bluuebunny/crossref_metadata_2025 huggingface dataset:
Increased Ischemia-Induced Angiogenesis in the Staggerer Mouse, a Mutant of the Nuclear Receptor Rorα
Authors: Besnard Sandrine , Silvestre Jean-Sébastien , Duriez Micheline , Bakouche Joëlle , Lemaigre-Dubreuil Yolande , Mariani Jean , Levy Bernard I. , Tedgui Alain
Published: September 2007

Rorα is an orphan nuclear receptor. In homozygous staggerer mutant mice (Rora
sg/sg
), a deletion within the Rora gene leads to an overexpression of inflammatory cytokines. Because inflammation and hypoxia are 2 key stimuli of ischemia-induced angiogenesis, we studied the role of Rorα in this setting. Ischemia was induced by ligation of the right femoral artery in C57BL/6 Rora
+/+
and Rora
sg/sg
mice. After 3 and 28 days, angiogenesis was evaluated by microangiography, measurement of capillary density using immunohistochemistry (anti-CD31), and measurement of blood flow by laser Doppler imaging. At day 3, angiographic score and blood flow were similar in Rora
sg/sg
mice and in Rora
+/+
littermates. Conversely, at day 28, Rora
sg/sg
mice showed a significant 2-fold increase in angiographic score and a 3-fold increase in capillary density within the ischemic hindlimb compared with control. Functionally, this coincided with a significant rise in leg perfusion in Rora
sg/sg
mice (0.83±0.05 for ischemic/nonischemic leg perfusion ratio) compared with Ror
+/+
mice (0.66±0.04,
P
<0.05). In addition, more extensive angiogenesis in Rora
sg/sg
mice correlated with an increased expression of eNOS protein by 83±12% and 71±24% at 3 and 28 days, respectively (
P
<0.05), whereas the level of the antiangiogenic cytokine IL-12 was significantly reduced by 38±10% at day 28 (
P
<0.05). Conversely, no changes in VEGF expression were observed. Our study identifies for the first time a new role for Rorα as a potent negative regulator of ischemia-induced angiogenesis.
Modulation of Endothelial Cell Growth Arrest and Apoptosis by Vascular Endothelial Growth Inhibitor
Authors: Yu Jingyi , Tian Song , Metheny-Barlow Linda , Chew Li-Jin , Hayes Andrew J. , Pan Hongguang , Yu Guo-Liang , Li Lu-Yuan
Published: September 2007

Vascular endothelial growth inhibitor (VEGI), a new member of the tumor necrosis factor family, is an endothelial cell–specific gene and a potent inhibitor of endothelial cell proliferation, angiogenesis, and tumor growth. We report here that VEGI mediates the following two activities in endothelial cells: early G
1
arrest in G
0
/G
1
cells responding to growth stimuli, and programmed death in proliferating cells. G
0
/G
1
-synchronized bovine aortic endothelial cells were treated with VEGI before and after the onset of the growth cycle. When the cells were stimulated with growth conditions but treated simultaneously with VEGI, a reversible, early-G
1
growth arrest occurred, evidenced by the lack of late G
1
markers such as hyperphosphorylation of the retinoblastoma gene product and upregulation of the
c-myc
gene. Additionally, VEGI treatment led to inhibition of the activities of cyclin-dependent kinases CDK2, CDK4, and CDK6. In contrast, VEGI treatment of cells that had entered the growth cycle resulted in apoptotic cell death, as evidenced by terminal deoxytransferase labeling of fragmented DNA, caspase 3 activation, and annexin V staining, all of which were lacking in nonproliferating cells treated with VEGI. Additionally, stress-signaling proteins p38 and JNK were not as fully activated by VEGI in quiescent as compared with proliferating populations. These findings suggest a dual role for VEGI, the maintenance of growth arrest and induction of apoptosis, in the modulation of the endothelial cell cycle.
Aortic Wall Mechanics and Composition in a Transgenic Mouse Model of Marfan Syndrome
Authors: Marque Valérie , Kieffer Pascal , Gayraud Barbara , Lartaud-Idjouadiene Isabelle , Ramirez Francesco , Atkinson Jeffrey
Published: September 2007

Abstract
—In Marfan syndrome, mutations of the fibrillin gene (
FBN1
) lead to aneurysm of the thoracic aorta, making the aortic wall more susceptible to dissection, but the precise sequence of events underlying aneurysm formation is unknown. We used a rodent model of Marfan syndrome, the mgR/mgR mouse (with mgR: hypomorphic
FBN1
mutation), which underexpresses FBN1, to distinguish between a defect in the early formation of elastic fibers and the later disruption of elastic fibers. The content of desmosine plus isodesmosine was used as an index of early elastogenesis; disruption of elastic fibers was analyzed by histomorphometry. Because disruption of the medial elastic fibers may produce aortic stiffening, so amplifying the aneurysmal process, we measured thoracoabdominal pulse wave velocity as an indicator of aortic wall stiffness. Both mgR/mgR and wild-type (C57BL/6J–129SV) strains were normotensive, and wall stress was not significantly modified because the increase in internal diameter (0.80±0.06 vs 0.63±0.03 mm in wild type,
P
<0.05) was accompanied by increased medial cross-sectional area. The aortic wall stiffened (4-fold increase in the elastic modulus–to–wall stress ratio). Desmosine content was not modified (mgR/mgR 432±31 vs wild type 492±42 μg/mg wet weight,
P
>0.05). Elastic fibers showed severe fragmentation: the percentage of the media occupied by elastic fibers was 18±3% in mgR/mgR mice vs 30±1% in wild-type mice, with the number of elastic segments being 1.9±0.2 vs 1.4±0.1×10
−6
/mm
2
in the wild type (both
P
<0.05). In conclusion, underexpression of FBN1 in mice leads to severe elastic network fragmentation but no change in cross-linking, together with aortic dilatation. This result suggests that fragmentation of the medial elastic network and not a defect in early elastogenesis is 1 of the determinants of aortic dilatation in Marfan syndrome.
HMG-CoA Reductase Inhibitor Modulates Monocyte–Endothelial Cell Interaction Under Physiological Flow Conditions In Vitro
Authors: Yoshida Masayuki , Sawada Taisuke , Ishii Hideto , Gerszten Robert E. , Rosenzweig Anthony , Gimbrone Michael A. , Yasukochi Yukio , Numano Fujio
Published: September 2007

Abstract
—3-Hydroxy-3-methylglutaryl coenzyme A reductase inhibitors, or statins, have been reported to exert actions independent of their lipid-lowering effects. To critically assess the effects of statins on monocyte–endothelial cell interactions, we used an in vitro model that mimicked physiological flow conditions. Monocytic U937 cells were incubated in the presence of cerivastatin for 48 hours. Adhesive interactions of statin-treated U937 cells were then analyzed by use of activated (interleukin-1β 10 U/mL, 4 hours) human umbilical vein endothelial cells in an in vitro flow apparatus. Flow cytometric analysis of adhesion molecules and measurement of F-actin content in U937 cells were performed before and after statin treatment. Preincubation with cerivastatin significantly decreased U937 firm adhesion to activated human umbilical vein endothelial cells, whereas U937 rolling was not decreased. Fluorescence-activated cell sorter analysis revealed downregulation of U937 surface expression of CD11a, CD18, and VLA4 after statin treatment. Cerivastatin significantly reduced F-actin content in U937 cells and inhibited RhoA translocation, whereas preincubation with C3 exoenzyme reduced U937 adhesion under flow. Cerivastatin reduces monocyte adhesion to vascular endothelium under physiological flow conditions via downregulation of integrin adhesion molecules and inhibition of actin polymerization via RhoA inactivation. Our findings have important implications for the lipid-independent effects of statins.
Phenotypic Heterogeneity Influences Apoptotic Susceptibility to Retinoic Acid and
cis
-Platinum of Rat Arterial Smooth Muscle Cells In Vitro
Authors: Orlandi Augusto , Francesconi Arianna , Cocchia Domenico , Corsini Alberto , Spagnoli Luigi Giusto
Published: September 2007

Abstract
—Rat aortic smooth muscle cells (SMCs) cultured from intimal thickening 15 days after endothelial injury (IT-15), unlike those of normal media, show a monolayered, epithelioid phenotype and high levels of cellular retinol binding protein-1 (CRBP). Epithelioid clones obtained from the normal media suggest a “mosaicism” of arterial SMCs. Intimal cell homeostasis from the balance of proliferation and apoptosis is critical for the progression of vascular lesions. All-
trans
retinoic acid (tRA) reduced [
3
H]thymidine incorporation and G
1
→S phase progression of IT-15 and epithelioid clone but not of normal media and IT 60 days after injury (IT-60) SMCs. Hoechst staining, flow cytometry, and ligation-mediated polymerase chain reaction showed an increased susceptibility of IT-15 and epithelioid clone to tRA and
cis
-diaminedichloroplatinum II (CDDP)–induced apoptosis and cytotoxicity compared with normal media and IT-60 cells. The latter retained an increased susceptibility to tRA-induced apoptosis compared with normal media SMCs. tRA-induced apoptosis associated with an increased ratio of bax to bcl-2 by bax overexpression and cleavage of caspase-3. Anti-CRBP but not anti-IgG antibody prevented tRA-induced apoptosis and changes in related signaling molecules but not CDDP effects. Our findings support the relevant role of phenotypic heterogeneity in the determining proliferative as well as apoptotic behavior of arterial SMCs.
Apoptotic Death of Inflammatory Cells in Human Atheroma
Authors: Li Wei , Dalen Helge , Eaton John W. , Yuan Xi-Ming
Published: September 2007

Abstract
—Although the accumulation of cholesterol and other lipidic material is unquestionably important in atherogenesis, the reasons why this material progressively accumulates, rather than being effectively cleared by phagocytic cells such as macrophages, are not completely understood. We hypothesize that atheromatous lesions may represent “death zones” that contain toxic materials such as oxysterols and in which monocytes/macrophages become dysfunctional and apoptotic. Indeed, cathepsins B and L, normally confined to the lysosomal compartment, are present in the cytoplasm and nuclei of apoptotic (caspase-3–positive) macrophages within human atheroma. The possible involvement of oxysterols is suggested by experiments in which cultured U937 and THP-1 cells exposed to 7-oxysterols similarly undergo marked lysosomal destabilization, caspase-3 activation, and apoptosis. Like macrophages within atheroma, intralysosomal cathepsins B and L are normally present in the cytoplasm and nuclei of these oxysterol-exposed cells. Lysosomal destabilization, cathepsin release, and apoptosis may be causally related, because inhibitors of cathepsins B and L suppress oxysterol-induced apoptosis. Thus, toxic materials such as 7-oxysterols in atheroma may impair the clearance of cholesterol and other lipidic material by fostering the apoptotic death of phagocytic cells, thereby contributing to further development of atherosclerotic lesions.
LDL-Activated p38 in Endothelial Cells Is Mediated by Ras
Authors: Zhu Yi , Liao Hailing , Wang Nanping , Ma Kuo-Sheng , Verna Lynne K. , Shyy John Y.-J. , Chien Shu , Stemerman Michael B.
Published: September 2007

Abstract
—Endothelial dysfunction is a major atherogenic proinflammatory event. LDL causes the activation and phenotypic changes of cultured vascular endothelial cells (ECs). We previously reported that LDL activates c-Jun and AP-1 in ECs. In this study, we demonstrated that p38–ATF-2 is activated by LDL in human ECs and that this activation is mediated by Ras. When ECs are incubated with LDL in pathophysiological concentrations, the p38-mediated ATF-2 phosphorylation and ATF-2 transactivation are increased in a time- and dose-dependent manner. To elucidate the upstream mechanism in LDL-activated p38 in ECs, we demonstrate that LDL increases Ras translocation from the cytoplasm to the cellular membrane, with concurrent increases in Ras binding activity to GST–Raf-1. Overexpression of RasN17, a dominant negative mutant of Ras, attenuates the LDL-induced increases in (1) phosphorylation of ATF-2, (2) phosphorylation of c-Jun, (3) AP-1 binding, and (4) AP-1–driven luciferase activity. To study the effect of p38 in the regulation of an LDL targeting gene, we show that a specific p38 inhibitor attenuates LDL-induced E-selectin at the mRNA level. Thus, LDL activates both p38 and JNK signaling pathways through Ras activation, and furthermore, these events may play an important role in LDL-induced endothelial activation.
Role of the Matrix Metalloproteinase and Plasminogen Activator–Plasmin Systems in Angiogenesis
Authors: Pepper Michael S.
Published: September 2007

Abstract—Extracellular proteolysis is an absolute requirement for new blood vessel formation (angiogenesis). This review examines the role of the matrix metalloproteinase (MMP) and plasminogen activator (PA)–plasmin systems during angiogenesis. Specifically, a role for gelatinases (MMP-2, MMP-9), membrane-type 1 MMP (MMP-14), the urokinase-type PA receptor, and PA inhibitor 1 has been clearly defined in a number of model systems. The MMP and PA-plasmin systems have also been implicated in experimental vascular tumor formation, and their role during this process will be examined. Antiproteolysis, particularly in the context of angiogenesis, has become a key target in therapeutic strategies aimed at inhibiting tumor growth and other diseases associated with neovascularization.
Population of the Vessel Wall by Leukocytes Binding to P-Selectin in a Model of Disturbed Arterial Flow
Authors: Skilbeck Christopher , Westwood Susan M. , Walker Peter G. , David Tim , Nash Gerard B.
Published: September 2007

We examined the hypothesis that disturbance of laminar flow promotes the attachment of leukocytes to the vessel wall in regions where the wall shear stress is otherwise too high. Isolated neutrophils, lymphocytes, or monocytes were perfused through chambers with backward-facing steps so that vortices occurred with well-defined reattachment of flow. Wall shear stresses downstream in reestablished flow equaled 0.07 Pa (low shear) or 0.3 Pa (high shear). In chambers coated with P-selectin, adherent leukocytes rolled. By use of a P-selectin-Fc fragment chimera, adhesion was predominantly stationary, enabling definition of initial attachment sites. Neutrophils adhered in all regions of the low-shear chamber, with a local maximum around the reattachment point. However, in the high-shear chamber, adhesion was restricted to the recirculation zone and immediately downstream from the reattachment point. Rolling at high shear stress allowed a population of regions where initial attachment could not occur. At high shear, lymphocytes and monocytes also formed attachments restricted to the region of the reattachment point. The results imply that all types of leukocytes might bind to a capture receptor in high-shear vessels with discontinuities in the wall and might then spread to other regions.
Electron Paramagnetic Resonance Investigation on Modulatory Effect of 17β-Estradiol on Membrane Fluidity of Erythrocytes in Postmenopausal Women
Authors: Tsuda Kazushi , Kinoshita Yukiko , Kimura Keizo , Nishio Ichiro , Masuyama Yoshiaki
Published: September 2007

Many studies have shown that estrogen may exert cardioprotective effects and reduce the risk of hypertension and coronary events. On the other hand, it has been proposed that cell membrane abnormalities play a role in the pathophysiology of hypertension, although it is not clear whether estrogen would influence membrane function in essential hypertension. The present study was performed to investigate the effects of 17β-estradiol (E
2
) on membrane fluidity of erythrocytes in normotensive and hypertensive postmenopausal women. We determined the membrane fluidity of erythrocytes by means of an electron paramagnetic resonance and spin-labeling method. In an in vitro study, E
2
significantly decreased the order parameter for 5-nitroxide stearate and the peak height ratio for 16-nitroxide stearate obtained from electron paramagnetic resonance spectra of erythrocyte membranes in normotensive postmenopausal women. The finding indicates that E
2
might increase the membrane fluidity of erythrocytes. The effect of E
2
was significantly potentiated by the NO donor,
S
-nitroso-
N
-acetylpenicillamine, and a cGMP analogue, 8-bromo-cGMP. In contrast, the change in the membrane fluidity evoked by E
2
was attenuated in the presence of the NO synthase inhibitor,
N
G
-nitro-
l
-arginine methyl ester, and asymmetric dimethyl-
l
-arginine. In hypertensive postmenopausal women, the membrane fluidity of erythrocytes was significantly lower than that in normotensive postmenopausal women. The effect of E
2
on membrane fluidity was significantly more pronounced in the erythrocytes of hypertensive postmenopausal women than in the erythrocytes of normotensive postmenopausal women. The results of the present study showed that E
2
significantly increased the membrane fluidity and improved the microviscosity of erythrocyte membranes, partially mediated by an NO- and cGMP-dependent pathway. Furthermore, the greater action of E
2
in hypertension might be consistent with the hypothesis that E
2
could have a beneficial effect in regulating rheological behavior of erythrocytes and could have a crucial role in the improvement of the microcirculation in hypertension.
Involvement of Rho-Kinase and the Actin Filament Network in Angiotensin II–Induced Contraction and Extracellular Signal–Regulated Kinase Activity in Intact Rat Mesenteric Resistance Arteries
Authors: Matrougui K. , Tankó L.B. , Loufrani L. , Gorny D. , Levy B.I. , Tedgui A. , Henrion D.
Published: September 2007

We have previously shown that angiotensin II (Ang II) and pressure increase extracellular signal-regulated kinase (ERK) 1/2 activity synergistically in intact, pressurized resistance arteries in vitro. However, the mechanisms by which pressure and Ang II activate ERK1/2 in intact resistance arteries remain to be determined. The purpose of the present study was to investigate the involvement of Rho-kinase and the actin filament network in Ang II- and pressure-induced ERK1/2 activation, as well as in the contractile response induced by Ang II. Mesenteric resistance arteries (200 to 300 μm) were isolated, mounted in an arteriograph, and stimulated by pressure, Ang II, or both. Activation of ERK1/2 was then measured by an in-gel assay. In mesenteric resistance arteries maintained at 70 mm Hg, Ang II (0.1 μmol/L) induced contraction (29±1.4% of phenylephrine, 10 μmol/L-induced contraction) and significantly increased ERK1/2 activity. Selective inhibition of Rho-kinase by Y-27632 (10 μmol/L) and selective disruption of the actin filament network by cytochalasin B (10 μmol/L) both decreased the Ang II-induced contraction by 78±1.2% and 87±1.9%, respectively, and significantly diminished ERK1/2 activity. In the absence of Ang II, increasing intraluminal pressure from 0 to 70 or 120 mm Hg increased ERK1/2 activity. ERK1/2 activity at 120 mm Hg was similar to that observed at 70 mm Hg in the presence of Ang II. Pressure-induced ERK1/2 activation was markedly attenuated by cytochalasin B but not by Y-27632. These results indicate that whereas pressure-induced ERK1/2 activation requires an intact actin filament network, but not Rho-kinase, the activation of ERK1/2 and the contraction induced by Ang II require both Rho-kinase and an intact actin filament network in isolated, intact mesenteric resistance arteries.
Cardiovascular Risk Factors Associated With Insulin Resistance Cluster in Families With Early-Onset Coronary Heart Disease
Authors: Kareinen Anu , Viitanen Laura , Halonen Pirjo , Lehto Seppo , Laakso Markku
Published: September 2007

Coronary heart disease (CHD) is a multifactorial disease caused by environmental and genetic factors. CHD clusters in families, but it is not known whether susceptibility to early-onset CHD is associated with the clustering of cardiovascular risk factors. Therefore, we determined the levels of cardiovascular risk factors among siblings with and without severe early-onset CHD drawn from 101 Finnish families. Probands with CHD, compared with their siblings without CHD, had, respectively, higher 2-hour insulin levels (475.7 versus 331.8 pmol/L,
P
=0.011) and 2-hour insulin areas (796.2 versus 640.4 pmol/L per hour,
P
=0.031) in an oral glucose tolerance test, lower high density lipoprotein cholesterol levels (1.22 versus 1.42 mmol/L,
P
=0.001), higher total triglyceride levels (1.91 versus 1.68 mmol/L,
P
=0.018), higher very low density lipoprotein triglyceride levels (1.25 versus 1.06 mmol/L,
P
=0.011), and higher fibrinogen levels (3.8 versus 3.4 g/L,
P
= 0.008). No significant differences were found in cardiovascular risk factors between affected siblings and probands with CHD. Environmental or lifestyle factors did not differ between siblings with or without early-onset CHD. We conclude that cardiovascular risk factors associated with the insulin resistance syndrome (hyperinsulinemia, low high density lipoprotein cholesterol, high total and very low density lipoprotein triglycerides, and high fibrinogen) are likely to contribute indirectly to early-onset CHD.
Local Gene Transfer of Tissue Inhibitor of Metalloproteinase-2 Influences Vein Graft Remodeling in a Mouse Model
Authors: Hu Yanhua , Baker Andrew H. , Zou Yiping , Newby Andrew C. , Xu Qingbo
Published: September 2007

Recently, we established a new mouse model of vein graft arteriosclerosis by grafting vena cava to carotid arteries. In many respects, the morphological features of this murine vascular graft model resemble those of human venous bypass graft disease. Using this model, we studied the effects of local gene transfer of tissue inhibitor of metalloproteinase-2 (TIMP-2) on vein graft remodeling. Mouse isogeneic vessels of the vena caval veins were grafted end to end into carotid arteries, then enveloped with the replication-defective recombinant adenoviruses overexpressing human TIMP-2 (RAdTIMP-2) or β-galactosidase (RAdLacZ) at 1×10
10
plaque-forming units/mL in a total volume of 50 μL, and incubated at room temperature for 20 minutes. In the untreated group, vessel wall thickening was observed as early as 1 week after surgery and progressed to 4- to 10-fold the original thickness in grafted veins at 4 and 8 weeks, respectively. RAdLacZ vector treatment significantly enhanced neointimal lesions at 8 weeks, which was completely blocked by RAdTIMP-2 gene overexpression. Interestingly, RAdTIMP-2 gene transfer resulted in a reduction in vessel diameter of grafted veins compared with ungrafted veins (819±96 versus 624±67 μm, respectively;
P
<0.05). Maximal β-galactosidase activity was found at 2 weeks and was detectable until 4 weeks after gene transfer. Double immunofluorescence studies demonstrated that cells overexpressing TIMP-2 were mostly localized in the adventitia and were MAC-1-positive monocytes/macrophages but not smooth muscle cells. Furthermore, the activity of matrix metalloproteinases was markedly decreased in the vessel walls treated with RAdTIMP-2 compared with that in the untreated control group and the RAdLacZ-treated group. Thus, this mouse model has been proven to be useful in gene transfer studies. Our findings demonstrate that local TIMP-2 gene transfer significantly reduces vein graft diameter, ie, remodeling to an artery-like vessel via inhibition of matrix metalloproteinase activity.
Extracellular Superoxide Dismutase Deficiency and Atherosclerosis in Mice
Authors: Sentman Marie-Louise , Brännström Thomas , Westerlund Sanna , Laukkanen Mikko O. , Ylä-Herttuala Seppo , Basu Samar , Marklund Stefan L.
Published: September 2007

Lipoprotein peroxidation in the arterial wall has been implicated in atherogenesis. The superoxide radical is formed in arteries and can induce such oxidation. Extracellular superoxide dismutase (EC-SOD) occurs in high concentration in the vascular wall interstitium, and in this study, we examined the importance of the enzyme in atherogenesis. On an apolipoprotein E–null background, the limited aortic lesions induced by a 1-month atherogenic diet were larger in EC-SOD wild-type mice than in EC-SOD–null mice, whereas there were no differences between the EC-SOD genotypes in the larger lesions seen after 3 months on the diet or after 8 months on normal chow. Despite smaller or equal lesions in the EC-SOD–null mice, their cholesterol levels were somewhat higher. Also, on a wild-type background, there were no effects produced by the absence or presence of EC-SOD on atherogenic diet–induced aortic root lesions. The urinary excretion of the lipid peroxidation biomarker 8-isoprostaglandin F
2α
was related to the rates of atherogenesis in the mice but was not influenced by the EC-SOD genotype. Likewise, the EC-SOD status had no effect on the staining for oxidized low density lipoprotein epitopes in aortic root sections. Our findings suggest that EC-SOD has little influence on atherogenesis in mice.
Intravenous Magnesium in Experimental Stent Thrombosis in Swine
Authors: Rukshin Vladimir , Azarbal Babak , Shah Prediman K. , Tsang Vivian T. , Shechter Michael , Finkelstein Ariel , Cercek Bojan , Kaul Sanjay
Published: September 2007

We investigated the effects of magnesium on acute platelet-dependent stent thrombosis in an ex vivo porcine arteriovenous shunt model of high-shear blood flow. Control nitinol stents were expanded to 2 mm in diameter in a tubular perfusion chamber interposed in the shunt and exposed to flowing arterial blood at a shear rate of 2100 s
−1
for 20 minutes (n=156 perfusion runs in 10 swine). Animals were treated with intravenous heparin or MgSO
4
alone (2 g bolus over 20 minutes, followed by 2 g/h infusion) and combined heparin plus MgSO
4
in random fashion. Effects on thrombus weight (TW), platelet aggregation, bleeding time, activated clotting time, mean arterial blood pressure, and heart rate were quantified. Data points in the magnesium-treated animals were examined within 20 minutes after bolus (Mg-early) and >40 minutes after bolus (Mg-late). Stent TW (20±3 mg, pretreatment) was reduced by 42±21%, 47±19%, 48±16%, 67±12%, and 86±8% in the groups treated with Mg-early alone, Mg-late alone, heparin alone, heparin+Mg-early, and heparin+Mg-late, respectively (all
P
<0.001 versus pretreatment,
P
<0.001 for heparin+Mg-early and Mg-late versus heparin or magnesium alone, and
P
<0.05 for heparin+Mg-late versus heparin+Mg-early, ANOVA). Magnesium had no significant effect on platelet aggregation, activated clotting time, or bleeding time. There were no significant effects on heart rate or mean arterial blood pressure. The serum magnesium level was inversely correlated with TW (
r
=−0.70,
P
=0.002). In conclusion, treatment with intravenous MgSO
4
produced a time-dependent inhibition of acute stent thrombosis under high-shear flow conditions without any hemostatic or significant hemodynamic complications. Thus, magnesium may be an effective agent for preventing stent thrombosis.
Protection From Obesity in Mice Lacking the VLDL Receptor
Authors: Goudriaan Jeltje R. , Tacken Paul J. , Dahlmans Vivian E.H. , Gijbels Marion J.J. , van Dijk Ko Willems , Havekes Louis M. , Jong Miek C.
Published: September 2007

It has previously been reported that mice lacking the VLDL receptor (VLDLR−/−) exhibit normal plasma lipid levels and a modest decrease in adipose tissue mass. In the present study, the effect of VLDLR deficiency on profound weight gain was studied in mice. Obesity was induced either by feeding of a high-fat, high-calorie (HFC) diet or by crossbreeding mice onto the genetically obese
ob
/
ob
background. After 17 weeks of HFC feeding, VLDLR−/− mice remained lean, whereas their wild-type littermates (VLDLR+/+) became obese. Similarly, the weight gain of
ob
/
ob
mice was less profound in the absence of the VLDLR. Moreover, VLDLR deficiency led to increased plasma triglycerides after HFC feeding. The protection from obesity in VLDLR−/− mice involved decreased peripheral uptake of fatty acids, because VLDLR−/− mice exhibited a significant reduction in whole-body free fatty acid uptake, with no clear differences in food intake and fat absorption. These observations were supported by a strong decrease in average adipocyte size in VLDLR−/− mice of both obesity models, implying reduced adipocyte triglyceride storage in the absence of the VLDLR. These results suggest that the VLDLR plays a role in the delivery of VLDL-derived fatty acids into adipose tissue.
Relation of Blood Pressure Variability to Carotid Atherosclerosis and Carotid Artery and Left Ventricular Hypertrophy
Authors: Roman Mary J. , Pickering Thomas G. , Schwartz Joseph E. , Pini Riccardo , Devereux Richard B.
Published: September 2007

The relationship of blood pressure (BP) variability to cardiovascular target-organ damage is controversial. Studies examining BP variability and left ventricular (LV) hypertrophy have been contradictory, and only limited data on the relation of BP variability to carotid atherosclerosis and carotid artery hypertrophy exist. BP variability was assessed as the standard deviation and coefficient of variation of awake and asleep pressures in 511 normotensive or untreated hypertensive subjects who underwent ambulatory BP monitoring and cardiac and carotid ultrasonography. Although the presence of focal carotid plaque was associated with an increase in ambulatory pressures and pressure variability, the differences in variability were eliminated by adjustment for age and absolute pressures. Similarly, LV mass was significantly related to BP variability, but the significance of this finding was eliminated after adjustment for important covariates. In multivariate analyses, age was the primary determinant of carotid artery cross-sectional area, with a weak but independent contribution from awake systolic and diastolic BP variability in addition to absolute pressure. BP variability was not independently related to either carotid or LV relative wall thickness, both measures of concentric remodeling. In the present study, awake BP variability was weakly but independently associated with carotid artery cross-sectional area, a measure of arterial hypertrophy. However, neither systolic nor diastolic BP variability was independently associated with carotid atherosclerotic plaque or LV mass.
Coronary Artery Complicated Lesion Area Is Related to Functional Polymorphism of
Matrix Metalloproteinase 9
Gene
Authors: Pöllänen Perttu J. , Karhunen Pekka J. , Mikkelsson Jussi , Laippala Pekka , Perola Markus , Penttilä Antti , Mattila Kari M. , Koivula Timo , Lehtimäki Terho
Published: September 2007

Matrix metalloproteinase 9 (MMP9) is expressed in human atherosclerotic plaques, and the protein is localized in human coronary atherosclerotic lesions. The
MMP9
gene has a C-to-T promoter polymorphism at position −1562, which affects transcription and leads to promoter low-activity (C/C) and high-activity (C/T, T/T) genotypes. To determine whether these genotypes exert an influence on the atherosclerotic lesion area, we investigated their association with different types of coronary lesions in an autopsy cohort of 276 men aged 33 to 69 years. Areas of the coronary wall covered with fatty streaks and fibrotic, calcified, and complicated lesions were measured, and the percentage of coronary narrowing was determined.
MMP9
genotypes were determined by polymerase chain reaction and restriction enzyme digestion. In men aged ≥53 years, the mean area of complicated lesions in 3 coronaries was significantly associated with the
MMP9
genotype (
P
=0.008). Subjects with high promoter activity genotypes had, on average, larger complicated lesion areas than did those with the low-activity genotype. The
MMP9
genotype persisted as an independent predictor of complicated lesion area after adjustment for age, body mass index, hypertension, diabetes, and smoking (
P
=0.012). These data provide evidence that the proposed effect of MMP9 in the process of atherosclerotic lesion development may be modified by the
MMP9
genotype.
Adenovirus-Mediated Overexpression of Dominant-Negative Mutant of c-Jun Prevents Intercellular Adhesion Molecule-1 Induction by LDL
Authors: Wang Nanping , Verna Lynne , Liao Hai-ling , Ballard Alex , Zhu Yi , Stemerman Michael B.
Published: September 2007

Low density lipoprotein (LDL) induces intercellular adhesion molecule-1 (ICAM-1) gene expression and leads to endothelial cell (EC) leukocyte adhesion. However, the transcriptional mechanism for LDL-induced EC perturbation remains to be fully explained. Activator protein-1 (AP-1) is induced after the exposure of ECs to LDL. In the present study, a regulated adenovirus expressing a dominant-negative mutant of c-Jun (TAM-67) was used to examine the role of AP-1 in the LDL-induced ICAM-1 activation. Overexpression of TAM-67 specifically inhibited AP-1 activation and prevented the LDL-activated surface expression of ICAM-1 protein in human umbilical vein ECs and human coronary artery ECs. Northern analyses and promoter transactivation assays indicated that this effect of TAM-67 was likely mediated through a suppression of the transcriptional regulation of the ICAM-1 gene. Functionally, TAM-67 attenuated leukocyte adherence to ECs in response to LDL. Furthermore, electrophoresis mobility shift assays and site-directed mutagenesis suggested that an AP-1–like motif in the promoter region of the human ICAM-1 gene was a critical
cis
element for LDL induction. These results, for the first time, provide evidence suggesting that AP-1 is a major regulatory mechanism leading to endothelial activation.
Characterization of Polymorphic Structure of Cathepsin G Gene
Authors: Herrmann Stefan-Martin , Funke-Kaiser Heiko , Schmidt-Petersen Klaus , Nicaud Viviane , Gautier-Bertrand Marion , Evans Alun , Kee Frank , Arveiler Dominique , Morrison Caroline , Orzechowski Hans-Dieter , Elbaz Alexis , Amarenco Pierre , Cambien François , Paul Martin
Published: September 2007

Cathepsin G (CTSG), a serine protease released from activated neutrophils, may cause platelet activation, leading to intravascular thrombosis, thus contributing to cardiovascular and cerebrovascular disease. Applying the candidate gene approach, we screened the 5′-flanking region and the entire coding region of the CTSG gene for genetic variation by using polymerase chain reaction/single-strand conformation polymorphism analysis from 96 patients at high risk for myocardial infarction (MI). We identified 4 polymorphisms in the 5′-flanking region (G-618C, G-315A, C-179T, and C-160T) and 1 polymorphism in the coding region (Asn125Ser) of the gene and genotyped the participants in the Etude Cas-Temoins sur l’Infarctus du Myocarde (ECTIM Study), a case-control study for MI, and in the Étude du Profil Génétique de l’Infarctus Cérébral (GENIC Study), a case-control study for brain infarction (BI), for all identified genetic variants. The potential in vitro functionality of the 4 variants in the 5′-flanking region was investigated with transient transfection analyses in U937 cells with different allelic promoter constructs by using a luciferase assay. Our in vitro analyses did not reveal any differences for the investigated allelic constructs with respect to promoter activity, and none of the polymorphisms in the 5′-flanking region was associated with the available phenotypes in either study. Allele and genotype distributions of all identified polymorphisms did not globally differ between cases and controls in the ECTIM Study. However, in patients from the ECTIM Study, the Ser125 allele was significantly associated with elevated plasma fibrinogen levels (
P
=0.006), but this effect was not seen in controls (case-control heterogeneity,
P
=0.04). There was a significant interaction between CTSG Asn125Ser and the β-fibrinogen gene polymorphism G-455A on plasma fibrinogen levels (
P
=0.04). In the GENIC Study, the odds ratio for BI associated with CTSG Ser125 carrying was 1.82 (95% CI 1.16 to 2.84,
P
=0.008) in patients without a history of cardiovascular or cerebrovascular diseases. Our results indicate that the CTSG Ser125 allele is associated with plasma fibrinogen levels in MI patients from the ECTIM Study and with BI in the GENIC Study. Further studies should be carried out to define the underlying mechanisms.
Overexpression of Allograft Inflammatory Factor-1 Promotes Proliferation of Vascular Smooth Muscle Cells by Cell Cycle Deregulation
Authors: Autieri Michael V. , Carbone Christopher M.
Published: September 2007

Allograft inflammatory factor-1 (AIF-1) is not present in normal arteries, but its expression is induced in vascular smooth muscle cells (VSMCs) in several models of arterial injury. The proliferation of VSMCs is a major component of neointimal hyperplasia in many arteriopathies, and the purpose of this study was to determine the role of AIF-1 in the growth of VSMCs. Transfection and constitutive expression of AIF-1 in a primary and a rat VSMC line results in enhanced growth of those cells as measured by cell number and is proportional to the amount of AIF-1 expressed. Constitutive expression of AIF-1 results in a shorter cell cycle, as measured by flow cytometry, and aberrant expression of cell cycle proteins, as determined by Western blot. AIF-1 overexpression also permits growth of these cells in serum-reduced media. Collectively, these data suggest that AIF-1 may participate in the progression of vascular proliferative disease on the basis of its ability to regulate the growth of VSMCs.
Formulation and Delivery Mode Affect Disposition and Activity of Tyrphostin-Loaded Nanoparticles in the Rat Carotid Model
Authors: Fishbein Ilia , Chorny Michael , Banai Shmuel , Levitzki Alexander , Danenberg Haim D. , Gao Jianchuan , Chen Xing , Moerman Evgeny , Gati Irith , Goldwasser Victoria , Golomb Gershon
Published: September 2007

Poor drug residence in the arterial wall hinders clinical implementation of local drug delivery strategies for the treatment of restenosis. A rat carotid model of vascular injury and intraluminal delivery of tyrphostin-containing polylactic acid (PLA) nanoparticles (NPs) were used to determine the relationship between residence properties and biological activity of different formulations and administration modes. The effects of delivery modes (denudation and delivery time) and formulation variables (adsorbed vs encapsulated drug, and NP size) on arterial drug/NP retention were examined. Antirestenotic effects of large (160 nm) and small (90 nm) tyrphostin-containing NPs, surface-absorbed tyrphostin, and systemic treatment were compared. Fluorescent NPs were used to study the spatial distribution of the carrier in the arterial wall. The decrease in arterial tyrphostin level over time fitted a biexponential model. Delivery time and pressure, endothelium integrity, particle size, and drug-polymer association affected local pharmacokinetics and the antirestenotic results after 14 days. The PLA-based tyrphostin NP formulation ensured a prolonged drug residence at the angioplasty site after single intraluminal application. Several readily adjustable formulation and procedural factors considerably modified arterial ingress of the drug-loaded NPs and governed their subsequent redistribution, tissue binding, elimination, and ensuing antirestenotic effect.
Persistence of Atherosclerotic Plaque but Reduced Aneurysm Formation in Mice With Stromelysin-1 (MMP-3) Gene Inactivation
Authors: Silence J. , Lupu F. , Collen D. , Lijnen H.R.
Published: September 2007

To investigate a potential role for stromelysin-1 (MMP-3) in the development and progression of atherosclerotic lesions and aneurysm formation, mice with a deficiency of apolipoprotein E (ApoE
−/−
:MMP-3
+/+)
) or with a combined deficiency of apoE and MMP-3 (ApoE
−/−
:MMP-3
−/−
) were kept on a cholesterol-rich diet for 30 weeks. Atherosclerotic lesions throughout the thoracic aorta were significantly larger in ApoE
−/−
:MMP-3
−/−
than in ApoE
−/−
:MMP-3
+/+
mice (
P
<0.05) and contained more fibrillar collagen (
P
<0.01). Aneurysms in the thoracic and abdominal aortas were less frequent in ApoE
−/−
:MMP-3
−/−
than in ApoE
−/−
:MMP-3
+/+
mice (8.5±1.7% vs 14±2.1% of sections, mean±SD,
P
<0.01). Immunocytochemistry revealed enhanced accumulation of macrophages in atherosclerotic lesions of ApoE
−/−
:MMP-3
+/+
mice (
P
<0.01) and expression of urokinase-type plasminogen activator (u-PA) and MMP-3 colocalizing with macrophages. Zymography confirmed the presence of u-PA and MMP-3 activity in extracts of atherosclerotic aortas. These data suggest that plasmin, generated by macrophage-secreted u-PA, activates pro-MMP-3 produced by accumulated macrophages. MMP-3 activity may then contribute to a reduction of plaque size, possibly by degradation of matrix components, and promote aneurysm formation by degradation of the elastica lamina.
Niacin, but Not Gemfibrozil, Selectively Increases LP-AI, a Cardioprotective Subfraction of HDL, in Patients With Low HDL Cholesterol
Authors: Sakai Takaaki , Kamanna Vaijinath S. , Kashyap Moti L.
Published: September 2007

Evidence indicates that the high density lipoprotein (HDL) subfraction containing apolipoprotein A-I without apolipoprotein AII (LP-AI) is more antiatherogenic than HDL particles containing apolipoprotein A-I and apolipoprotein A-II (LP-AI+AII). This study examined the effect of extended-release niacin (niacin-ER) and gemfibrozil on LP-AI and LP-AI+AII particles in patients with low levels of HDL cholesterol (HDL-C). Mechanisms by which these agents modulate HDL particles were investigated by in vitro studies using human hepatoblastoma (Hep G2) cells. A total of 139 patients with low HDL-C (≤40 mg/dL) were randomized to niacin-ER or gemfibrozil in a multicenter double-blind trial. Patients were dose-escalated with once-nightly niacin-ER (1 to 2 g) or gemfibrozil (1.2 g) for 19 weeks. Niacin-ER had a greater effect in raising HDL-C and apolipoprotein A-I levels than did gemfibrozil. Niacin-ER at 1- and 2-g doses increased LP-AI levels by 8.7±4.0% (
P
=0.033) and 24.0±4.4% (
P
<0.001), respectively. Gemfibrozil had no consistent effect on LP-AI levels. LP-AI+AII levels increased 5% to 8% by both agents. In vitro studies showed that niacin, but not gemfibrozil, selectively decreased the uptake of
125
I-labeled LP-AI holoparticles by Hep G2 cells. The uptake of [
3
H]cholesterol ester was ≈75% greater from LP-AI versus LP-AI+AII particles, but neither niacin nor gemfibrozil affected cholesterol ester uptake. These data indicate that unlike gemfibrozil, niacin selectively increases LP-AI compared with LP-AI+AII particle concentration in patients with low HDL-C levels. The mechanism of action of increased LP-AI concentration appears to be mediated by decreased hepatic removal of LP-AI particles, which are more efficient in reverse cholesterol transport, thus suggesting an additional mechanism by which niacin mediates its antiatherogenic properties.
Adhesion of Monocytes to Arterial Endothelium and Initiation of Atherosclerosis Are Critically Dependent on Vascular Cell Adhesion Molecule-1 Gene Dosage
Authors: Dansky Hayes M. , Barlow Courtenay B. , Lominska Chris , Sikes John L. , Kao Catherine , Weinsaft Jonathan , Cybulsky Myron I. , Smith Jonathan D.
Published: September 2007

Vascular cell adhesion molecule-1 (VCAM-1/
Vcam1
) is a cytokine-inducible member of the immunoglobulin gene superfamily that is expressed by arterial endothelial cells in regions predisposed to atherosclerosis and at borders of atherosclerotic plaques. To determine whether VCAM-1 expression regulates atherosclerotic lesion formation, we crossed
Vcam1
domain 4–deficient (D4D) mice, which partially circumvent the embryonic lethality of
Vcam1
null mice, with apolipoprotein E null (
Apoe
−/−
) mice, which spontaneously develop hypercholesterolemia and atherosclerosis. In the
Apoe
−/−
background, mice homozygous for the
Vcam1
D4D allele had markedly reduced arterial VCAM-1 expression, monocyte adherence in the aortic root, and fatty streak formation. Heterozygous
Vcam1
D4D mice revealed a
Vcam1
gene-dosage effect and had intermediate, yet significant, reductions in these parameters. Our data demonstrate that VCAM-1 plays a pivotal role in the initiation of atherosclerosis in
Apoe
−/−
mice.
Insulin Inhibits the Maturation Phase of VLDL Assembly via a Phosphoinositide 3-Kinase—Mediated Event
Authors: Brown Anna-Marie , Gibbons Geoffrey F.
Published: September 2007

LY 294002 (80 μmol/L), an inhibitor of phosphoinositide 3-kinase, was used to investigate the involvement of this enzyme in the insulin-mediated regulation of very low density lipoprotein (VLDL) apolipoprotein B (apoB) output from cultured rat hepatocytes. Newly synthesized apoB was pulse-labeled with [
35
S]methionine and was then allowed to assemble, via an intermediate precursor stage, into mature VLDL during subsequent chase periods. Brefeldin A (BFA, 0.2 μg/mL) was used to discriminate between the role of insulin in the regulation of the early, compared with the later, events of VLDL assembly, including apoB degradation. Insulin (78 nmol/L), when present during the pulse-labeling and subsequent chase periods, inhibited the secretion of apoB-100 and apoB-48 as VLDL by 53% and 56%, respectively. Degradation of both was concomitantly increased. Secretion of high density lipoprotein apoB, derived from VLDL precursors, was relatively unaffected under these conditions, as was the net synthesis of apoB-100 and apoB-48. The presence of BFA during the pulse-labeling period and subsequent chase period prevented the maturation of VLDL in the insulin-treated and the non–insulin-treated cells. BFA was then removed, allowing the maturation of VLDL to proceed. Removal of insulin at this stage reversed the overall inhibitory effect of insulin. Furthermore, when insulin remained present during this period, the simultaneous presence of LY 294002 also reversed the inhibitory effect of insulin on VLDL apoB output and abolished the increase in apoB degradation. The results suggest that insulin signaling via phosphoinositide 3-kinase inhibited the maturation phase of VLDL assembly by preventing bulk lipid transfer to a VLDL precursor, thus enhancing the degradation of apoB. There was no inhibition of the conversion of newly synthesized apoB into the VLDL precursor form.
Plasma Fibrin D-Dimer Levels and Risk of Stable Coronary Artery Disease
Authors: Koenig Wolfgang , Rothenbacher Dietrich , Hoffmeister Albrecht , Griesshammer Martin , Brenner Hermann
Published: September 2007

Abstract—

Increased levels of fibrin D-dimer are indicative of a hypercoagulable state, as found in acute coronary syndromes. Few well-controlled studies have assessed D-dimers in patients with stable coronary artery disease (CAD). We measured levels of D-dimers (in ng/mL by enzyme-linked immunosorbent assay) in 312 patients with angiographically proved CAD and stable angina pectoris and in 477 age- and sex-matched healthy blood donors. Demographic characteristics were assessed by a standardized questionnaire, and a complete lipid profile was performed for all subjects. In addition, a variety of other markers of hemostasis and inflammation were measured. The distribution of D-dimer levels was skewed to the right, and plasma median levels were higher in cases than in controls (median: 11.2 vs 2.8 ng/mL;
P
<0.001). In controls, correlations of D-dimer were found with fibrinogen, plasma viscosity, and interleukin-6. In logistic regression analysis, the age- and sex-adjusted odds ratio (OR) for the presence of CAD was 2.6 (95% confidence interval [CI], 1.9 to 3.5) when the highest quartile of the D-dimer distribution was compared with the combined lower 3 quartiles. The OR did not change appreciably after controlling for nonlipid risk factors (OR, 2.7; 95% CI, 1.9 to 3.9) and remained significant after further adjustment for other hemostatic parameters (OR, 2.4; 95% CI, 1.7 to 3.3) and markers of inflammation (OR, 2.1; 95% CI, 1.5 to 2.9). Plasma D-dimer levels are strongly and independently associated with the presence of CAD in patients with stable angina pectoris. These results support the concept of a contribution of intravascular fibrin to atherothrombogenesis.