerotic lesions from patients with native (n=12) and transplant (n=5) coronary disease by using antibodies to Cox-2, iNOS, and nitrotyrosine (an indicator of peroxynitrite production). Control tissue was obtained from unused donor hearts and at the time of autopsy. Cox-2 and iNOS colocalized predominantly in macrophages/foam cells in both types of atherosclerosis. Cox-2 expression was also detected in medial smooth muscle cells and endothelial cells, including those of the vasa vasorum. Nitrotyrosine was found in the same distribution as that of iNOS and was colocalized with Cox-2 in macrophages. Cox-2 and iNOS are coexpressed in native and transplant atherosclerosis, possibly allowing for interaction between the enzymes and suggesting an alternative mechanism for the benefits of aspirin via inhibition of Cox-2 activity.
Effects of γ-Tocotrienol on ApoB Synthesis, Degradation, and Secretion in HepG2 Cells
Authors: Theriault Andre , Wang Qi , Gapor Abdul , Adeli Khosrow
Published: June 2011

Abstract
—γ-Tocotrienol (γ-T3), a naturally occurring analog of tocopherol (vitamin E), has been shown to have a hypocholesterolemic effect in animals and humans. Unlike tocopherol, it has also been shown to reduce plasma apoB levels in hypercholesterolemic subjects. The aim of this study was to define the mechanism of action of γ-T3 on hepatic modulation of apoB production using cultured HepG2 cells as the model system. HepG2 cells preincubated with γ-T3 were initially shown to inhibit the rate of incorporation of [
14
C]acetate into cholesterol in a concentration- and time-dependent manner, with a maximum 86±3% inhibition at 50 μmol/L observed within 6 hours. γ-T3, on the other hand, had no significant effect on the uptake of [
14
C]glycerol into pools of cellular triacylglycerol and phospholipid relative to untreated control. The rate of apoB synthesis and secretion was then studied by an [
35
S]methionine pulse-labeling experiment and quantified by immunoprecipitating apoB on chasing up to 3 hours. An average reduction of 24±3% in labeled apoB in the media was apparent with γ-T3 despite a 60±2% increase in apoB synthesis. Fractionation of secreted apoB revealed a relatively denser lipoprotein particle, suggesting a less stable particle. Using a digitonin-permeabilized HepG2 cell system, the effects of γ-T3 on apoB translocation and degradation in the endoplasmic reticulum were further investigated. The generation of a specific N-terminal 70-kDa proteolytic fragment proved to be a sensitive measure of the rate of apoB translocation and degradation. The abundance of this fragment increased significantly in γ-T3-treated cells relative to untreated control cells (50±21%) after 2 hours of chase. In addition, the presence of γ-T3 resulted in an average decrease of 64±8% in intact apoB. Taken together, the data suggest that γ-T3 stimulates apoB degradation possibly as the result of decreased apoB translocation into the endoplasmic reticulum lumen. It is speculated that the lack of cholesterol availability reduces the number of secreted apoB-containing lipoprotein particles by limiting translocation of apoB into the endoplasmic reticulum lumen.
Lipid and Apolipoprotein Predictors of Atherosclerosis in Youth
Authors: Rainwater David L. , McMahan C. Alex , Malcom Gray T. , Scheer W. Douglas , Roheim Paul S. , McGill Henry C. , Strong Jack P.
Published: June 2011

Abstract
—We compared serum lipid and apolipoprotein predictors of atherosclerosis in cases from the multicenter study, Pathobiological Determinants of Atherosclerosis in Youth (PDAY). The lipid measures included HDL cholesterol (HDL-C) and non-HDL-C, and the apolipoprotein measures included concentrations of apoA1, apoB, and Lp(a), and sizes of the apo(a) proteins. We tested whether the apolipoprotein measures predicted atherosclerotic lesions as well as the more traditional lipid measures. We estimated extent of lesions as fatty streaks or raised lesions (fibrous plaques, complicated or calcified lesions) in 3 sites: thoracic aorta, abdominal aorta, and right coronary artery. Neither apoA1 nor apoB measures were as strongly or consistently correlated with extent of lesions as the corresponding lipid measure (HDL-C and non-HDL-C, respectively). Beyond the basic model that included sex, age, race, smoking status, hypertension, and the lipid measures, apoA1 and apoB added only an average 1.3% increased explanatory ability to the model, whereas HDL-C plus non-HDL-C added an average 2.5%. The results suggest that the traditional lipid measures are more useful than apolipoprotein measures for detecting young persons at high risk of precocious atherosclerosis. Because of large racial differences, the two Lp(a)-related measures, Lp(a) concentrations and apo(a) size, were evaluated in blacks and whites separately. Under these circumstances, neither of the Lp(a)-related measures was strongly or consistently correlated with extent of lesions.
Possible Induction of Renal Dysfunction in Patients With Lecithin:Cholesterol Acyltransferase Deficiency by Oxidized Phosphatidylcholine in Glomeruli
Authors: Jimi Shiro , Uesugi Noriko , Saku Keijiro , Itabe Hiroyuki , Zhang Bo , Arakawa Kikuo , Takebayashi Shigeo
Published: June 2011

Abstract—To clarify the causes of renal dysfunction in familial lecithin:cholesterol acyltransferase (LCAT) deficiency, kidney samples from 4 patients with LCAT deficiency (3 homozygotes and 1 heterozygote) were examined immunohistochemically. All of the patients exhibited corneal opacities, anemia, renal dysfunction, deficiencies in plasma high density lipoprotein and LCAT activity and mass, and an increase in the ratio of plasma unesterified cholesterol to esterified cholesterol. Renal lesions began with the deposition of lipidlike structures in the glomerular basement membrane, and these structures accumulated in the mesangium and capillary subendothelium. By electron microscopy, 2 types of distinctive structure were found in glomerular lesions: vacuole structures and cross-striated, membranelike structures. The plasma oxidized phosphatidylcholine (oxPC) –modified low density lipoprotein (LDL) levels in LCAT-deficient subjects were significantly (P<0.01) higher than those in controls (1.30±0.82 versus 0.42±0.32 ng/5 μg LDL, respectively), and a significant (P<0.01) difference was observed even after adjustment for confounding factors by an analysis of covariance. The patient with the highest plasma oxPC-modified LDL had the most membranelike structures in the glomeruli and showed the greatest renal deterioration from a young age. In glomerular lesions, although there was an abundance of apoB and apoE, oil red O–positive lipids, macrophages, apoA1, and malondialdehyde were scarce. OxPC was found extracellularly in glomerular lesions, and although its distribution differed from that of apolipoproteins, it was quite similar to that of phospholipids. In conclusion, these results indicate that oxPC in plasma and glomeruli is distinctive for patients with LCAT deficiency. Therefore, oxPC may be a factor in the deterioration of kidneys in patients with familial LCAT deficiency.
Adhesive Interaction Between P-Selectin and Sialyl LewisxPlays an Important Role in Recurrent Coronary Arterial Thrombosis in Dogs
Authors: Ikeda Hisao , Ueyama Takahisa , Murohara Toyoaki , Yasukawa Hideo , Haramaki Nobuya , Eguchi Hiroyuki , Katoh Atsushi , Takajo Yoshinori , Onitsuka Ichiro , Ueno Takafumi , Tojo Shinichiro J. , Imaizumi Tsutomu
Published: June 2011

Abstract—Cell adhesion molecules may play an important role in the disease process of acute coronary syndromes. We have shown a neutralizing anti-P-selectin monoclonal antibody and a sialyl Lewisx-containing oligosaccharide (SLex-OS), an analogue of selectin ligand on leukocytes, reduce cyclic flow variations (CFVs) in a canine model of recurrent coronary arterial thrombosis, suggesting the important interaction between P-selectin and SLexfor the pathophysiology of these syndromes. However, the functional role of these adhesion molecules in the thrombotic process remains unclear. Therefore, we investigated effects of SLex-OS on CFVs, platelet P-selectin expression, and morphology of the stenotic site in the same model. Anesthetized open-chest dogs (n=34) were randomly divided into 4 groups after developing CFVs. Dogs intravenously received saline or graded doses of SLex-OS (5, 20, or 40 mg/kg bolus) infusion followed by a continuous infusion (5 mg · kg−1· h−1) for 60 minutes. By flow cytometric analysis, P-selectin expression on platelets after CFVs was significantly upregulated during CFVs. Immunohistochemical analysis revealed the incorporation of platelets with upregulated P-selectin within thrombi at the stenotic site. Microscopic observations revealed the presence of numerous platelets adhered to leukocytes at the stenotic site on the damaged endothelium. SLex-OS significantly reduced CFVs, inhibited the P-selectin expression on platelets, and prevented the adherence of platelets and leukocytes. These findings further support the notion that the adhesive interaction between P-selectin on platelets and SLexon leukocytes plays an important role in platelet-mediated thrombus formation in this model.
Mutations in Promoter Region of Thrombomodulin and Venous Thromboembolic Disease
Authors: Le Flem Léna , Picard Véronique , Emmerich Joseph , Gandrille Sophie , Fiessinger Jean-Noël , Aiach Martine , Alhenc-Gelas Martine
Published: June 2011

Abstract
—The present study was designed to analyze the thrombomodulin proximal promoter region spanning nucleotides −293 to −12 to search for polymorphisms that could modify thrombomodulin gene expression in patients with venous thromboembolic disease. The study population comprised 205 patients and 394 healthy subjects of similar age and sex distribution. No polymorphisms and only 1 point mutation (G-33A) were found. The G-33A mutation was present at the heterozygous state in 2 patients and in 1 control. Being more frequent in the patients (0.97%) than in the controls (0.25%), the G-33A mutation might be a risk factor for venous thrombosis. To investigate the effect of this mutation on the thrombomodulin promoter activity, the proximal promoter region of the gene (bearing or not bearing the G-33A mutation) was inserted into a promotorless expression vector, upstream of the firefly luciferase gene, and transiently transfected into EA.hy926 endothelial cells. Under the conditions of the assay, the G-33A mutation mildly decreased the promoter activity. This study confirms that abnormalities of the thrombomodulin proximal promoter are not frequent in patients with venous thromboembolism.
Temporal Gradient in Shear But Not Steady Shear Stress Induces PDGF-A and MCP-1 Expression in Endothelial Cells
Authors: Bao Xuping , Lu Chuanyi , Frangos John A.
Published: June 2011

Abstract—Three well-defined laminar flow profiles were created to distinguish the influence of a gradient in shear and steady shear on platelet-derived growth factor A (PDGF-A) and monocyte chemoattractant protein-1 (MCP-1) expression in human endothelial cells. The flow profiles (16 dyne/cm2maximum shear stress) were ramp flow (shear stress smoothly transited at flow onset), step flow (shear stress abruptly applied at flow onset), and impulse flow (shear stress abruptly applied for 3 s only). Ramp flow induced only minor expression of PDGF-A and did not increase MCP-1 expression. Step flow increased PDGF-A and MCP-1 mRNA levels 3- and 2-fold at 1.5 hours, respectively, relative to ramp flow. In contrast, impulse flow increased PDGF-A and MCP-1 expression 6- and 7-fold at 1.5 hours, and these high levels were sustained for at least 4 hours. These results indicate that a temporal gradient in shear (impulse flow and the onset of step flow) and steady shear (ramp flow and the steady component of step flow) stimulates and diminishes the expression of PDGF-A and MCP-1, respectively. NO synthase inhibitorNG-amino-l-arginine (L-NAA) was found to markedly enhance MCP-1 and PDGF-A expression induced by step flow, but decrease their expression induced by impulse flow, in a dose-dependent manner. NO donor spermine-NONOate (SPR/NO) dose-dependently reduced the MCP-1 and PDGF-A expression induced by impulse flow. Moreover, impulse flow was found to stimulate sustained (4 hours) IκB-α degradation andegr-1 mRNA induction. L-NAA prevented IκB-α degradation, whereas SPR/NO increased IκB-α resynthesis 2 hours after impulse flow. Both L-NAA and SPR/NO inhibited the impulse flow inducibility ofegr-1 4 hours after the flow stimulation. The results show that both NO induced by steady shear and NO donor inhibit temporal gradient in shear-induced MCP-1 and PDGF-A expression by downregulation of their respective transcription factors NFκB andegr-1, whereas NO induced by impulse flow stimulates MCP-1 and PDGF-A expression by upregulation of the transcription factors. The above findings suggest distinct roles of temporal gradient in shear and steady shear in atherogenesis in vivo.
Lipolytic Modification of LDL by Phospholipase A
2
Induces Particle Aggregation in the Absence and Fusion in the Presence of Heparin
Authors: Hakala Jukka K. , Öörni Katariina , Ala-Korpela Mika , Kovanen Petri T.
Published: June 2011

Abstract
—One of the first events in atherogenesis is modification of low density lipoprotein (LDL) particles in the arterial wall with ensuing formation of aggregated and fused lipid droplets. The accumulating particles are relatively depleted in phosphatidylcholine (PC). Recently, secretory phospholipase A
2
(PLA
2
), an enzyme capable of hydrolyzing LDL PC into fatty acid and lysoPC molecules, has been found in atherosclerotic arteries. There is also evidence that both LDL and PLA
2
bind to the glycosaminoglycan (GAG) chains of extracellular proteoglycans in the arterial wall. Here we studied the effect of heparin GAG on the lipolytic modification of LDL by PLA
2
. Untreated LDL, heparin-treated LDL, and heparin-bound LDL were lipolyzed with bee venom PLA
2
. In the presence of albumin, lipolysis resulted in aggregation in all 3 preparations of the LDL particles. Lipolysis of untreated LDL did not result in aggregation if albumin was absent from the reaction medium, and the lipolytic products accumulated in the particles rendering them negatively charged. However, heparin-treated and heparin-bound lipolyzed LDL particles aggregated even in the absence of albumin. Importantly, in the presence of albumin, some of the heparin-treated and heparin-bound lipolyzed LDL particles fused, the proportion of fused particles being substantially greater when LDL was bound to heparin during lipolysis. In summary, lipolysis of LDL PC by PLA
2
under physiological conditions, which allow transfer of the lipolytic degradation products to albumin, leads to fusion of LDL particles in the presence, but not in the absence, of heparin. Thus, it is possible that within the GAG meshwork of the arterial intima, PLA
2
-induced modification of LDL is one source of the lipid droplets during atherogenesis.
Role of Group II Secretory Phospholipase A2in Atherosclerosis
Authors: Ivandic Boris , Castellani Lawrence W. , Wang Xu-Ping , Qiao Jian-Hua , Mehrabian Margarete , Navab Mohamad , Fogelman Alan M. , Grass David S. , Swanson Mark E. , de Beer Maria C. , de Beer Frederick , Lusis Aldons J.
Published: June 2011

Abstract—Some observations have suggested that the extracellular group IIa phospholipase A2(sPLA2), previously implicated in chronic inflammatory conditions such as arthritis, may contribute to atherosclerosis. We have examined this hypothesis by studying transgenic mice expressing the human enzyme. Compared with nontransgenic littermates, the transgenic mice exhibited dramatically increased atherosclerotic lesions when maintained on a high-fat, high-cholesterol diet. Surprisingly, the transgenic mice also exhibited significant atherosclerotic lesions when maintained on a low-fat chow diet. Immunohistochemical staining indicated that sPLA2was present in the atherosclerotic lesions of the transgenic mice. On both chow and atherogenic diets, the transgenic mice exhibited decreased levels of HDLs and slightly increased levels of LDLs compared with nontransgenic littermates. These data indicate that group IIa sPLA2may promote atherogenesis, in part, through its effects on lipoprotein levels. These data also provide a possible mechanism for the observation that there is an increased incidence of coronary artery disease in many chronic inflammatory diseases.
Role of Group II Secretory Phospholipase A
2
in Atherosclerosis
Authors: Leitinger Norbert , Watson Andrew D. , Hama Susan Y. , Ivandic Boris , Qiao Jian-Hua , Huber Joakim , Faull Kym F. , Grass David S. , Navab Mohamad , Fogelman Alan M. , de Beer Frederick C. , Lusis Aldons J. , Berliner Judith A.
Published: June 2011

Abstract
—Secretory nonpancreatic phospholipase A
2
(group II sPLA
2
) is induced in inflammation and present in atherosclerotic lesions. In an accompanying publication we demonstrate that transgenic mice expressing group II sPLA
2
developed severe atherosclerosis. The current study was undertaken to determine whether 1 mechanism by which group II sPLA
2
might contribute to the progression of inflammation and atherosclerosis is by increasing the formation of biologically active oxidized phospholipids. In vivo measurements of bioactive lipids were performed, and in vitro studies tested the hypothesis that sPLA
2
can increase the accumulation of bioactive phospholipids. We have shown previously that 3 oxidized phospholipids derived from the oxidation of 1-palmitoyl-2-arachidonoyl-
sn
-glycero-3-phosphorylcholine (PAPC) stimulated endothelial cells to bind monocytes, a process that is known to be an important step in atherogenesis. We now show that these 3 biologically active phospholipids are significantly increased in livers of sPLA
2
transgenic mice fed a high-fat diet as compared with nontransgenic littermates. We present in vitro evidence for several mechanisms by which these phospholipids may be increased in sPLA
2
transgenics. These studies demonstrated that polyunsaturated free fatty acids, which are liberated by sPLA
2
, increased the formation of bioactive phospholipids in LDL, resulting in increased ability to stimulate monocyte-endothelial interactions. Moreover, sPLA
2
-treated LDL was oxidized by cocultures of human aortic endothelial cells and smooth muscle cells more efficiently than untreated LDL. Analysis by electrospray ionization–mass spectrometry revealed that the bioactive phospholipids, compared with unoxidized PAPC, were less susceptible to hydrolysis by human recombinant group II sPLA
2
. In addition, HDL from the transgenic mice and human HDL treated with recombinant sPLA
2
in vitro failed, in the coculture system, to protect against the formation of biologically active phospholipids in LDL. This lack of protection may in part relate to the decreased levels of paraoxonase seen in the HDL isolated from the transgenic animals. Taken together, these studies show that levels of biologically active oxidized phospholipids are increased in sPLA
2
transgenic mice; they also suggest that this increase may be mediated by effects of sPLA
2
on both LDL and HDL.
Determinants of Fasting and Post-Methionine Homocysteine Levels in Families Predisposed to Hyperhomocysteinemia and Premature Vascular Disease
Authors: de Jong S. C. , Stehouwer C. D. A. , van den Berg M. , Kostense P. J. , Alders D. , Jakobs C. , Pals G. , Rauwerda J. A.
Published: June 2011

Abstract
—Elevated plasma total homocysteine (tHcy) levels, either measured in the fasting state or after oral methionine loading, are associated with an increased risk of atherothrombotic disease. Fasting and post-methionine hyperhomocysteinemia (HHC) overlap to a limited extent; both can occur as familial traits. We investigated determinants of fasting, postmethionine and delta (ie, post-methionine minus fasting levels) tHcy levels in 510 subjects of 192 HHC-prone families including 161 patients with clinical vascular disease and 349 without vascular disease. We focused on tHcy levels in relation to levels of vitamin B
12
, B
6
and folate and the methylenetetrahydrofolate reductase (MTHFR) C677T mutation. Multivariate linear analyses adjusted for the presence of vascular disease showed that fasting tHcy was significantly related to folate and vitamin B
12
, and the presence of the MTHFR
TT
genotype and the
T
allele, and to age, smoking habits, and serum levels of creatinine. Both post-methionine and delta tHcy levels were related to serum folate levels, and the presence of the MTHFR
TT
genotype and the
T
allele, and to postmenopausal status, and body mass index. An interaction was found between MTHFR
TT
genotype and serum folate levels for both fasting and post-methionine tHcy, ie, for a given decrease in serum folate, homocysteine levels increased more in subjects with the
TT
genotype than in those with the
CC
genotype. Fasting, post-methionine and delta tHcy were higher in patients with vascular disease than in their healthy siblings, but these levels were less dependent on serum folate levels (
P
<0.05), whereas the effect of MTHFR genotype was stronger (
P
=0.01). This study found evidence that post-methionine and delta tHcy levels are not only influenced by factors affecting homocysteine transsulfuration but also by factors that affect remethylation. The explained variances of fasting, post-methionine and delta tHcy were 49%, 62%, and 78%, respectively. We also found evidence, in patients with premature vascular disease but not in their healthy siblings, for a factor that increases tHcy levels but weakens the normal inverse relation between folate and tHcy and amplifies the effect of the MTHFR genotype.
Relationship Between Lipoprotein- and Oxidation-Related Variables and Atheroma Lipid Composition in Subjects Undergoing Coronary Artery Bypass Graft Surgery
Authors: Craig Wendy Y. , Rawstron Ming W. , Rundell Clark A. , Robinson Elizabeth , Poulin Sue E. , Neveux Louis M. , Nishina Patsy M. , Keilson Leonard M.
Published: June 2011

Abstract
—The relationship between atheroma lipid composition and serum lipoprotein and oxidation measurements has not been fully explored. To address this question, we studied serum, plasma, and aortic wall specimens from 66 subjects undergoing coronary artery bypass graft surgery. The lipid composition of aortic specimens was characterized in terms of cholesterol ester and cholesterol crystal plus phospholipid by using hot-stage polarizing light microscopy; tissue oxidation status was assessed by measuring conjugated dienes. Serum lipoprotein–related measurements included total cholesterol, triglyceride, low density lipoprotein (LDL) cholesterol, high density lipoprotein (HDL) cholesterol, apolipoproteins B and AI, and lipoprotein(a). Oxidation status was assessed by measuring LDL mobility, thiobarbituric acid–reactive substances, LDL conjugated dienes, and IgG and IgM autoantibodies against oxidized LDL. Fasting blood glucose was also determined. Lesion cholesterol crystal plus phospholipid content was associated inversely with serum HDL cholesterol levels
(r
=−0.279,
P
=0.029) and positively with fasting blood glucose (
r
=0.359,
P
=0.016), LDL mobility (0.276,
P
<0.05), and IgM autoantibodies against oxidized LDL (
r
=0.272,
P
=0.037). There was also a significant relationship between the level of aortic tissue conjugated dienes and plasma LDL mobility (
r
=0.332,
P
=0.007). In multivariate analysis, IgM autoantibodies against oxidized LDL, fasting blood glucose, and LDL mobility, in descending order of significance, together accounted for 35% of the variability in aortic lesion cholesterol crystal plus phospholipid content. These data support direct and independent roles for oxidation and hyperglycemia in the pathophysiology of atherosclerosis.
Differential Expression of Connexin43 and Desmin Defines Two Subpopulations of Medial Smooth Muscle Cells in the Human Internal Mammary Artery
Authors: Ko Yu-Shien , Yeh Hung-I , Haw Marcus , Dupont Emmanuel , Kaba Riyaz , Plenz Gabriele , Robenek Horst , Severs Nicholas J.
Published: June 2011

Abstract
—Upregulation of connexin43-gap junctions is associated with transition of contractile vascular smooth muscle cells (SMCs) to the synthetic state. To determine whether phenotypically distinct subpopulations of medial SMCs differentially express connexin43, we investigated the human distal internal mammary artery, a structurally heterogeneous vessel with features ranging from elastic to elastomuscular to muscular. Immunoconfocal microscopy combined with quantitative analysis and complemented by in situ hybridization showed that SMCs in the elastic medial regions expressed high levels of connexin43 but low levels of desmin, whereas those of muscular medial regions expressed low levels of connexin43 but high levels of desmin. Ultrastructurally, SMCs of both regions were of the contractile phenotype, but the former cells were irregular in shape with relatively prominent synthetic organelles whereas the latter were spindle shaped with fewer synthetic organelles. Vimentin, smooth muscle α-actin, calponin, h-caldesmon, and myosin heavy chains (SM1 and SM2) were equally highly expressed by most cells in both subpopulations. The connexin43/desmin expression pattern of SMCs in regions of intimal thickening resembled those of elastic medial regions. These findings refine the view suggested from previous studies that high levels of connexin43 expression are associated with SMCs of a less contractile/more synthetic phenotype. In the internal mammary artery, the 2 subpopulations of SMCs with markedly different connexin43 expression levels both represent a differentiated contractile phenotype, but the subpopulation showing high levels of connexin43-gap junctions is characterized by low levels of desmin and structural features that reflect a more synthetic tendency.
Plasma Concentrations of Interleukin-6 and Abdominal Aortic Diameter Among Subjects Without Aortic Dilatation
Authors: Rohde Luis E. P. , Arroyo Luis H. , Rifai Nader , Creager Mark A. , Libby Peter , Ridker Paul M. , Lee Richard T.
Published: June 2011

Abstract
—Previous studies suggest that cytokine-induced tissue inflammation may participate in the pathogenesis of abdominal aortic aneurysms. Serum inflammatory markers may reflect arterial inflammation in asymptomatic phases of the aneurysmal disease. We studied 120 outpatients (62 men; age, 65±9 years) by ultrasound evaluation of the abdominal aorta to evaluate the association of circulating levels of interleukin-6 (IL-6) with abdominal aortic diameter in subjects with normal aortic size. Aortic diameter was measured at the infrarenal level and indexed for body surface area. Seven patients with abdominal aortic dilatation (indexed aortic diameter, >1.3 cm/m
2
) were also identified. Plasma concentrations of IL-6, serum amyloid A (SAA), C-reactive protein (CRP), total homocysteine, and lipids were measured. Among the 113 subjects without aortic dilatation, indexed aortic diameter was positively associated with serum levels of IL-6 (
P
<0.01), SAA (
P
<0.01), and total homocysteine (
P
=0.01). IL-6 levels increased in a stepwise fashion among dichotomized groups of aortic size (low and high aortic diameters) and peaked in patients with aortic dilatation (2.3±1.2 versus 2.7±0.9 versus 3.2±0.9 pg/mL, respectively;
P
for trend=0.039). None of the serum lipid measurements correlated with abdominal aortic diameter. Although CRP levels were associated with SAA levels (
r
=0.60;
P
<0.001), associations between CRP and aortic diameter were nonsignificant. In multivariate analysis, levels of IL-6 (
P
=0.02), SAA (
P
=0.001), and total homocysteine (
P
<0.001) were independent correlates of indexed aortic diameter. In conclusion, circulating levels of IL-6, SAA, and total homocysteine may reflect processes involved in the early phases of abdominal aortic aneurysm formation, before dilation of the abdominal aorta is established. These data support a role for chronic inflammation in the progression of asymptomatic aortic disease.
Overexpression of Human Catalase Gene Decreases Oxidized Lipid-Induced Cytotoxicity in Vascular Smooth Muscle Cells
Authors: Santanam Nalini , Augé Nathalie , Zhou Mimi , Keshava Channa , Parthasarathy Sampath
Published: June 2011

Abstract
—Reactive oxygen metabolites such as hydrogen peroxide (H
2
O
2
) and oxidized fatty acids are proinflammatory and are involved in the pathophysiology of various diseases including atherosclerosis. The effects of these oxidants could be inhibited by the external addition of an antioxidant, suggesting the promotion or propagation of further oxidation. In this study, we describe the stable overexpression of human catalase in smooth muscle cells and the resistance of these cells to cytotoxicity induced not only by the addition of H
2
O
2
but also by the addition of 13-hydroperoxyoctadecadienoic acid (13-HPODE). The results pose an intriguing possibility of the generation of H
2
O
2
from a peroxidized fatty acid. Accordingly, incubation of cells with both 13-HPODE and 13-hydroxyoctadecadienoic acid resulted in the generation of intracellular H
2
O
2
. To explain the observed results by which catalase could overcome the effects of 13-HPODE, we propose that oxidized fatty acids are degraded in the cellular peroxisomes, resulting in the generation of H
2
O
2
. In other words, the cellular effects of peroxidized fatty acids could be attributed to the generation of H
2
O
2
.
Membrane Active Lipids in Remnant Lipoproteins Cause Impairment of Endothelium-Dependent Vasorelaxation
Authors: Doi Hideki , Kugiyama Kiyotaka , Ohgushi Masamichi , Sugiyama Seigo , Matsumura Toshiyuki , Ohta Yasutaka , Oka Hideki , Ogata Nobuhiko , Hirata Akira , Yamamoto Yorihiro , Yasue Hirofumi
Published: June 2011

Abstract
—We have recently found that remnant lipoproteins (RLPs) and their lipid fractions impair endothelium-dependent vasorelaxation (EDR). This study was aimed at clarifying mechanisms responsible for RLP-induced endothelial dysfunction in isolated rabbit aortas. RLPs were isolated from plasma in hyperlipidemic subjects by use of the immunoaffinity gel mixture of anti-ApoA1 and anti-ApoB100 monoclonal antibodies and ultracentrifugation. Organ chamber experiments showed that EDR impairment was restored by addition of reduced glutathione (GSH) or
N
-acetylcysteine, antioxidants, into the incubation buffer containing isolated rabbit aortas and RLPs (0.75 mg of triglyceride/mL). Furthermore, the incubation of isolated human red blood cells (RBCs) with RLP and its lipids converted the normal shape of RBCs to echinocytes, but coincubation with antioxidants suppressed the RLP-induced RBC transformation, suggesting that they exerted oxidative damage on RBC surface membranes. Studies with HPLC and the postcolumn chemiluminescence method showed that RLPs contain a substantial amount of phosphatidylcholine hydroperoxides. Peroxidized phosphatidylcholine also impaired EDR and had echinocytogenic action, both of which were suppressed by
N
-acetylcysteine. RLPs isolated from the plasma of patients under treatment with α-tocopherol, an antioxidant, had a lower level of phosphatidylcholine hydroperoxides (15% of the amount in nontreated patients), which was associated with a lack of the inhibitory action on EDR and with lesser effect on RBC transformation. Oxidative damage caused by lipid components in RLPs, especially peroxidized phospholipids, deteriorates cell surface membrane and may be at least partly responsible for RLP-induced impairment of EDR.
Heart Rate Variability and Progression of Coronary Atherosclerosis
Authors: Huikuri Heikki V. , Jokinen Vesa , Syvänne Mikko , Nieminen Markku S. , Airaksinen K. E. Juhani , Ikäheimo Markku J. , Koistinen Juhani M. , Kauma Heikki , Kesäniemi Antero Y. , Majahalme Silja , Niemelä Kari O. , Frick M. Heikki
Published: June 2011

Abstract
—Low heart rate (HR) variability is associated with increased risk of cardiovascular morbidity and mortality, but the causes and mechanisms of this association are not well known. This prospective study was designed to test the hypothesis that reduced HR variability is related to progression of coronary atherosclerosis. Average HR and HR variability were analyzed in 12-hour ambulatory ECG recordings from 265 qualified patients participating in a multicenter study to evaluate the angiographic progression of coronary artery disease in patients with prior coronary artery bypass surgery and low high-density lipoprotein cholesterol concentrations (<1.1 mmol/L). Participants were randomized to receive a placebo or gemfibrozil therapy. The progression of coronary atherosclerosis was estimated by quantitative, computer-assisted analysis of coronary artery stenoses from the baseline angiograms and from repeated angiograms performed an average of 32 months later. The progression of focal coronary atherosclerosis of the patients randomized to placebo therapy was more marked in the tertile with the lowest standard deviation of all normal to normal R-R intervals (SDNN, 74±13 ms; mean decrease in the per-patient minimum luminal diameter −0.17 mm; 95% confidence interval [CI], −0.23 to −0.12 mm) than in the middle tertile (SDNN, 107±7 ms; mean decrease −0.05 mm; 95% CI, −0.08 to −0.01 mm) or highest tertile (SDNN, 145±25 ms; mean change 0.01 mm; 95% CI, −0.04 to 0.02 mm) (
P
<0.001 between the tertiles). This association was abolished by gemfibrozil. SDNN was lower (
P
<0.001) and minimum HR was faster (
P
<0.01) in the patients with marked progression than in those with regression of focal coronary atherosclerosis. In multiple regression analysis including HR variability, minimum HR, demographic and clinical variables, smoking, blood pressure, glucose, lipid measurements and lipid-modifying therapy, progression of focal coronary atherosclerosis was independently predicted by the SDNN (β=0.24;
P
=0.0001). Low HR variability analyzed from ambulatory ECG predicts rapid progression of coronary artery disease. HR variability provided information on progression of focal coronary atherosclerosis beyond that obtained by traditional risk markers of atherosclerosis.
Subunit Antigen and Activity Levels of Blood Coagulation Factor XIII in Healthy Individuals
Authors: Ariëns R. A. S. , Kohler H. P. , Mansfield M. W. , Grant P. J.
Published: June 2011

Abstract
—Factor (F) XIII covalently cross-links and stabilizes the fibrin-clot. Recent evidence suggests a role for FXIII in atherothrombotic diseases, but no information is available regarding the association of FXIII with common risk factors. The aim of this study was to investigate the relationship of FXIII with age, sex, smoking, and hypertension. Plasma levels of FXIII A-subunit antigen, FXIII B-subunit antigen, and FXIII cross-linking activity were measured in 612 healthy individuals (250 men and 362 women). FXIII A- and B-subunit levels were correlated significantly with age in both men (
r
=0.21,
P
=0.001, and
r
=0.17,
P
=0.008, respectively) and women (
r
=0.20,
P
<0.0005, and
r
=0.13,
P
=0.011, respectively). FXIII B-subunit levels and activity were correlated significantly with FXIII A-subunit levels (
r
=0.60,
P
<0.0005, and
r
=0.14,
P
<0.0005, respectively) and fibrinogen (
r
=0.26,
P
<0.0005, and
r
=0.14,
P
=0.001, respectively). Women had higher levels of FXIII A-subunit (111.8% versus 105.2%,
P
<0.01) and B-subunit (109.5% versus 103.8%,
P
<0.01) than did men. FXIII A-subunit was significantly increased in smokers (117.0% versus 104.6%,
P
<0.0005) and in subjects with hypertension (114.9% versus 107.8%,
P
<0.05). In a multiple regression model, FXIII A-subunit was significantly increased by female sex (+6.4%,
P
<0.007), smoking (+12.3%,
P
<0.0005), and increasing age (+3.7% per 10 years,
P
<0.0005). FXIII B-subunit was significantly related to female sex and fibrinogen, and FXIII activity was significantly related to fibrinogen levels. In conclusion, the FXIII A-subunit level increases significantly with female sex, age, and smoking, whereas FXIII B-subunit and FXIII activity are associated with FXIII A-subunit level and fibrinogen. Although evidence for a causal relationship between FXIII A-subunit and vascular disease is not available, these results might suggest a role for elevated FXIII A-subunit levels in the pathogenesis of vascular disease.
Neonatal Intima Formation in the Human Coronary Artery
Authors: Ikari Yuji , McManus Bruce M. , Kenyon Jennifer , Schwartz Stephen M.
Published: June 2011

Abstract
—Intimal masses develop in the human coronary arteries of all humans, becoming atherosclerotic in later life either because of focal accumulation of lipid or the resulting response to injury. We evaluated the time course of formation of the intimal mass in the proximal left anterior descending coronary artery in autopsy specimens from 91 patients between 17 weeks’ gestation and 23 months of postnatal age. Intima was rarely found before 30 weeks’ gestation; however, the frequency with which at least some intimal cells were observed increased to 35% between 36 weeks’ gestation and birth. By 3 months after birth, all patients had an intimal mass at this coronary location. The mean intima/media ratio was 0.1 just after birth and increased continuously to the second postnatal year. Replication of medial smooth muscle cells, indicated by proliferating cell nuclear antigen staining, was high before birth and decreased between birth and 2 years of age. However, the replication index of the intima remained at 2% to 5%. Thus, coronary intimal cells appearing in the perinatal period may arise by migration after replication of medial smooth muscle, as is seen in models of carotid artery balloon injury. In conclusion, formation of the coronary artery intima is a rapid process, beginning in the peripartum or postpartum period. Given the clonality of the adult lesion and the lack of proliferation in later stages of lesion formation, it is intriguing to speculate that this event may form the basis for atherosclerosis in later life.
Phosphatidylinositol 3-Kinase and Focal Adhesion Kinase Are Early Signals in the Growth Factor–Like Responses to Thrombospondin-1 Seen in Human Vascular Smooth Muscle
Authors: Lymn Joanne S. , Rao Sarafina J. , Clunn Gerard F. , Gallagher Karen L. , O’Neil Clive , Thompson Neil T. , Hughes Alun D.
Published: June 2011

Abstract
—Thrombospondin-1 (TSP-1) is a matricellular protein that is expressed in negligible amounts in normal blood vessels but is markedly upregulated in vascular injury. Although TSP-1 can act as a pleiotropic regulator for human vascular smooth muscle cells (HVSMCs), the intracellular signaling pathways stimulated by this protein remain obscure. In cultured HVSMCs derived from saphenous vein, TSP-1 induces tyrosine phosphorylation of a number of cellular proteins, with a complex temporal pattern of activation. Immunoprecipitation techniques have identified the early tyrosine-phosphorylated signals as being the p85 regulatory subunit of phosphatidylinositol 3-kinase (PI 3-K) and focal adhesion kinase (FAK). Tyrosine phosphorylation of the p85 subunit of PI 3-K showed a biphasic response to TSP-1 stimulation, which corresponded to a biphasic activation of the lipid kinase. Treatment with both wortmannin and LY294002 inhibited PI 3-K activity of HVSMCs but did not affect tyrosine phosphorylation of the p85 regulatory subunit. TSP-1–stimulated FAK phosphorylation, however, was substantially reduced by these inhibitors, as was the TSP-1–induced chemotaxis of these cells. These results suggest that activation of PI 3-K is an early signal induced by TSP-1 and is critical for chemotaxis. Activation of this kinase precedes and may occur upstream from FAK phosphorylation, although the nature of the interaction between these 2 enzymes remains obscure.
Local Hypomethylation in Atherosclerosis Found in Rabbit
ec-sod
Gene
Authors: Laukkanen Mikko O. , Mannermaa Sanna , Hiltunen Mikko O. , Aittomäki Saara , Airenne Kari , Jänne Juhani , Ylä-Herttuala Seppo
Published: June 2011

Abstract
—Extracellular superoxide dismutase (EC-SOD) protects arteries against deleterious effects of superoxide anions and the development of atherosclerosis. In this study, we cloned and characterized rabbit
ec-sod
gene. We identified 6 rabbit C-elements and 5 CpG clusters in the cloned sequence. One of the CpG clusters is located on the coding sequence. Because CpG clusters are potential sites for methylation and may explain the occurrence of mutations, methylation status of each of the CpG dimers located in the coding sequence CpG cluster was characterized using direct genomic sequencing. Unexpectedly, a marked reduction in the amount of methylated CpG dinucleotides in
ec-sod
gene was detected in atherosclerotic aortas as compared with normal aortic intima-media. Although alterations in DNA methylation are well characterized in malignant tumors, the presence of methylation changes in atherosclerosis has not been studied even though both diseases are characterized by excess cellular proliferation and alterations in gene expression. Further analysis of the whole genomic methylation by high-pressure liquid chromatography in normal and atherosclerotic aortas revealed a tendency for a decreased 5-methylcytosine (5-mC) content in atherosclerotic aortas as compared with normal arteries. Hypomethylation in atherosclerotic aortas occurred at the same level as has been reported from malignant tumors. Although a causal relationship between the methylation level and expression of EC-SOD cannot be proven, our results show that
ec-sod
hypomethylation is associated with the development of atherosclerosis and suggest that it may affect structure and function of
ec-sod
and other genes possibly involved in the development of atherosclerotic lesions.
Endotoxin Induces a Second Window of Protection in the Rat Heart as Determined by Using
p
-Nitro-Blue Tetrazolium Staining, Cardiac Troponin T Release, and Histology
Authors: Zacharowski Kai , Otto Mike , Hafner Gerd , Chatterjee Prabal K. , Thiemermann Christoph
Published: June 2011

Abstract
—Pretreatment of rats with small doses of lipopolysaccharide (LPS), eg, for 24 hours, attenuates the cardiac dysfunction caused by subsequent period of myocardial ischemia. This phenomenon of enhanced tolerance to an ischemic insult has been termed “second window of protection.” Although the cardioprotective effects of LPS were first reported in 1989, it is still unclear whether the observed attenuation by LPS of the ischemia-induced cardiac dysfunction is indeed secondary to the protection of cardiac myocytes against ischemic cell injury and death. This study was designed to investigate the effects of “preconditioning” with LPS on cell injury caused by regional myocardial ischemia and reperfusion in the anesthetized rat. Thirty-five Wistar rats were subjected to 25 minutes occlusion of the left anterior descending coronary artery followed by 2 hours of reperfusion. Hemodynamic parameters were continuously recorded, and at the end of the experiments, infarct size (using
p
-nitro-blue tetrazolium staining), cardiac troponin T release, and histological markers of cell injury and death were determined. In rats pretreated with a bolus of saline (vehicle for LPS) 2 or 24 hours before left anterior descending coronary artery occlusion and reperfusion, the infarct size was 59±4% (2 hours saline-control, n=6) and 61±3% (24 hours saline-control, n=6), respectively. Pretreatment of animals with a bolus of LPS (1 mg/kg IP) 24 hours before the onset of myocardial ischemia and reperfusion reduced both infarct size (to 18±7%;
P
<0.05, n=6) as well as histological signs of cell injury. Pretreatment (24 hours, as above) of rats with LPS also reduced the release of cardiac troponin T from 58±13 ng/mL (saline-control) to 16±9 ng/mL. In contrast, pretreatment of rats with LPS (2 hours, as above) did not affect infarct size (56±8%, n=6), cardiac troponin T release, or the histological parameters of cell injury. These data provide the first conclusive evidence that pretreatment of rats with a bolus of LPS 24 hours before intervention reduces the cell injury and death caused by a subsequent period of myocardial ischemia and reperfusion.
Aortic endothelial and smooth muscle histamine metabolism in experimental diabetes.
Authors: Orlidge A , Hollis T M
Published: June 2011

We studied histamine metabolism, i.e., histidine decarboxylase (HD)-mediated synthesis and histaminase-mediated catabolism, in relation to intracellular histamine content in both aortic endothelial and subjacent smooth muscle cells of control and diabetic rats. Diabetes was induced by a single jugular vein injection of streptozotocin (55 mg/kg in acidified saline, pH 4.5), and animals were held for either 2 or 4 weeks following overt manifestation of diabetes. An additional 4-week diabetic group received insulin (Iletin NPH, 10 U per 24 hour) during the last week. With respect to control values, the histamine content of aortic endothelial cells increased 138%, HD activity increased 250%, and histaminase activity decreased 50% over the 4-week period. In subjacent smooth muscle cells, the histamine content increased in excess of 150%, HD activity increased more than 300%, and histaminase activity decreased in excess of 30%. Insulin treatment for the last week resulted in complete reversal of all these changes. These results support the concept that a large vessel response similar to the microcirculatory prolonged phase of inflammation occurs in experimental diabetes, a change similar to that occurring in experimental atherosclerosis. They also indicate that both synthetic and catabolic changes occur in histamine metabolism under these conditions, changes that alter arterial wall histamine pools, and suggest that insulin administration under conditions of experimental diabetes may modulate aortic histamine metabolism and the resultant intraaortic histamine pools.