bluuebunny/crossref_metadata_2025 huggingface dataset:
Fc-γ Receptor Cross-Linking by Immune Complexes Induces Matrix Metalloproteinase-1 in U937 Cells via Mitogen-Activated Protein Kinase
Authors: Huang Yan , Fleming Andrew J. , Wu Shan , Virella Gabriel , Lopes-Virella Maria F.
Published: June 2011

Abstract—Matrix metalloproteinase-1 (MMP-1) secreted by macrophages potentially contributes to plaque rupture. Because large quantities of immunoglobulin G and ICs (ICs), including low density lipoprotein–containing ICs (LDL-ICs), are present in atherosclerotic lesions, we examined the effect of LDL-ICs on macrophage MMP-1 expression. With the use of ICs prepared with human LDL and rabbit anti-LDL antiserum, our enzyme-linked immunosorbent assays and Northern blots showed that MMP-1 secretion and expression by U937 histiocytes were induced by LDL-ICs. Furthermore, our results showed that blocking of Fc-γ receptor I and II (FcγRI and FcγRII) inhibited 70% and 55%, respectively, of the LDL-IC–induced secretion of MMP-1. Finally, our data showed that both PD98059, an inhibitor of the mitogen-activated protein kinase pathway, and Ro-31-2880, an inhibitor of protein kinase C, inhibited LDL-IC–stimulated MMP-1 secretion in a dose-dependent manner, with 96% and 95% inhibition, respectively, at the respective doses of 50 μmol/L and 80 nmol/L. In conclusion, this study demonstrated for the first time that LDL-ICs induce macrophage MMP-1 secretion by cocross-linking FcγRI and FcγRII and triggering a protein kinase C–dependent mitogen-activated protein kinase pathway. These results suggest that LDL-ICs and other ICs localized in atherosclerotic plaques may be potent stimulators for macrophage MMP-1 expression and may contribute to plaque rupture and acute coronary events.
Endoglin Is Overexpressed After Arterial Injury and Is Required for Transforming Growth Factor-β–Induced Inhibition of Smooth Muscle Cell Migration
Authors: Ma Xiaoli , Labinaz Marino , Goldstein Jason , Miller Harvey , Keon Wilbert J. , Letarte Michelle , O’Brien Edward
Published: June 2011

Abstract—Endoglin is a homodimeric membrane glycoprotein primarily expressed on endothelial cells. In association with transforming growth factor (TGF)-β receptors I and II, it can bind TGF-β1 and -β3 and form a functional receptor complex. There is increasing evidence that endoglin can modulate the cellular response to TGF-β, a factor implicated in vascular lesion formation in human and experimental models. The purpose of this study was to analyze the expression of endoglin in normal and balloon-injured porcine coronary arteries and in normal and atherosclerotic human coronary arteries and to determine its ability to mediate the effects of TGF-β on the migration of vascular smooth muscle cells (SMCs). In normal porcine coronary arteries, endoglin was of low abundance and was found primarily on endothelial cells and adventitial fibroblasts, as well as on a minority of medial SMCs. On days 3, 7, and 14 after angioplasty, endoglin was present not only on endothelial cells but also on adventitial myofibroblasts and medial SMCs of porcine coronary arteries. By day 28, few or no cells expressed endoglin. In situ hybridization revealed that endoglin mRNA expression appeared to be highest in endothelial cells on days 3, 7, and 14 days after injury and absent thereafter. With a second balloon injury, a similar pattern of endoglin protein and mRNA expression was observed. In human vascular tissue, endoglin immunolabeling was higher in endarterectomy specimens removed from diseased coronary arteries than in normal internal mammary arteries. In vitro, antisense oligonucleotides to endoglin decreased its expression and antagonized the TGF-β–mediated inhibition of human and porcine SMC migration. In summary, upregulation of endoglin occurs during arterial repair and in established atherosclerotic plaques and may be required for modulation of SMC migration by TGF-β.
Coadministration of Angiopoietin-1 and Vascular Endothelial Growth Factor Enhances Collateral Vascularization
Authors: Chae Jei Keon , Kim Injune , Lim Seok Tae , Chung Myoung Ja , Kim Won Ho , Kim Hwan Gyu , Ko Jae Ki , Koh Gou Young
Published: June 2011

Abstract
—Using growth factors to induce vasculogenesis is a promising approach in the treatment of ischemic legs and myocardium. Because the vasculogenesis requires a cascade of growth factors, their receptors, and intracellular signals, such therapies may require the application of more than a single growth factor. We examined the effect of 2 endothelial cell–specific growth factors, angiopoietin-1 (Ang1) and vascular endothelial growth factor (VEGF), on primary cultured porcine coronary artery endothelial cells. VEGF, but not Ang1, increased DNA synthesis and cell number. Ang1 or VEGF induced migration and sprouting activity, increased plasmin and matrix metalloproteinase-2 secretion, and decreased tissue inhibitors of metalloproteinase type 2 secretion. A combination of the submaximal doses of Ang1 and VEGF enhanced these effects and was more potent than the maximal dose of either alone. In a rabbit ischemic hindlimb model, a combination of Ang1 and VEGF gene delivery produced an enhanced effect on resting and maximal blood flow and capillary formation that was greater than that of either factor alone. Angiographic analyses revealed that larger blood vessels were formed after gene delivery of Ang1 or Ang1 plus VEGF than after VEGF gene delivery. These results suggest that combined treatment of Ang1 and VEGF could be used to produce therapeutic vascularization.
High Levels of Lp(a) With a Small Apo(a) Isoform Are Associated With Coronary Artery Disease in African American and White Men
Authors: Paultre Furcy , Pearson Thomas A. , Weil Henry F. C. , Tuck Catherine H. , Myerson Merle , Rubin Jill , Francis Charles K. , Marx Herbert F. , Philbin Edward F. , Reed Roberta G. , Berglund Lars
Published: June 2011

Abstract
—Elevated levels of lipoprotein(a) [Lp(a)] and the presence of small isoforms of apolipoprotein(a) [apo(a)] have been associated with coronary artery disease (CAD) in whites but not in African Americans. Because of marked race/ethnicity differences in the distribution of Lp(a) levels across apo(a) sizes, we tested the hypothesis that apo(a) isoform size determines the association between Lp(a) and CAD. We related Lp(a) levels, apo(a) isoforms, and the levels of Lp(a) associated with different apo(a) isoforms to the presence of CAD (≥50% stenosis) in 576 white and African American men and women. Only in white men were Lp(a) levels significantly higher among patients with CAD than in those without CAD (28.4 versus 16.5 mg/dL, respectively;
P
=0.004), and only in this group was the presence of small apo(a) isoforms (<22 kringle 4 repeats) associated with CAD (
P
=0.043). Elevated Lp(a) levels (≥30 mg/dL) were found in 26% of whites and 68% of African Americans, and of those, 80% of whites but only 26% of African Americans had a small apo(a) isoform. Elevated Lp(a) levels with small apo(a) isoforms were significantly associated with CAD (
P
<0.01) in African American and white men but not in women. This association remained significant after adjusting for age, diabetes mellitus, smoking, hypertension, HDL cholesterol, LDL cholesterol, and triglycerides. We conclude that elevated levels of Lp(a) with small apo(a) isoforms independently predict risk for CAD in African American and white men. Our study, by determining the predictive power of Lp(a) levels combined with apo(a) isoform size, provides an explanation for the apparent lack of association of either measure alone with CAD in African Americans. Furthermore, our results suggest that small apo(a) size confers atherogenicity to Lp(a).
A Locus Influencing Total Serum Cholesterol on Chromosome 19p
Authors: Imperatore Giuseppina , Knowler William C. , Pettitt David J. , Kobes Sayuko , Fuller John H. , Bennett Peter H. , Hanson Robert L.
Published: June 2011

Abstract
—A genome-wide linkage study was analyzed to identify loci that influence serum lipid concentrations in Pima Indians. Linkage analyses were conducted for total cholesterol measured in 998 siblings from 292 nuclear families, for total triglycerides in 547 siblings from 188 families, and for high density lipoprotein (HDL) cholesterol in 590 siblings from 201 families. Genotypes were generated for 516 autosomal microsatellite markers. Multipoint variance components methods were used to assess linkage. The strongest evidence for linkage with total cholesterol was on chromosome 19p (lod score 3.89), in the vicinity of the marker
D19S1034,
which is near the low density lipoprotein receptor gene. The strongest evidence for linkage with HDL cholesterol was on chromosome 3q (lod score 2.64) near
D3S3053
. For triglycerides, the strongest evidence for linkage was on chromosome 2p near
D2S1788
(lod score 1.70) and on chromosome 3p near
D3S2406
(lod score 1.77). This genomic scan provides evidence for a locus influencing total cholesterol concentration on chromosome 19p. It also suggests a locus influencing HDL cholesterol on chromosome 3q.
Stromelysin-1 and Interleukin-6 Gene Promoter Polymorphisms Are Determinants of Asymptomatic Carotid Artery Atherosclerosis
Authors: Rauramaa Rainer , Väisänen Sari B. , Luong Le-Anh , Schmidt-Trücksäss Arno , Penttilä Ilkka M. , Bouchard Claude , Töyry Jari , Humphries Steve E.
Published: June 2011

Abstract
—The functional 5A/6A polymorphism of the stromelysin-1 promoter has been implicated as a potential genetic marker for the progression of angiographically determined atherosclerosis in patients with coronary artery disease. Recently, a novel interleukin-6
(IL-6)
gene functional G/C polymorphism at −174 in the promoter has also been reported. In this study, we analyzed the relation of these two polymorphisms with carotid artery atherosclerosis in 109 randomly selected, middle-aged men without exercise-induced ischemia. Atherosclerosis was quantified as intima-media thickness (IMT) by high-resolution ultrasonography. Univariately, stromelysin genotype was significantly (
P
=0.015) associated with IMT, and this relation remained (
P
=0.033) after adjustments for age, cardiorespiratory fitness, body mass index, smoking, LDL cholesterol, and systolic blood pressure and for sonographers. The 5A/6A polymorphism independently explained 7% of the variance in carotid bifurcation IMT. The IL-6 polymorphism was also significantly associated (
P
=0.036) with increased IMT, with men homozygous for the G allele having IMT that was 11% greater than men homozygous for the C allele. Men who were homozygous for both the 6A and G alleles had an covariate adjusted IMT that was 36% greater than men who were homozygous for neither allele (
P
<0.003). These data suggest that genetic factors that predispose to reduced matrix remodeling (stromelysin 6A allele) and to increased inflammation (IL-6 G allele) combine to increase susceptibility for intima-media thickening in the carotid bifurcation, a predilection site for atherosclerosis.
Distribution of ApoA-I–Containing HDL Subpopulations in Patients With Coronary Heart Disease
Authors: Asztalos Bela F. , Roheim Paul S. , Milani Richard L. , Lefevre Michael , McNamara Judith R. , Horvath Katalin V. , Schaefer Ernst J.
Published: June 2011

Abstract
—High density lipoproteins (HDLs) and their subspecies play a role in the development of coronary heart disease (CHD). HDL subpopulations were measured by 2-dimensional nondenaturing gel electrophoresis in 79 male control subjects and 76 male CHD patients to test the hypothesis that greater differences in apolipoprotein (apo)A-I–containing HDL subpopulations would exist between these 2 groups than for traditional lipid levels. In CHD subjects, HDL cholesterol (HDL-C) was lower (−14%, P<0.001), whereas total cholesterol and the low density lipoprotein cholesterol/HDL-C ratio were higher (9% [
P
<0.05] and 21% [
P
<0.01], respectively) compared with control levels. No significant differences were found for low density lipoprotein cholesterol, triglyceride, and apoA-I levels. In CHD subjects, there were significantly (
P
<0.001) lower concentrations of the large lipoprotein (Lp)A-I α
1
(−35%), pre-α
1
(−50%), pre-α
2
(−33%), and pre-α
3
(−31%) subpopulations, whereas the concentrations of the small LpA-I/A-II α
3
particles were significantly (
P
<0.001) higher (20%). Because α
1
was decreased more than HDL-C and plasma apoA-I concentrations in CHD subjects, the ratios of HDL-C to α
1
and of apoA-I to α
1
were significantly (
P
<0.001) higher by 36% and 57%, respectively, compared with control values. Subjects with low HDL-C levels (≤35 mg/dL) have different distributions of apoA-I–containing HDL subpopulations than do subjects with normal HDL-C levels (>35 mg/dL). Therefore, we stratified participants according to HDL-C concentrations into low and normal groups. The differences in lipid levels between controls and HDL-C–matched cases substantially decreased; however, the significant differences in HDL subspecies remained. Our research findings support the concept that compared with control subjects, CHD patients not only have HDL deficiency but also have a major rearrangement in the HDL subpopulations with significantly lower α
1
and pre-α
1–3
(LpA-I) and significantly higher α
3
(LpA-I/A-II) particles.
Quantification of Cholesteryl Esters in Human and Rabbit Atherosclerotic Plaques by Magic-Angle Spinning
13
C-NMR
Authors: Peng Shaoqing , Guo Wen , Morrisett Joel D. , Johnstone Michael T. , Hamilton James A.
Published: June 2011

Abstract
—Accumulation of cholesteryl esters (CEs) is a key event in the formation of atherosclerotic plaques. More recent work suggests a role for CEs in plaque rupture leading to thrombosis, which can result in an acute event such as myocardial infarction or stroke. In this study, we present nuclear magnetic resonance (NMR) protocols for quantification of CEs in plaques in situ. Total CEs quantified by
13
C magic-angle spinning (MAS) NMR in excised plaques from human carotid arteries and rabbit aortic arteries were in good agreement with the amounts determined by subsequent standard chemical assays. The latter analysis is disadvantageous because it requires that plaque lipids be extracted from the tissue, resulting in the loss of all phase information of CEs as well as other major plaque components. With our MAS-NMR protocol, the plaque components are preserved in their native phases. Combining MAS and off-MAS NMR, we were able to quantitatively distinguish isotropic (liquid) CEs from anisotropic (liquid-crystalline) CEs in plaque tissues. In a recent study, we applied a different
13
C MAS-NMR protocol to quantify crystalline cholesterol monohydrate in plaques. Together, these 2 studies describe a new, noninvasive MAS-NMR strategy for the identification and quantification of the major lipid components in plaques in situ. This approach will be useful for investigation of the relationship between plaque rupture and specific lipids in their biologically relevant phases.
Role of Matrix Metalloproteinases in Blood Flow–Induced Arterial Enlargement
Authors: Tronc François , Mallat Ziad , Lehoux Stéphanie , Wassef Michel , Esposito Bruno , Tedgui Alain
Published: June 2011

Abstract
—Tears in the internal elastic lamina (IEL) can be observed after chronic increases in arterial blood flow, suggesting a potential role for matrix metalloproteinases (MMPs) in flow-induced vascular remodeling. We undertook to study this phenomenon by constructing an arteriovenous fistula (AVF) between the left common carotid artery (CCA) and the external jugular vein in rabbits. The diameter of the flow-loaded left CCA increased by 13.6±1.8% by day 3 after construction of the AVF compared with the right CCA (n=4,
P
<0.01) and by 40.7±7.5% by day-15 (n=10,
P
<0.0001). Increased CCA diameter also coincided with IEL fragmentation. Three days after construction of the AVF, gelatin zymography of protein extracts from left CCAs of untreated rabbits showed a significant increase in the 62-kDa (active MMP-2) activity and the appearance of a lytic band at 92 kDa (pro–MMP-9). In further experiments, MMP activity was inhibited by treatment with doxycycline (DOX) or BB-94, a specific MMP inhibitor. The increase in the 62-kDa gelatinolytic band was abolished in DOX- and BB-94–treated rabbits. The 92-kDa gelatinolytic band was also reduced in DOX-treated animals. Furthermore, both increased left CCA diameter and IEL fragmentation were abolished in DOX- and BB-94–treated rabbits. To evaluate whether nitric oxide was involved in blood flow–induced MMP activation, the rabbits were treated with
N
G
-nitro-
l
-arginine methyl ester to inhibit nitric oxide synthesis. MMP activities were significantly decreased in the left CCAs of
N
G
-nitro-
l
-arginine methyl ester–treated animals. Hence, blood flow–induced MMP activation is critical in flow-induced vascular enlargement and IEL fragmentation, and blood flow–induced nitric oxide participates in MMP activation.
Expression of Fas Ligand in Arteries of Hypercholesterolemic Rabbits Accelerates Atherosclerotic Lesion Formation
Authors: Schneider Darren B. , Vassalli Giuseppe , Wen Shan , Driscoll Robert M. , Sassani André B. , DeYoung Mary Beth , Linnemann Ruth , Virmani Renu , Dichek David A.
Published: June 2011

Abstract
—Fas ligand (FasL) is expressed by cells of the arterial wall and is present in human atherosclerotic lesions. However, the role of FasL in modifying the initiation and progression of atherosclerosis is unclear. To investigate the role of arterial FasL expression in the development of atherosclerosis, we first established a model of primary lesion formation in rabbit carotid arteries. In this model, infusion of adenoviral vectors into surgically isolated, nondenuded arteries of hypercholesterolemic rabbits leads to the formation of humanlike early atherosclerotic lesions. Expression of FasL in arterial endothelium in this model decreased T-cell infiltration and expression of vascular cell adhesion molecule-1 but did not affect expression of intercellular adhesion molecule-1. Intimal lesions grew more rapidly in FasL-transduced arteries than in arteries transduced with a control adenovirus that did not express a transgene. Total intimal macrophage accumulation was increased in FasL-transduced arteries; however, the proportion of lesion area occupied by macrophages was not elevated. The accelerated lesion growth was primarily due to the accumulation of intimal smooth muscle cells with a synthetic proliferative phenotype. There was no significant apoptosis in FasL-transduced or control arteries and no granulocytic infiltrates. Thus, the net result of elevated FasL expression is to accelerate atherosclerotic lesion growth by increasing lesion cellularity. Vascular expression of FasL may contribute to the progression of atherosclerosis.
Mouse Model of Transplant Arteriosclerosis
Authors: Dietrich Hermann , Hu Yanhua , Zou Yiping , Dirnhofer Stephan , Kleindienst Roman , Wick Georg , Xu Qingbo
Published: June 2011

Abstract
—Transplant-accelerated arteriosclerosis in coronary arteries is the major limitation to long-term survival of patients with heart transplantation. The pathogenesis of this disease is not fully understood. Herein, we describe a simplified model of artery allografts in the mouse that allows us to take advantage of transgenic, knockout, or mutant animals. Common carotid arteries or aortic vessels were end-to-end allografted into carotid arteries between C57BL/6J and BALB/c mice. Neointimal lesions were observed as early as 2 weeks after surgery and had progressed at 4 and 6 weeks postoperatively. The lumen of grafted arteries was significantly narrowed due to neointima hyperplasia 4 weeks after transplantation. Using this model, we studied the role of intercellular adhesion molecule-1 (ICAM-1) in the development of transplant arteriosclerosis in ICAM-1–deficient mice. Neointimal lesions of artery grafts from ICAM-1 –/– C57BL/6J to BALB/c mice were reduced up to 60% compared with wild-type controls. MAC-1 (CD11b/18)-positive cells adhering to the surface of ICAM-1 –/– artery grafts were significantly less as identified by en face immunofluorescence, and these positive cells were more abundant in intimal lesions of artery grafts in wild-type mice. Furthermore, the major cell component of neointimal lesions 4 weeks after surgery was found to be α-actin–positive smooth muscle cells, which were significantly reduced in lesions of ICAM-1 –/– artery grafts. Thus, this model has been proven to be useful for understanding the mechanism of transplant arteriosclerosis. Our findings demonstrate that ICAM-1 is critical in the development of allograft arteriosclerosis via mediation of leukocyte adhesion to, and infiltration into, the vessel wall.
Adhesion of Monocyte Very Late Antigen-4 to Endothelial Vascular Cell Adhesion Molecule-1 Induces Interleukin-1β–Dependent Expression of Interleukin-6 in Endothelial Cells
Authors: Zohlnhöfer Dietlind , Brand Korbinian , Schipek Katharina , Pogatsa-Murray Gisela , Schömig Albert , Neumann Franz-Josef
Published: June 2011

Abstract
—In atheroma, T cell–derived interferon-γ (INF-γ) stimulates endothelial cells and facilitates recruitment of monocytes. We investigated potential mechanisms by which these interactions could contribute to local and systemic inflammatory responses. Specifically, we analyzed the expression of interleukin (IL)-1β and IL-6 in both cell types after coculture, the relevant adhesion molecules in this interaction, and transcriptional control by NF-κB. We studied coculture of purified peripheral blood monocytes with human umbilical vein endothelial cells (HUVECs), which were stimulated with INF-γ (10
6
U/L) to model the activated endothelium of atherosclerotic lesions. Coculture of monocytes with activated HUVECs resulted in release of IL-1β (40.6±3 pg/24 h,
P
=0.002) and IL-6 (46.6±7 ng/24 h,
P
=0.0015). Electrophoretic mobility gel shift assay and Northern blotting in each cell type separately revealed NF-κB activation in both cell types, IL-1β mRNA expression predominantly in monocytes, and IL-6 mRNA expression predominantly in HUVECs. The endothelial IL-6 release was IL-1–dependent, because it was suppressed by IL-1 receptor antagonist. Experiments with blocking antibodies demonstrated that binding of monocyte very late antigen-4 (VLA-4) to endothelial vascular cell adhesion molecule-1 (VCAM-1) was necessary for the induction of IL-1β in monocytes. Binding of monocyte VLA-4 to endothelial VCAM-1 induces NF-κB activation in both cell types with expression and release of IL-1β by monocytes, which in turn stimulates endothelial release of IL-6. The β
1
-integrin–mediated expression of IL-1β and IL-6 could contribute to local and systemic inflammatory reactions in atherosclerosis.
ACE
Gene Polymorphism in Cardiovascular Disease
Authors: Agerholm-Larsen Birgit , Nordestgaard Børge G. , Tybjærg-Hansen Anne
Published: June 2011

Abstract
—The objective of this study was to assess the influence of the
ACE
gene insertion (I)/deletion (D) polymorphism on plasma ACE activity; blood pressure; and risk of myocardial infarction, ischemic heart disease, and ischemic cerebrovascular disease by comparing small and large studies. The meta-analyses are based on a literature search of MEDLINE up until April 1998 and assessment of bibliographies of published studies and reviews. Forty-six studies were selected, including a total of 32 715 white individuals. Plasma ACE activity was increased 40% and 71% for
ID
and
DD
versus
II
in small studies and 21% and 48% in large studies (small versus large:
P
<0.001 and
P
<0.001). Blood pressure was not influenced by genotype. Risk of myocardial infarction and ischemic heart disease was increased by 47% and 29%, respectively, for
DD
versus
ID
and
II
genotypes in small studies but not in large studies (small versus large:
P
<0.001 for risk of myocardial infarction and
P
=0.01 for risk of ischemic heart disease). Risk of ischemic cerebrovascular disease was not increased either in the small or in the largest study. In conclusion, the
ACE
gene polymorphism affects plasma ACE activity but not blood pressure and is not associated with increased risk of myocardial infarction, ischemic heart disease, or ischemic cerebrovascular disease in the largest studies.
p53, p21
WAF1/CIP1
, and MDM2 Involvement in Proliferation and Apoptosis in an In Vitro Model of Conditionally Immortalized Human Vascular Smooth Muscle Cells
Authors: Hsieh J.-Kuang , Kletsas Dimitris , Clunn Gerard , Hughes Alun D. , Schachter Michael , Demoliou-Mason Catherine
Published: June 2011

Abstract
—Using an in vitro model of a conditionally immortalized cell line, we investigated how human vascular smooth muscle cells (VSMCs) are affected by the expression of simian virus 40 (SV40) large T antigen (LT antigen), which binds to cell cycle regulators, such as the tumor suppressor protein p53. Cells were obtained after infection of saphenous vein–derived VSMCs with a nonreplicative retroviral vector containing a temperature-sensitive (ts) mutant of SV40 LT antigen and were shown to have maintained some characteristics and responses of VSMCs. Under permissive temperature conditions (36°C), the increased rate of cell proliferation was shown to be associated with expression of LT antigen and with LT-antigen binding to and inactivation of p53. p53 inactivation failed to block apoptosis induced by serum withdrawal or by UV irradiation. Downregulation of LT-antigen expression at the nonpermissive temperature (39°C) was shown to be associated with growth arrest, increased expression of the cell cycle inhibitor p21
WAF1/CIP1
, increased MDM2-promoter activity, and differential expression of MDM2 gene products, suggesting that p53-induced transcription/transactivation may be involved in VSMC cell cycle control but not necessarily apoptosis. The established SMC line HVTs-SM1 may be a useful model for the study of processes involved in myointimal hyperplasia and cellular aging, as well as for the study of cell cycle control in general.
Cyclooxygenase-2 Regulates Granulocyte-Macrophage Colony–Stimulating Factor, but Not Interleukin-8, Production by Human Vascular Cells
Authors: Stanford Salome J. , Pepper John R. , Mitchell Jane A.
Published: June 2011

Abstract
—Vascular smooth muscle is now recognized as an important site of mediator generation under inflammatory conditions. Indeed, the release of leukocyte activators, such as granulocyte-macrophage colony–stimulating factor (GM-CSF) and interleukin (IL)-8, by human arterial smooth muscle cells has recently been demonstrated. However, the potential for venous cells to release GM-CSF has not been addressed. We have shown that human vascular smooth muscle cells express the “inflammatory” form of cyclooxygenase (COX), cyclooxygenase-2 (COX-2), when stimulated with cytokines. In some nonvascular cell types, the COX activity has been shown to regulate the release of GM-CSF and IL-8, although the nature of the isoform responsible was not addressed. We show that human venous smooth muscle cells, like their arterial counterparts, release GM-CSF after stimulation with IL-1β. Similarly, both cell types released IL-8. Under the same conditions, we found that COX-2 activity suppressed GM-CSF, but not IL-8, release by both types of human vascular cells. Moreover, the prostacyclin mimetic, cicaprost, and the cAMP analogue, dibutyryl cAMP, inhibited GM-CSF release from these cells. These observations suggest that COX-2 activity suppresses GM-CSF release via a cAMP-dependent pathway in human vascular cells and illustrates a novel mechanism by which this enzyme can modulate immune and inflammatory events.
Association of the C−514T Polymorphism in the Hepatic Lipase Gene With Variations in Lipoprotein Subclass Profiles
Authors: Couture Patrick , Otvos James D. , Cupples L. Adrienne , Lahoz Carlos , Wilson Peter W. F. , Schaefer Ernst J. , Ordovas Jose M.
Published: June 2011

Abstract
—Hepatic lipase is involved in the metabolism of several lipoproteins and has a key role in reverse cholesterol transport. A common C-to-T substitution at position −514 of the hepatic lipase promoter has been associated with variations in plasma high density lipoprotein cholesterol (HDL-C) levels and hepatic lipase activity. The aim of the current study was to investigate the association of this polymorphism to lipoprotein levels in a population-based sample of 1314 male and 1353 female Framingham Offspring Study participants. In men and women, carriers of the −514
T
allele had higher HDL-C and apolipoprotein A-I (apoAI) concentrations compared with noncarriers. The higher HDL-C levels associated with the −514
T
allele was due to an increase in the HDL
2
-C subfraction, and this association was stronger in women compared with men (
P
=0.0043 versus 0.0517). To gain further understanding about the metabolic basis of these effects, HDL and low density lipoprotein (LDL) subclass profiles were measured by using automated nuclear magnetic resonance spectroscopy and gradient gel electrophoresis, respectively. The association of the −514
T
allele with higher HDL-C levels seen in men and women was primarily due to significant increases in the large HDL subfractions (size range 8.8 to 13.0 nm). In contrast, there was no relationship between the hepatic lipase polymorphism at position −514 and the LDL particle size distribution after adjustment for familial relationships, age, body mass index, smoking, alcohol intake, use of β-blockers, apoE genotype, and menopausal status and estrogen therapy in women. Moreover, multiple regression analyses suggested that the C−514T polymorphism contributed significantly to the variability of HDL particle size in men and women (
P
<0.04). Thus, our results show that the C−514T polymorphism in the hepatic lipase gene is associated with significant variations in the lipoprotein profile in men and women.
Local Plasminogen Activator Inhibitor Type 1 Overexpression in Rat Carotid Artery Enhances Thrombosis and Endothelial Regeneration While Inhibiting Intimal Thickening
Authors: Hasenstab David , Lea Holly , Clowes Alexander W.
Published: June 2011

Abstract
—Elevated levels of plasminogen activator inhibitor type 1 (PAI-1) are found in advanced atherosclerotic plaque compared with normal vessel and may contribute to plaque progression and complications associated with plaque rupture. Increased expression of PAI-1 probably contributes to the thrombotic properties of advanced atherosclerotic plaque by impeding plasmin generation and degradation of fibrin. To test this hypothesis, we have deliberately created synthetic neointimas by seeding onto the denuded luminal surface of rat carotid arteries smooth muscle cells transduced with replication-defective retrovirus encoding rat PAI-1. This cell-based gene transfer method results in stable, long-term, and localized gene expression. PAI-1 overexpression increases mural thrombus accumulation at 4 days but decreases neointimal area by 30% and 25% at 1 week and 2 weeks, respectively. PAI-1 overexpression accelerates reendothelialization of injured arteries compared with control arteries at 1 week, 2 weeks, and 1 month. PAI-1 overexpression does not alter matrix accumulation at 1 week. Increased PAI-1 expression in the rat carotid artery enhances thrombosis and endothelial regeneration while inhibiting intimal thickening. These results suggest that PAI-1 could play a direct role in the development of advanced atherosclerotic plaque and in the repair of the diseased vessel after fibrous cap disruption.
No Association Between Dehydroepiandrosterone Sulfate and Development of Atherosclerosis in a Prospective Population Study (Bruneck Study)
Authors: Kiechl Stefan , Willeit Johann , Bonora Enzo , Schwarz Siegfried , Xu Qingbo
Published: June 2011

Abstract
—Antiatherogenic properties of dehydroepiandrosterone (DHEA) have been postulated for >40 years. Large-scale epidemiological studies on this important issue, however, are still sparse, and those available have yielded contradictory results. The Bruneck Study involved a large random sample of men and women aged 40 to 79 years that were enrolled in 1990 and reevaluated 5 years later. Baseline DHEA sulfate (DHEAS) levels were measured in 867 subjects after an overnight fast. Development and progression of carotid atherosclerosis was monitored by high-resolution duplex ultrasound. DHEAS levels declined with advancing age (29% and 44% per decade in men and women) and showed a complex sex-specific association with various vascular risk attributes and factors conferring protection against atherosclerosis. Age- and sex-adjusted DHEAS baseline levels did not differ between subjects with or without incident/progressive atherosclerosis (geometric mean 1161 versus 1253 μg/L). After adjustment for vascular risk factors and potential confounders, the odds ratio of incident/progressive atherosclerosis comparing a 50% increase in DHEAS levels was 0.99 (95% CI 0.89 to 1.11). Lack of an association between DHEAS and atherogenesis was confirmed in sex-specific and a variety of supplementary analyses. Statistical power would be high enough to detect differences in DHEAS between outcome categories as low as 15% (α=0.05). This prospective community-based study does not support a role for endogenous DHEA(S) in the development of human atherosclerosis.
Hypoxia Induces Transcription of the Plasminogen Activator Inhibitor-1 Gene Through Genistein-Sensitive Tyrosine Kinase Pathways in Vascular Endothelial Cells
Authors: Uchiyama Tsuyoshi , Kurabayashi Masahiko , Ohyama Yoshio , Utsugi Toshihiro , Akuzawa Nobuhiro , Sato Mahito , Tomono Shouichi , Kawazu Shoji , Nagai Ryozo
Published: June 2011

Abstract
—A decline in oxygen concentration perturbs endothelial function, which promotes local thrombosis. In this study, we determined whether hypoxia in the range of that observed in pathophysiological hypoxic states stimulates plasminogen activator inhibitor-1 (PAI-1) production in bovine aortic endothelial cells. PAI-1 production, measured by ELISA, was increased by 4.7-fold (
P
<0.05 versus normoxic control, n=4) at 12 hours after hypoxic stimulation. Northern blot analysis showed the progressive time-dependent increase in the steady-state level of PAI-1 mRNA expression by hypoxia, which reached a 7.5-fold increase (
P
<0.05 versus control, n=4) at 12 hours. Deferoxamine, which has been known to bind heme protein and to reproduce the hypoxic response, induced PAI-1 production at both the mRNA and protein levels. The half-life of PAI-1 mRNA, as determined by a standard decay assay, was not affected by hypoxia, suggesting that induction of PAI-1 mRNA was regulated mainly at the transcriptional level. Transient transfection assays of the human PAI-1 promoter–luciferase construct indicates that a hypoxia-responsive region lies between −414 and −107 relative to the transcription start site, where no putative hypoxia response element is found. The hypoxia-mediated increase in PAI-1 mRNA levels was attenuated by the tyrosine kinase inhibitors genistein (50 μmol/L) and herbimycin A (1 μmol/L), whereas PD98059 (50 μmol/L, MEK1 inhibitor), SB203580 (10 μmol/L, p38 mitogen-activated protein kinase inhibitor), and calphostin C (1 μmol/L, protein kinase C inhibitor) had no effect on the induction of PAI-1 expression by hypoxia and deferoxamine. Genistein but not daidzein blocked the production of hypoxia- and deferoxamine-induced PAI-1 protein. Thus, we conclude that hypoxia stimulates PAI-1 gene transcription and protein production through a signaling pathway involving genistein-sensitive tyrosine kinases in vascular endothelial cells.
Comparative In Vitro Efficacy of Different Platelet Glycoprotein IIb/IIIa Antagonists on Platelet-Mediated Clot Strength Induced by Tissue Factor With Use of Thromboelastography
Authors: Mousa Shaker A. , Khurana Sandeep , Forsythe Mark S.
Published: June 2011

Abstract
—In the present study, the in vitro efficacy of different platelet glycoprotein IIb/IIIa (GPIIb/IIIa) antagonists on platelet-fibrin–mediated clot strength under shear was compared with the antiaggregatory efficacy by using tissue factor (TF) thromboelastography (TEG). The ability of platelets to augment the elastic properties of blood clots under shear conditions was measured by computerized TEG under conditions of maximal platelet activation accelerated by recombinant TF. Under these conditions, platelets significantly enhance clot strength 8-fold (relative to platelet-free fibrin clots). This effect was inhibited to a different extent by various platelet GPIIb/IIIa receptor antagonists; this inhibition appears to be dependent on the transmission of platelet contractile force to fibrin via the GPIIb/IIIa receptors. The GPIIb/IIIa antagonists with high binding affinity for resting and activated platelets and slow platelet dissociation rates (class I) but not those with fast platelet dissociation rates (class II) demonstrated potent and comparable inhibition of platelet aggregation and TF-TEG clot strength. Platelet GPIIb/IIIa antagonists of class I, such as XV459 (free-acid form of roxifiban), DMP802, XV454, and c7E3, demonstrated comparable inhibitory dose responses of TF-TEG clot strength and platelet aggregation, with an IC
50
of 50 to 70 nmol/L. In contrast, platelet GPIIb/IIIa antagonists from class II, with comparable antiaggregatory efficacy, such as DMP728, YZ202 (free-acid form of orbofiban), YZ211 (free-acid form of sibrafiban), YZ751, and other antagonists, have a much lower efficacy in altering the strength of TF-mediated clot formation (IC
50
>1.0 μmol/L). These data suggest differential efficacy among different GPIIb/IIIa antagonists in inhibiting platelet-fibrin clot retraction despite of equivalent antiaggregatory potency.
Antisense Oligodeoxynucleotide Inhibition of Vascular Angiotensin-Converting Enzyme Expression Attenuates Neointimal Formation
Authors: Morishita Ryuichi , Gibbons Gary H. , Tomita Naruya , Zhang Lunan , Kaneda Yasufumi , Ogihara Toshio , Dzau Victor J.
Published: June 2011

Abstract
—It has been proposed that vascular angiotensin-converting enzyme (ACE) plays an important role in regulating vascular growth. Indeed, ACE inhibitors have been reported to prevent neointimal formation after vascular injury in a rat carotid artery model. However, classic pharmacological experiments cannot exclude the potential contributions of hemodynamics and the circulating renin-angiotensin system (RAS). In this study, we used antisense oligodeoxynucleotide (ODN) to obtain local blockade of vascular ACE expression without effects on systemic hemodynamics and circulating RAS. To increase the effectiveness of antisense action, we modified the hemagglutinating virus of Japan–liposome ODN delivery method by cotransfection with nuclear protein (high mobility group 1 [HMG-1]) and RNase H. In vitro experiments showed the enhanced efficacy of antisense ODN by cotransfection of HMG-1 and RNase H compared with ODN alone. In vivo transfection of antisense ACE ODNs into intact uninjured rat carotid artery resulted in a significant reduction of vascular ACE activity, and cotransfection of HMG-1 and RNase H showed further reduction. We examined the effects of local blockade of vascular ACE expression on neointimal formation after vascular injury. Transfection of antisense ACE ODNs resulted in the attenuation of neointimal formation, whereas sense and scrambled ODNs did not. Blood pressure, heart rate, and serum ACE activity were not affected by antisense treatment. The magnitude of vascular ACE inhibition correlated with the suppression of the neointimal size. Overall, this study demonstrates that local antisense ODN inhibition of vascular ACE expression attenuates neointimal formation independent of hemodynamics and circulating RAS. The results support the existence of a functional tissue angiotensin system in the rat vessel wall.
Blockade of Angiotensin II Type 1 Receptor and Not of Endothelin Receptor Prevents Hypertension and Cardiovascular Disease in Transgenic (mREN2)27 Rats via Adrenocortical Steroid–Independent Mechanisms
Authors: Rossi Gian Paolo , Sacchetto Alfredo , Rizzoni Damiano , Bova Sergio , Porteri Enzo , Mazzocchi Giuseppina , Belloni Anna S. , Bahcelioglu Meltem , Nussdorfer Gastone G. , Pessina Achille C.
Published: June 2011

Abstract
—We investigated the role of angiotensin II (Ang II) and endothelin-1 (ET-1) in transgenic (mREN2)27 rats, a model of the monogenic renin-dependent form of severe hypertension and cardiovascular disease. Four-week-old heterozygous male transgenic (mREN2)27 rats (n=24) were matched according to body weight (BW) and blood pressure (BP) and randomly allocated to receive a placebo (group P), the mixed endothelin type A and B receptor antagonist bosentan (100 mg/kg BW PO, group B), the Ang II type 1–specific receptor antagonist irbesartan (50 mg/kg BW PO, group I), or the endothelin type A–selective antagonist BMS-182874 (52 mg/kg BW PO, group BMS). After 4 weeks of treatment, during which BW and BP were measured weekly, animals were euthanized, and the heart, left ventricle, right ventricle, adrenal gland, brain, and kidney were weighed. The plasma levels of adrenocortical steroids were measured by high-performance liquid chromatography. The tension responses of ET-free segments of the thoracic aorta to 5×10
−6
mmol/L phenylephrine, 60 mmol/L KCl, and cumulative doses of ET-1 were assessed. The density of ET-1 receptor subtypes in the aorta and vascular structural changes in the mesenteric arterioles (100 to 200 μm ID) were also measured with autoradiography and myography, respectively. Compared with all other groups, group I rats showed significantly (
P
<0.001) lower systolic BP (group I, 161±8 mm Hg; group P, 269±23 mm Hg; group B, 275±17 mm Hg; and group BMS, 254±21 mm Hg), left ventricular weight (2.28±0.15 versus 3.71±0.26, 3.38±0.27, and 3.96±0.51 mg/g BW, respectively), tension responses to vasoconstrictors, and normalized media thickness of the mesenteric arterioles (22.3±0.6 versus 25.3±0.5, 25.5±0.7, and 24.1±1.5 μm, respectively). Compared with levels in group P (78±25 pmol/mL), plasma aldosterone levels were significantly decreased in group B (51±11 pmol/mL) and group I (40±16 pmol/mL). Thus, endogenous ET-1 and Ang II contribute to the regulation of aldosterone, but only Ang II is crucial for the development of hypertension and related target organ damage via the Ang II type 1 receptor. Endogenous Ang II does not appear to enhance cardiovascular production of ET-1 in this model of hypertension within the time span of our experiment.
Apoptosis and Regulation of Bax and Bcl-X Proteins During Human Neonatal Vascular Remodeling
Authors: Kim Hyo-Soo , Hwang Kyung-Kuk , Seo Jeong-Wook , Kim So-Young , Oh Byung-Hee , Lee Myoung-Mook , Park Young-Bae
Published: June 2011

Abstract
—To verify that apoptosis is one of the possible mechanisms of neonatal vascular remodeling during the transition from fetal to neonatal circulation, we assayed for apoptosis and evaluated the expression of apoptosis-regulatory proteins in umbilical vessel versus ascending aorta, ductus arteriosus (DA) versus adjacent pulmonary artery and aorta, or aorta versus its branching arteries. Twenty-two umbilical cords (UCs), 6 DAs with adjacent aortas and pulmonary arteries, and 4 aortic arches with their branching great arteries were obtained from neonates. Smooth muscle cell (SMC) apoptosis in umbilical vessels was identified in all UCs. The expressions of Bax and Bcl-X were stronger in umbilical artery than in the neonatal aorta, but Bcl-2 was weak in both arteries in immunohistochemistry. In the immunoblot analysis of UCs, the expression of the proapoptotic short isoform of Bcl-X was stronger than in other tissue, and caspase-3 was selectively activated, whereas it was not in the other components of the cardiovascular system. In contrast, the expression patterns of the FasAg and Fas ligand were similar in umbilical artery and aorta. Regulation of Bcl-2 family proteins was also observed in other vascular sites at which SMCs undergo apoptosis on hemodynamic changes during birth, such as the DA and the branching points of the great arteries from the aortic arch. Apoptosis is involved in the regression of human umbilical vessels and the DA and in the remodeling of the branching great arteries during the neonatal period, when Bcl-2 family proteins are likely to play a key role.
Heme Oxygenase-1 Is a cGMP-Inducible Endothelial Protein and Mediates the Cytoprotective Action of Nitric Oxide
Authors: Polte Tobias , Abate Aida , Dennery Phyllis A. , Schröder Henning
Published: June 2011

Abstract
—Inducible heme oxygenase (HO-1) has recently been recognized as an antioxidant and cytoprotective gene. By use of Western blotting, cell viability analysis, and antisense technique, the present study investigates the involvement of HO-1 in endothelial protection induced by the clinically used nitric oxide (NO) donor molsidomine (specifically, its active metabolite 3-morpholinosydnonimine [SIN-1]) and the second messenger cGMP. In bovine pulmonary artery endothelial cells, SIN-1 and
S
-nitroso-
N
-acetyl-
d,l
-penicillamine (SNAP) at 1 to 100 μmol/L induced the synthesis of HO-1 protein in a concentration-dependent fashion up to 3-fold over basal levels. HO-1 induction by SIN-1 was inhibited in the presence of the NO scavenger phenyl-4,4,5,5,-tetramethylimidazoline-1-oxyl-3-oxide and the soluble guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazole[4,3-
a
]quinoxalin-1-one. 8-Bromo-cGMP (1 to 100 μmol/L) and dibutyryl cGMP (1 to 100 μmol/L) as well as the activator of particulate guanylyl cyclase atrial natriuretic peptide (1 to 100 nmol/L) produced increases in HO-1 protein similar to those produced by SIN-1. SIN-1 and 8-bromo-cGMP increased heme oxygenase activity (bilirubin formation). Cytoprotection by NO donors was abrogated in the presence of the heme oxygenase inhibitor tin protoporphyrin IX. Pretreatment of cells with a phosphorothioate-linked HO-1 antisense oligonucleotide prevented protection by SIN-1 or 8-bromo-cGMP against tumor necrosis factor-α cytotoxicity, whereas sense and scrambled HO-1 were without effect under these conditions. Our results show for the first time that HO-1 is a cGMP-sensitive endothelial gene and establish conclusively a causal relationship between HO-1 induction and endothelial protection by the NO/cGMP system. By targeting cytoprotective HO-1, NO donors may therefore be expected to induce antioxidant, antiatherogenic, and anti-inflammatory effects.
Intracellular Ca
2+
Handling in Vascular Smooth Muscle Cells Is Affected by Proliferation
Authors: Vallot Olivier , Combettes Laurent , Jourdon Philippe , Inamo Jocelyn , Marty Isabelle , Claret Michel , Lompré Anne-Marie
Published: June 2011

Abstract
—Despite intensive interest in the dedifferentiation process of vascular smooth muscle cells, very little data are available on intracellular Ca
2+
signaling. The present study was designed to investigate the evolution of the intracellular Ca
2+
pools when rat aortic smooth muscle cells (RASMCs) proliferate and to define the mechanisms involved in the functional alterations. RASMCs were cultured in different conditions, and [Ca
2+
]
i
was measured by use of fura 2. Expression of the sarco(endo)plasmic reticulum Ca
2+
pumps (SERCA2a and SERCA2b), Ca
2+
channels, the ryanodine receptor (RyR), and the inositol trisphosphate receptor (IP3R) was studied by reverse transcription–polymerase chain reaction and immunofluorescence. Antibodies specific for myosin heavy chain isoforms were used as indicators of the differentiation state of the cell, whereas an anti–proliferating cell nuclear antigen antibody was a marker of proliferation. SERCA2a, SERCA2b, RyR3, and IP3R-1 mainly were present in the aorta in situ and in freshly isolated RASMCs. These cells used the 2 types of Ca
2+
channels to release Ca
2+
from a common thapsigargin-sensitive store. Proliferation of RASMCs, induced by serum or by platelet-derived growth factor-BB, resulted in the disappearance of RyR and SERCA2a mRNAs and proteins and in the loss of the caffeine- and ryanodine-sensitive pool. The differentiated nonproliferative phenotype was maintained in low serum or in cells cultured at high density. In these conditions, RyR and SERCA2a were also present in RASMCs. Thus, expression of RyR and SERCA2a is repressed by cell proliferation, inducing loss of the corresponding Ca
2+
pool. In arterial smooth muscle, Ca
2+
release through RyRs is involved in vasodilation, and suppression of the ryanodine-sensitive pool might thus alter the control of vascular tone.