bluuebunny/crossref_metadata_2025 huggingface dataset:
Patients With Early-Onset Peripheral Vascular Disease Have Increased Levels of Autoantibodies Against Oxidized LDL
Authors: Bergmark C. , Wu R. , Faire U. de , Lefvert A. K. , Swedenborg J.
Published: June 2011

Abstract
Oxidative modification of LDL has been proposed as an early and crucial step in the development of atherosclerosis, and antibodies against such modified LDL are found in both healthy individuals and patients with atherosclerosis. In this study, 62 patients who were surgically treated for peripheral arterial occlusive disease below the age of 50 were investigated and compared with age- and sex-matched healthy individuals in a case-control study. Autoantibodies against oxidized LDL were measured with an enzyme-linked immunosorbent assay method. Risk factors such as smoking, hypertension, family history of premature cardiovascular events, and lipoprotein levels were also determined. The patients had significantly higher levels of autoantibodies against oxidized LDL; significantly higher levels of total cholesterol, LDL cholesterol, triglycerides, and apo A-I; and significantly lower levels of HDL cholesterol than did control subjects. In multivariate analyses autoantibodies against oxidized LDL discriminated better between patients and control subjects than did any of the different lipoprotein analyses. Among patients, the presence of hypertension and a family history of cardiovascular events were the only factors significantly associated with increased levels of autoantibodies against oxidized LDL.
The Enhanced Association of Apolipoprotein E With Apolipoprotein B–Containing Lipoproteins in Serum-Stimulated Hepatocytes Occurs Intracellularly
Authors: Fazio Sergio , Yao Zemin
Published: June 2011

Abstract
Synthesis and secretion of VLDL in HepG2 cells are stimulated by several lipogenic factors, including serum. We previously found that the amount of apolipoprotein (apo) E associated with large lipoproteins such as VLDL increased under conditions of stimulated lipogenesis. The present study was designed to determine if the increased apoE association with VLDL occurs intracellularly or after secretion. In addition to HepG2, we studied rat hepatoma McA-RH7777 cells for production of endogenous rat apoE and transfected human apoE3. In both hepatoma cell lines stimulation of lipogenesis and production of large apoB-containing lipoproteins by serum resulted in increased apoE association with these particles and in decreased apoE association with HDL without affecting the total apoE output. Although evidence of apoE redistribution was seen among lipoproteins in the media, the apoE newly secreted under conditions of stimulated lipogenesis mainly associated with apoB-containing lipoproteins at the expense of its association with HDL. However, this effect was not attributable to reduced HDL lipid and apoA-I mass. Finally, redistribution of apoE from HDL to apoB-containing lipoproteins was also observed intracellularly in both HepG2 and transfected McA-RH7777 cells expressing human apoE3. These data suggest that the redistribution of apoE from HDL to apoB-containing lipoproteins upon activated lipogenesis in hepatoma cells occurs intracellularly and is not attributable to a decrease in HDL production.
Whole-Blood Platelet Aggregation Predicts In Vitro and In Vivo Primary Hemostatic Function in the Elderly
Authors: Emery Jonathan D. , Leifer David W. , Moura Glaci L. , Southern Patricia , Morrissey James H. , Lawrence Jeffry B.
Published: June 2011

AbstractIncreased platelet aggregation is associated with higher coronary artery disease mortality. Enhanced platelet aggregation in platelet-rich plasma has also been described in the elderly. To define age-related changes in primary hemostasis, we studied 37 elderly and 31 young blood donors. There were no significant age-related differences in whole-blood platelet aggregation, platelet adherence and thrombus formation on human umbilical artery segments, or bleeding time. Plasma fibrinogen was significantly higher in elderly men and women, whereas activated factor VII was elevated only in elderly women. Collagen-induced platelet aggregation was significantly correlated with platelet adherence to the subendothelium in elderly (r=.488,P=.002) but not in young donors. Accordingly, collagen-induced platelet aggregation showed a significant inverse correlation with bleeding time only in the elderly (r=−.401,P=.014). Arachidonic acid–induced platelet aggregation was significantly associated with platelet adherence to the subendothelium (r=.658,P=.003) and bleeding time (r=−.540,P=.021) only in elderly men. In young donors, ADP-induced platelet aggregation was significantly correlated with platelet adherence to the thrombogenic adventitial surface (r=.395,P=.031); in the elderly this association only approached significance (r=.315,P=.058). Whole-blood platelet aggregation in response to collagen and arachidonic acid may be more useful in predicting primary hemostatic function in the elderly than in the young. Furthermore, in the elderly, the correlation between platelet aggregation in whole blood and platelet–arterial wall interactions in vitro and in vivo may contribute to the ability of this test to predict coronary risk.
Coronary Heart Disease Risk Factors in Women With Polycystic Ovary Syndrome
Authors: Talbott Evelyn , Guzick David , Clerici Annette , Berga Sarah , Detre Katherine , Weimer Karl , Kuller Lewis
Published: June 2011

Abstract
The goal of the study was to compare cardiovascular heart disease risk factors in women with polycystic ovary syndrome (PCOS) and matched control subjects. Women with PCOS have risk factors, including anovulation, hyperandrogenism, and insulin resistance, that suggest a male coronary heart disease risk-factor profile. A total of 206 women with PCOS were recruited by using records from a large reproductive endocrinology practice. A clinical diagnosis of PCOS was made if there was a history of chronic anovulation in association with either clinical evidence of androgen excess (hirsutism) or if total testosterone level was >2 nm/L or the luteinizing hormone/follicle-stimulating hormone ratio was greater than 2. The overall response rate for cases was 76%. A control population was obtained by using a combination of area voters’ registration tapes and directories of households. A control subject was matched to each case subject by age±5 years, race, and neighborhood. The response rate for recruitment of the first or second eligible control subject was 83.6%. The average age at initial interview was 35.9±7.4 years for case and 37.2±7.8 years for control subjects. Women with PCOS had significantly increased cardiovascular disease risk factors compared with control women. These included increases in body mass index, insulin, and triglyceride levels (
P
<.001), decreased total HDL and HDL
2
levels (
P
<.01), and increased total cholesterol and fasting LDL levels, waist/hip ratio, and systolic blood pressure (
P
<.05). After controlling for age, body mass index, and other confounding variables, differences in total cholesterol, total HDL, HDL
2
, LDL cholesterol, and triglycerides were still significant between case and control subjects. These risk factors were especially elevated in PCOS women in the early premenopausal years compared with control women, indicating that women with PCOS should be monitored for early detection and considered for appropriate clinical interventions.
Antithrombotic Effects of Activated Protein C and Protein S in a Rabbit Model of Microarterial Thrombosis
Authors: Arnljots Björn , Dahlbäck Björn
Published: June 2011

Abstract
The antithrombotic properties of activated protein C (APC) and protein S were investigated in a rabbit model of microarterial thrombosis. The study focused on the ability of intact and thrombin-cleaved bovine protein S to potentiate the biological effects of bovine APC in vivo. Segments of the central arteries of the ears were subjected to arteriotomy, deep-vessel wall trauma, and arteriotomy suture. Five minutes before vascular reperfusion, groups of rabbits were infused with boluses of 0.1 mg/kg bovine APC alone or combined with different doses of intact (0.5, 0.1, or 0.05 mg/kg) or thrombin-cleaved (0.5 mg/kg) protein S. APC in combination with the two higher doses of protein S produced a potent antithrombotic response, as judged by assessment of vessel patency rates, while only the group receiving APC+0.5 mg/kg protein S showed significant reduction of thrombus weights as well. The biological effect depended on the active site in APC, as the antithrombotic effect was lost on pretreatment of APC with the serine protease inhibitor
d
-phenylalanyl-
l
-prolyl-
l
-arginine chloromethylketone. The potentiation of the APC response by protein S depended on the structural integrity of the protein, and cleavage of the thrombin-sensitive region in protein S by thrombin resulted in a loss of biological response. No hemorrhagic side effects were noted by the APC–protein S combination, and the anticoagulant response was mild, even to the highest doses of APC and protein S. The study demonstrates the specificity of the APC–protein S interaction in vivo and shows both the necessity of an active catalytic site in APC and the physiological importance of the APC–protein S interaction. APC appears to be a suitable antithrombotic agent when fully potentiated by its cofactor protein S, as it produces only mild plasma anticoagulation and no hemorrhagic side effects.
A Leukocyte Type of 12-Lipoxygenase Is Expressed in Human Vascular and Mononuclear Cells
Authors: Kim Jeong A. , Gu Jia-Li , Natarajan Rama , Berliner Judith A. , Nadler Jerry L.
Published: June 2011

Abstract
The lipoxygenase (LO) pathway has been implicated in leading to accelerated atherosclerosis. However, the precise type of LO present in unstimulated human aortic smooth muscle cells (HSMC), endothelial cells (HAEC), and monocytes (MO) is not clear. In this study, we used a specific reverse-transcriptase polymerase chain reaction (RT-PCR) method to analyze the type of LO mRNA expressed in normal HSMC, HAEC, and MO. In all three cell types, a 333-base-pair band was seen when primers and probes specific for the leukocyte type of 12-LO were used, suggesting that a leukocyte type of 12-LO is expressed in these cell types. Western immunoblotting analysis in cultured HSMC, HAEC, and MO using a polyclonal peptide antibody to the leukocyte type of 12-LO showed a specific 72-kD band that is identical to the molecular weight of the leukocyte type of 12-LO. These results indicate that a leukocyte type of 12-LO RNA and protein are expressed in HSMC, HAEC, and MO. Further, angiotensin II upregulates 12-LO activity and expression in HSMC, supporting a role for this 12-LO pathway in human vascular disease.
Renal and Systemic Transvascular Albumin Leakage in Severe Atherosclerosis
Authors: Jensen Jan Skov
Published: June 2011

Abstract
Microalbuminuria was recently proposed as a novel atherogenic risk factor. The pathophysiological link between microalbuminuria and atherosclerosis may be mediated through an increased generalized transvascular leakage of albumin. To investigate this hypothesis, urinary albumin excretion and clearance and systemic transvascular albumin leakage (TER
alb
) were measured in 23 patients with severe clinical atherosclerosis and 25 healthy controls. In addition, renal clearances of three other endogenous plasma proteins (IgG, IgG
4
, and β
2
-microglobulin) and of creatinine were measured. Measurements of urine and serum proteins were done by enzyme-linked immunosorbent assays. TER
alb
was measured by the fractional disappearance rate of
125
I-albumin from the total intravascular compartment in 1 hour after intravenous injection. Glomerular filtration rate was estimated as creatinine clearance. Urinary albumin excretion (geometric means [95% confidence intervals], 10.5 [6.1 to 18.3] versus 5.7 [4.7 to 6.9] μg/min;
P
<.05), fractional urinary albumin clearance (2.8 [1.6 to 4.8] ×10
−6
versus 1.3 [1.0 to 1.6] ×10
−6
;
P
<.05), and TER
alb
(6.0 [5.5 to 6.5] versus 5.1 [4.5 to 5.8] %/h;
P
<.05) were higher in patients than in control subjects. Glomerular charge selectivity (ratio of IgG clearance to IgG
4
clearance) was lower in patients than in control subjects (1.5 [1.1 to 2.0] versus 2.3 [2.0 to 2.6];
P
<.05). These alterations were independent of blood pressure, glomerular filtration rate, tubular function, and smoking status. It is concluded that atherosclerotic vascular disease is associated with renal and systemic transvascular leakiness for albumin. Theoretically, such leakiness may in addition allow for an increased lipid insudation into the large vessel wall, thereby linking microalbuminuria to atherogenesis. The lower glomerular charge selectivity is suggestive of decreased concentration of anionic components in the vessel walls.
Longitudinal Study of Associations of Microalbuminuria With the Insulin Resistance Syndrome and Sodium-Lithium Countertransport in Nondiabetic Subjects
Authors: Foyle Wendy-Jane , Carstensen Elin , Fernández Maryam C. , Yudkin John S.
Published: June 2011

Abstract
Microalbuminuria in diabetic patients is associated with ischemic heart disease and insulin resistance. We previously found a 9% prevalence of microalbuminuria in a nondiabetic population that we have reassessed, investigating associations of microalbuminuria with hypertension, dyslipidemia, hyperinsulinemia, and sodium-lithium countertransport. Of 125 subjects reexamined, 42 had been microalbuminuric 3 years previously. Twelve of these (29%) were microalbuminuric on at least one sample at follow-up, and 30 (76%) were normoalbuminuric on two. Of the 79 previously normoalbuminuric subjects, 12 (15%) became microalbuminuric on one sample, while 67 (85%) remained normoalbuminuric. Subjects who were microalbuminuric at both screening and recall were older (mean±SD, 65.9±11 versus 59.1±10.2 years,
P
=.04), with a higher waist-to-hip ratio (0.93±0.09 versus 0.86±0.08,
P
=.008) and at recall, on univariate analysis, had higher specific insulin (44.2 [range, 16.9 to 157.0] versus 28.4 [7.4 to 129.0] pmol/L,
P
=.005), intact proinsulin (5.1 [1.5 to 11.0] versus 3.0 [0.8 to 14.6] pmol/L,
P
=.003), and des-31,32-proinsulin (5.0 [0.5 to 9.8] versus 1.0 [0.5 to 12.2] pmol/L,
P
=.004) concentrations. There was also a significant difference in des-31,32-proinsulin concentration, after adjustment for covariates (
P
=.013), between subjects classified either as microalbuminuric or as normoalbuminuric at screening. There was no difference in body mass index; fasting blood glucose; systolic or diastolic blood pressure; total, HDL, or LDL cholesterol; triglycerides; plasminogen activator inhibitor-1; or sodium-lithium countertransport activity between consistently normoalbuminuric and persistently microalbuminuric subjects. We found a positive relationship of changes in albumin excretion rate with those in HDL cholesterol concentrations over the follow-up period (
r
=.25,
P
=.009) but none with changes in fasting blood glucose, blood pressure, other lipids, insulin, or proinsulin-like molecules. In conclusion, microalbuminuria is an unstable phenomenon over a period of 3 years in nondiabetic subjects, with a coefficient of variation of ≈60% on two paired samples over this time. It is associated with increased concentrations of insulin and of proinsulin-like molecules but not with other features of the insulin resistance syndrome.
HMG-CoA Reductase Inhibitors Reduce Acetyl LDL Endocytosis in Mouse Peritoneal Macrophages
Authors: Bernini Franco , Scurati Nicoletta , Bonfadini Giuliana , Fumagalli Remo
Published: June 2011

Abstract
We previously reported that mevalonate starvation elicited by hydroxymethyl glutaryl coenzyme A (HMG-CoA) reductase inhibitors reduced cholesterol accumulation promoted in murine macrophages by acetylated LDL (AcLDL). In the present study we investigated the cellular mechanism of this effect. Our results indicate that the HMG-CoA reductase inhibitors fluvastatin and simvastatin reduce, in a concentration-dependent manner, more then 50% of the
125
I-AcLDL degradation by macrophages. This effect was not due to a decrease of lysosomal enzyme activity, and it was paralleled by the retention of AcLDL-associated cholesteryl ester in the incubation medium. The ability of fluvastatin to inhibit AcLDL degradation was completely overcome by mevalonate and its derivative geranylgeraniol. Evaluation at 4°C of
125
I-AcLDL binding to plasma membrane suggested that the inhibitory effect of fluvastatin on lipoprotein catabolism was not due to a decreased expression of scavenger receptors. Fluorescent microscope analysis of cellular internalization of AcLDL labeled with the fluorochrome 3,3′-dioctadecyl indocarbocyanine demonstrated that fluvastatin inhibits lipoprotein endocytosis, an effect reversed by mevalonate. Studies performed with native
125
I-LDL indicated that fluvastatin did not inhibit but rather increased the degradation of LDL taken up by the normal LDL receptor. These results exclude a generalized depression of the cellular endocytotic activity by the drug. The ability of fluvastatin to reduce AcLDL catabolism and cholesterol esterification was more pronounced in cholesterol-enriched macrophages compared with normal cells. In conclusion, the present results demonstrate that HMG-CoA reductase inhibitors may reduce the in vitro cholesterol accumulation in macrophages by inhibiting AcLDL endocytosis.
Activation of Phosphatidylinositol-Specific Phospholipase C in Response to HDL
3
and LDL Is Markedly Reduced in Cultured Fibroblasts From Tangier Patients
Authors: Drobnik Wolfgang , Möllers Christoph , Resink Therese , Schmitz Gerd
Published: June 2011

Abstract
We compared HDL
3
- and LDL-induced signal transduction in normal and Tangier fibroblasts to elucidate whether impaired signal transduction responses to lipoproteins might contribute to disturbed cellular lipid and lipoprotein metabolism in Tangier disease, a rare autosomal disorder of cellular lipid and lipoprotein metabolism. In several cell types HDL and LDL activate a currently unknown isoform of phosphatidylinositol-specific phospholipase C (PI-PLC) that results in the generation of 1,2-diacylglycerol and inositol 1,4,5-trisphosphate. Compared with normal fibroblasts, Tangier fibroblasts stimulated with HDL
3
or LDL resulted in a significantly reduced accumulation of inositol phosphates and 1,2-diacylglycerol formation. Furthermore, in Tangier fibroblasts both lipoproteins failed to mobilize calcium from internal pools, and the cytosol-to-membrane redistribution of protein kinase C (in both the α and ε isoforms) was markedly reduced. Thus, the data indicate an impaired PI-PLC activation in response to lipoproteins in Tangier fibroblasts.
Plasminogen Activator System in Human Coronary Atherosclerosis
Authors: Raghunath P.N. , Tomaszewski John E. , Brady Stephen T. , Caron Robert J. , Okada S. Steve , Barnathan Elliot S.
Published: June 2011

Abstract
Altered coronary artery expression of plasminogen activator (PA) system components may predispose to thrombosis and modulate the vascular response to injury. By immunohistochemistry, we studied the expression of PAs (tPA and uPA), their major physiological inhibitor (PAI-1), and a receptor for uPA (uPAR) in human coronary arteries with either pure fibrointimal proliferation (n=15) or developed atherosclerotic plaques (n=10). Overall, the degree of staining showed the following rank order: PAI-1>tPA>uPAR>uPA. A similar pattern was seen in two normal coronary arteries. There were no significant differences in the extent of staining in any vascular compartment between atherosclerotic arteries and those with only fibrointimal proliferation. However, the ratio of intimal to medial expression of tPA (
P
=.001) and uPAR (
P
=.004) was significantly increased in atherosclerotic arteries, with a similar trend for uPA (
P
=.069) but not for PAI-1 (
P
=.73). Four of 10 atherosclerotic arteries had higher uPAR expression in the intima than in the media, whereas none of the 15 arteries with only fibrointimal proliferation had this pattern (
P
<.01). Dual labeling studies demonstrated colocalization of all four PA system components in endothelial cells, smooth muscle cells, and macrophages, with a predominance of PAI-1. Thus, coronary arteries with a wide range of vascular pathology express an abundance of antifibrinolytic potential with enhanced local expression of profibrinolytic proteins, mainly within atherosclerotic plaques.
Surface Recruitment but Not Activation of Integrin α
IIb
β
3
(GPIIb-IIIa) Requires a Functional Actin Cytoskeleton
Authors: Addo John B. , Bray Paul F. , Grigoryev Dmitriy , Faraday Nauder , Goldschmidt-Clermont Pascal J.
Published: June 2011

Abstract
Binding of integrin α
IIb
β
3
(glycoprotein [GP] IIb-IIIa) to soluble fibrinogen requires that the receptor undergo a conformational change (receptor activation), which occurs rapidly in agonist-stimulated platelets. Agonist stimulation of platelets also results in α
IIb
β
3
recruitment from intracellular membranes (α-granules and open canalicular system) to the platelet surface. Once activated and accessible, the receptor can engage, a process that corresponds to the binding of the receptor to its soluble fibrinogen ligand, leading to intracellular signaling reactions and centripetal migration of bound receptor molecules. Because these processes occur concurrently with a marked reorganization of the actin cytoskeleton, we investigated the role of actin in fibrinogen receptor activation and surface recruitment. We used a flow cytometric assay to directly quantitate the binding of α
IIb
β
3
to fluorescently labeled fibrinogen on the platelet surface. Cytochalasin D, which inhibits elongation of actin filaments, was used to prevent the actin response to platelet agonists. Despite its ability to inhibit the actin response and α
IIb
β
3
binding to the actin cytoskeleton, cytochalasin D did not alter the agonist-induced intramolecular changes resulting in increased affinity of α
IIb
β
3
for soluble fibrinogen and therefore did not inhibit ADP-induced aggregation. Thus, disruption of the actin network with cytochalasin D had no effect on the dissociation constant of the complex between activated α
IIb
β
3
and fibrinogen (K
d
=0.26 to 0.28 μmol/L). However, cytochalasin D suppressed the recruitment of cryptic α
IIb
β
3
molecules to the platelet surface. While the physiological consequence of exposing additional α
IIb
β
3
molecules on the surface of platelets is unclear, it is tempting to speculate that this process plays an important role in consolidating intra-arterial platelet thrombi, despite the shear strain generated by the arterial blood flow.
Plasma Homocysteine and Severity of Atherosclerosis in Young Patients With Lower-Limb Atherosclerotic Disease
Authors: van den Berg Michiel , Stehouwer Coen D.A. , Bierdrager Edwin , Rauwerda Jan A.
Published: June 2011

Abstract
Elevated plasma homocysteine levels are recognized as an independent risk factor for atherosclerotic disease. It is not known (1) whether the severity of atherosclerotic disease is related to hyperhomocyst(e)inemia or (2) whether any such relation differs between fasting and post–methionine loading plasma homocysteine levels. Therefore, in 171 consecutive patients under 55 years of age with first symptoms of lower-limb disease, we examined the relation between severity of atherosclerosis and plasma homocysteine concentration. Severity of atherosclerotic disease was estimated from the prevalence of coronary artery disease and cerebrovascular disease and from the angiographic extent of lower-limb disease. Plasma homocysteine was measured after a period of fasting and in response to methionine loading (0.1 g/kg). In multivariate analysis, the prevalence of coronary artery disease plus cerebrovascular disease was related to both fasting and postmethionine homocysteine levels (odds ratio [OR] for the upper quartile versus the lower three quartiles, 2.8, 95% confidence interval [CI], 1.1 to 7.5; and OR 3.0, 95% CI, 1.1 to 7.8, respectively). The extent of lower-limb disease was weakly related to the fasting homocysteine level (partial correlation coefficient, .12;
P
=.17) and more strongly related to the postmethionine homocysteine level (partial correlation coefficient, .25;
P
=.003). These relations tended to be more pronounced in women than in men. They were independent of age, total serum cholesterol, blood pressure, and smoking habit. We concluded that the severity of atherosclerotic disease in young patients with lower-limb atherosclerotic disease is associated with high postmethionine and fasting homocysteine concentrations.
Blood Velocity Profiles in the Human Renal Artery by Doppler Ultrasound and Their Relationship to Atherosclerosis
Authors: Yamamoto Tokunori , Ogasawara Yasuo , Kimura Akihiro , Tanaka Hiroyoshi , Hiramatsu Osamu , Tsujioka Katsuhiko , Lever M. John , Parker Kim H. , Jones Christopher J.H. , Caro Colin G. , Kajiya Fumihiko
Published: June 2011

Abstract
Blood velocity profiles were measured in the renal branch (diameter 5.9±1.3 mm) of the aortorenal bifurcation using a 20-MHz 80-channel pulsed Doppler velocimeter during retroperitoneal surgery in 10 patients. The peak Reynolds number was 1145±140 and the frequency parameter (Wormersley parameter) was 3.0±0.8. Immediately distal to the ostium of the renal artery, reverse flow, indicating flow separation, was observed near the cranial wall mainly during the first part of the cardiac cycle. There were flows from the cranial to the caudal side of the artery at this location, indicating the presence of strong secondary flows. Two diameters downstream of the ostium, the velocity profiles were skewed to the caudal side in all patients. Four diameters downstream, the flow profile was symmetrical (3 patients) or only slightly skewed (7 patients) and virtually parabolic throughout the cardiac cycle. These observations mean that the flow on the cranial side of the renal branch of the human aortorenal bifurcation is characterized by (1) a bidirectional oscillation of the flow, (2) separation of the flow during systole, and (3) low time-averaged shear rate. These blood velocity patterns may be related to the localization and development of atheromatous plaque that occurs preferentially in this region of the renal artery. Conversely, the unidirectional, axisymmetrical flow found in more distal parts of the renal artery are associated with a very low incidence of lesions.
Hormone Replacement Therapy Lowers Plasma Lp(a) Concentrations
Authors: Taskinen M.-R. , Puolakka J. , Pyörälä T. , Luotola H. , Björn M. , Kääriäinen J. , Lahdenperä S. , Ehnholm C.
Published: June 2011

To study the responses of serum lipoproteins, apoproteins (apo's), and lipoprotein(a) (Lp[a]) to two frequently used hormone replacement therapies (HRTs), 120 postmenopausal women were randomly allocated to receive either transdermal therapy consisting of 28-day cycles with patches that delivered 17β-estradiol (50 μg/d) combined with cyclic oral medroxyprogesterone acetate (10 mg/d for 12 days per cycle) or continuous oral 17β-estradiol (2 mg/d) together with norethisterone acetate (1 mg/d) for 12 months. Blood samples were taken before and at 6 and 12 months of HRT. Concentrations of serum total, low density lipoprotein (LDL) and high density lipoprotein (HDL) cholesterol decreased by 14% (
P
<.001), 17% (
P
<.001), and 9% (
P
<.001) in the oral HRT group. Respective changes were 5.7% (
P
<.001), 4.8% (
P
<.05), and 4.7% (NS) in the transdermal group. Serum triglycerides remained unchanged in the oral group but decreased by 15.7% (
P
<.001) in the transdermal group. We observed only trivial changes in serum apo B levels. The changes in apo A-I levels paralleled those of HDL cholesterol in the oral HRT group. The concentration of serum Lp(a) decreased by 31% (
P
<.001) and 16% (
P
<.001) in the two groups. The combination of progestin and transdermal estrogen was not associated with any further change of Lp(a). The decrement in Lp(a) during therapy was positively associated with baseline Lp(a) levels in both groups (
r
=.96,
P
<.001 and
r
=.88,
P
<.001). Thus, both HRT regimens were highly effective in lowering elevated Lp(a) levels in postmenopausal women. The divergent responses of LDL and HDL cholesterol in the two HRT groups may influence the potential cardioprotective effects of the two HRT regimens. Prospective trials are needed to define the long-term effects with respect to coronary heart disease risk.
Endothelial Cells Inhibit NO Generation by Vascular Smooth Muscle Cells
Authors: Farré Antonio López , Mosquera Juan R. , Sánchez de Miguel Lourdes , Millás Inmaculada , de Frutos Trinidad , Montón Mercedes , Sierra María P. , Riesco Amparo , Casado Santos
Published: June 2011

Endothelial cell (EC)–released agents are active regulators of vascular smooth muscle cell (VSMC) functions. The first aim of the present work was to analyze the effect of ECs on interleukin-1β (IL-1β)–induced NO production by SMCs. Bovine aortic ECs (BAECs) and BVSMCs in culture were used for the study. IL-1β (0.03 U/L) stimulated nitrite production by BVSMCs. This increase was smaller in the presence of BAECs. This effect was accompanied by reduced expression of inducible NO synthase (iNOS) in BVSMCs coincubated with BAECs, as analyzed by Western blot analysis. The reduction in iNOS protein expression was partially reversed by a polyclonal antibody against transforming growth factor-β (TGF-β). Furthermore, we examined the cytotoxic effect of the NO released from BVSMCs on both BAECs and the BVSMCs themselves. Incubation of BAECs with IL-1β–prestimulated BVSMCs induced EC toxicity, which was partially inhibited by an inhibitor of NO synthesis,Nωnitro-l-arginine methyl ester, or an inhibitor of iNOS expression, dexamethasone. No cytotoxic effect of IL-1β on BVSMCs themselves was detected. ECs modulate iNOS expression in SMCs by mechanisms that include a TGF-β–dependent pathway. The NO released from SMCs exerts cytotoxic effects on the adjacent endothelium without altering the viability of the SMCs.
Identification of Two Novel Point Mutations in the Human Protein S Gene Associated With Familial Protein S Deficiency and Thrombosis
Authors: Li Muyao , Long George L.
Published: June 2011

Individuals with thrombosis who were believed to possess associated familial protein S deficiency were analyzed for mutations in the protein S gene by a two-step process. First, the individuals were analyzed for protein S Pro
626
A/G dimorphism in both their genomic DNA and reverse-transcribed (RT) polymerase chain reaction (PCR)–amplified cDNA from peripheral blood cell mRNA. If a heterozygote expressed both alleles at the mRNA level at this site in genomic DNA, a search for point mutations was made by direct cDNA sequencing. RT-PCR amplification of exons 1-6 with mRNA from two twin sisters, each of whom has severe type I protein S deficiency, revealed both larger and smaller fragments in addition to the expected 504–base pair fragment in normal individuals. A donor splice-site mutation at position +4 of the 5′ end of intron A was subsequently identified in both sisters and their mother. This mutation would lead to incorrect precursor mRNA splicing and the observed cDNA products. Translation of the altered mRNAs would result in a truncated protein without biological activity. In a second family, cDNA sequencing revealed a T→G mutation at codon 603 (Ile→Ser) in exon 15 of the protein S gene in an individual with protein S deficiency (mixed type) and a history of thrombosis. The same mutation was also detected in the proband's mother and grandmother, both of whom also exhibit protein S deficiency and thrombotic disease. This mutation occurs within a disulfide loop of protein S that is believed to be responsible for binding to C4b binding protein and may result in greater affinity between protein S and C4b, consequently leading to thrombotic disease.
Mouse Peritoneal Macrophages Contain Abundant ω-6 Lipoxygenase Activity That Is Independent of Interleukin-4
Authors: Cornicelli Joseph A. , Welch Kathryn , Auerbach Bruce , Feinmark Steven J. , Daugherty Alan
Published: June 2011

The action of an ω-6 lipoxygenase (LO) has been implicated in the development of atherosclerosis through a mechanism involving oxidation of LDL, and its regulation in macrophages may have important implications for the disease process. Human monocyte–derived macrophages (HMDMs) showed no demonstrable LO protein or activity unless they were incubated with interleukin-4 (IL-4). In contrast, mouse peritoneal macrophages (MPMs) possessed significant basal levels of LO activity and protein that were augmented by IL-4 treatment. Interferon gamma prevented the induction of LO in both HMDMs and MPMs. Whereas interferon gamma could completely block the IL-4 induction of LO in human cells, it did not suppress basal LO activity in MPMs. Both HMDMs and MPMs exhibited similar concentration-response relationships for stimulation of LO activity and protein, with maximal induction at 1 ng/mL IL-4. The time course of IL-4 induction of LO activity was markedly different in human and murine cells. IL-4 induced LO activity and protein in human cells by 48 hours that were maximal by 72 hours; there was a decline to a new baseline by 96 hours. MPMs have a significant amount of LO activity at baseline, which declined with time by nearly 10-fold in the absence of IL-4. IL-4 blunted the decline of LO activity with time and restored activity to that found at baseline by 48 hours. IL-4 was not responsible for the LO activity present in freshly isolated MPMs since both activity and protein content were similar in cells harvested from IL-4+/+and IL-4−/−mice. Therefore, whereas IL-4 may be an important modulator of LO production in vitro, it is not essential for the in vivo expression of this protein. Further, these studies demonstrate that significant differences exist between monocyte-derived macrophages matured in vitro and tissue macrophages that have matured in vivo.
Selective Inhibition of Tumor Necrosis Factor–Induced Vascular Cell Adhesion Molecule-1 Gene Expression by a Novel Flavonoid
Authors: Wölle Joachim , Hill Russell R. , Ferguson Erika , Devall Larry J. , Trivedi B.K. , Newton Roger S. , Saxena Uday
Published: June 2011

In the present studies, we examined the effect of flavonoids on the endothelial cell expression of adhesion molecules, an early step in inflammation and atherogenesis. Addition of tumor necrosis factor-α (TNF) to human aortic endothelial cells (HAECs) led to the induction of vascular cell adhesion molecule-1 (VCAM-1) expression and enhancement in expression of intercellular adhesion molecule-1 (ICAM-1). A flavonoid, 2-(3-amino-phenyl)-8-methoxy-chromene-4-one (PD 098063), markedly inhibited TNF-induced VCAM-1 cell-surface expression in a concentration-dependent fashion with half-maximal inhibition at 19 μmol/L but had no effect on ICAM-1 expression. Another structurally distinct flavonoid, 2-phenyl-chromene-4-one, similarly selectively decreased VCAM-1 expression. The inhibition in cell-surface expression of VCAM-1 by PD 098063 correlated with decreases in steady-state mRNA levels, but there was no effect on ICAM-1 mRNA levels. The decrease in VCAM-1 mRNA levels was not due to changes in mRNA stability but rather resulted from a reduction in the rate of transcription of the gene. However, electrophoretic mobility shift assays using nuclear extracts from TNF-induced HAECs treated with PD 098063 failed to show a decrease in the activation of NF-κB, indicating that inhibition of activation of this transcription factor may not be its mode of action. Similarly, PD 098063 did not affect chloramphenicol acetyltransferase reporter gene activity in TNF-inducible minimal VCAM-1 promoter constructs containing two NF-κB sites, suggesting that the compound does not affect the transactivation driven by these sites. We conclude that this compound selectively blocks agonist-induced VCAM-1 protein and gene expression in HAECs by NF-κB–independent mechanism
Heparin-Binding Domain of Fibrin Mediates Its Binding to Endothelial Cells
Authors: Odrljin Tatjana M. , Francis Charles W. , Sporn Lee Ann , Bunce Leslie A. , Marder Victor J. , Simpson-Haidaris Patricia J.
Published: June 2011

Spreading of human umbilical vein endothelial cells (ECs) on fibrin requires thrombin cleavage of fibrinopeptide B (FPB) and subsequent exposure of the new β15-42
N
-terminus. To further understand the interactions between ECs and fibrin β15-42 sequences, binding of fibrin(ogen) to EC monolayers was measured with polyclonal anti-fibrinogen (FBG) in parallel with monoclonal anti-FBG (18C6, β1-21; J88B, γ63-78) and anti-fibrin (T2G1, β15-21) antibodies in an indirect enzyme-linked immunosorbent assay. To accomplish this, large, soluble fragments of fibrin were prepared by cyanogen bromide (CNBr) cleavage (fibrin-CNBr); CNBr-cleaved FBG (FBG-CNBr) served as the control ligand.
N
-terminal fibrin-CNBr bound to EC monolayers and cells in suspension in a dose-dependent and saturable manner. By contrast, FBG-CNBr bound only 50% as well to EC monolayers, with no significant binding of intact FBG,
C
-terminal FBG plasmic fragment D, or
N
-terminal plasmic fragment E, which lacks β1-53. ECs bound the peptide β15-42–bovine serum albumin (BSA) conjugate but neither a scrambled β15-42 peptide conjugate nor conjugates of β24-42, β18-27, or β18-31. Binding of fibrin-CNBr was inhibited 54% by the β15-42–BSA conjugate and 17% by the Bβ1-42-BSA conjugate but not by free β15-42 peptide or RGDS-cell binding peptide. Binding of fibrin-CNBr was inhibited >95% by heparin in a concentration-dependent manner; the same concentrations of heparin inhibited binding of β15-42–BSA by >75% but not the dose-dependent binding of fibronectin to ECs. These data suggest that in their native conformation, FBG Bβ15-42 sequences are unavailable for binding to ECs and that thrombin-induced exposure of β15-42 is required for binding by a heparin-dependent, RGD-independent mechanism at the new
N
-terminus of fibrin.
Active Oxygen Species and Lysophosphatidylcholine Are Involved in Oxidized Low Density Lipoprotein Activation of Smooth Muscle Cell DNA Synthesis
Authors: Stiko Ann , Regnström Jan , Shah Prediman K. , Cercek Bojan , Nilsson Jan
Published: June 2011

AbstractIt has recently been shown that oxidative modification of LDL enhances the mitogenic effect of LDL on smooth muscle cell (SMC) DNA synthesis. However, because of its complex chemical structure, the mitogenic components have not been well characterized. Exposure of LDL to the oxidant Cu2+is followed by a rapid accumulation of peroxides that peaks after 8 to 12 hours and a conversion of the phospholipid phosphatidylcholine into lysophosphatidylcholine that continues for up to 48 hours. Most of the mitogenic activity is formed during the first 4 hours of oxidation. Both superoxide dismutase and catalase effectively inhibit the mitogenic activity of oxidized LDL, suggesting involvement of reactive oxygen intermediates. In the presence of 1% serum, low concentrations of hydrogen peroxide activated SMC DNA synthesis in a dose-dependent manner, with a maximal effect at a concentration of 200 μmol/L, whereas higher concentrations were inhibitory. Lysophosphatidylcholine also enhanced SMC DNA synthesis, with a maximal stimulation at a concentration of 10 μmol/L. Oxysterols, which also accumulate in oxidized LDL, effectively inhibited DNA synthesis. These results demonstrate that oxidation of LDL is associated with formation of several substances affecting the growth of SMCs. Among these substances, low levels of reactive oxygen intermediates and lysophosphatidylcholine stimulate DNA synthesis, whereas at a higher concentration they, as well as oxysterols, are inhibitory.
Effects of Colestipol Alone and in Combination With Simvastatin on Apolipoprotein B Metabolism
Authors: Gaw Allan , Packard Christopher J. , Lindsay Grace M. , Murray Elizabeth F. , Griffin Bruce A. , Caslake Muriel J. , Colquhoun Ian , Wheatley David J. , Lorimer A. Ross , Shepherd James
Published: June 2011

Abstract
The effects of colestipol therapy alone (20 g/d) or combined with simvastatin (20 mg/d) were examined in a group of eight male patients with primary moderate hypercholesterolemia (total cholesterol ≥6.5 mmol/L [≥250 mg/dL]) who had undergone coronary artery bypass grafting more than 3 months previously. Colestipol therapy decreased total cholesterol by 14% (
P
<.001) and LDL cholesterol (LDL-C) by 23% (
P
<.001), while dual therapy decreased total cholesterol by 38% and LDL-C by 52% (both
P
<.001 versus baseline). No significant changes were observed in plasma triglyceride, VLDL cholesterol, or HDL cholesterol levels. VLDL subfraction turnovers were conducted at baseline and again on each regimen. ApoB kinetic parameters derived from a multicompartmental model suggested that colestipol therapy resulted in an expansion of the total VLDL apoB pool (36%,
P
<.05) that was largely due to a fall in the clearance rate of VLDL
1
apoB (49%), while the LDL apoB pool decreased 23% as a result of diminished direct LDL input. The model used also revealed that addition of simvastatin to the resin therapy caused increases in the fractional transfer rates of VLDL
2
to IDL and IDL to LDL together with a 37% increment in the LDL apoB fractional catabolic rate. Compared with baseline, combined therapy generated falls in both IDL (35%,
P
=.01) and LDL (37%,
P
<.04) apoB pools due to enhanced clearance of IDL (214%,
P
<.03) and reduced total input of LDL (39%,
P
<.003).
In Hypercholesterolemia, Lower Peripheral Monocyte Count Is Unique Among the Major Predictors of Atherosclerosis
Authors: Huang Zei-Shung , Wang Chiu-Hwa , Yip Ping-Keung , Yang Chi-Yu , Lee Ti-Kai
Published: June 2011

Abstract
Many studies have shown that enhanced monocyte adherence is an important factor in the initiation of atherosclerosis. Because the relationships between circulating monocyte count and atherosclerosis or its major predictors have received little attention, we conducted this study with the aim of clarifying these relationships. The study included 409 men and women who underwent a carotid artery duplex study and white blood cell analysis (Sysmex Cell Counter) during a 2-day health check at our hospital in 1994. We found no correlation between preexisting carotid atherosclerosis and monocyte count. After adjustment for age and sex, hypercholesterolemia, among the major predictors of atherosclerosis, showed a unique correlation with both lower monocyte count and percentage (
P
<.001,
P
<.0001, respectively), whereas smoking was correlated with a higher monocyte count (
P
<.001). There was a slight but nonsignificant increase in monocyte count in hypertension, diabetes, and hypertriglyceridemia. Our results imply that: (1) hypercholesterolemia has a strong, peripheral monocyte–reducing effect, probably due to direct enhancement of monocyte adhesion to the endothelium, which subsequently initiates the atherosclerotic process, and (2) the mechanisms of other predictor(s)-induced atherosclerosis may be quite different from that of hypercholesterolemia. Another possible explanation for the inverse correlation between monocyte count and serum cholesterol level is that decreased monocyte levels might lead to hypercholesterolemia because of decreased uptake of cholesterol from the plasma by less monocyte-derived macrophages. The reasons why preexisting carotid atherosclerosis did not correlate with monocyte count are also discussed.
Vital Exhaustion, Anger Expression, and Pituitary and Adrenocortical Hormones
Authors: Keltikangas-Järvinen Liisa , Räikkönen Katri , Hautanen Aarno , Adlercreutz Herman
Published: June 2011

Abstract
This study was undertaken to examine whether there are psychological factors that can incline an individual toward coronary heart disease and that can in turn identify a pattern of pituitary and adrenocortical responses that is associated with the Insulin Resistance Syndrome (IRS). The study was performed with 69 normotensive and 21 unmedicated borderline hypertensive men (age range, 30 to 55 years). Type A behavior, hostility (defined as cynicism, pessimism, and paranoia), vital exhaustion, and anger expressions were the behavioral variables studied. Among these, only the vital exhaustion–anger-out factor identified the neuroendocrine pattern that predicted the IRS. This neuroendocrine pattern consisted primarily of an adrenal responsiveness to ACTH and secondarily of a high mean basal cortisol–to–mean basal ACTH ratio. The contribution of this last variable was, however, slightly questionable. Instead of the traditional coronary-prone factors, ie, type A behavior and hostility, the findings emphasize the significance of vital exhaustion and emotional distress. The findings have been discussed in terms of defeat reaction, hypocortisolemia, and visceral obesity.
Genetic Analysis of the IRS
Authors: Mitchell Braxton D. , Kammerer Candace M. , Mahaney Michael C. , Blangero John , Comuzzie Anthony G. , Atwood Larry D. , Haffner Steven M. , Stern Michael P. , MacCluer Jean W.
Published: June 2011

Abstract
Insulin resistance is part of a metabolic syndrome that also includes non–insulin-dependent diabetes mellitus, dyslipidemia, obesity, and hypertension. It has been hypothesized that insulin resistance represents the primary physiological defect underlying this syndrome. Since insulin resistance is at least partially genetically determined, we hypothesized that genes influencing insulin resistance would have pleiotropic effects on a number of other traits, including triglyceride (TG) and HDL cholesterol levels, body mass index (BMI) and body fat distribution, and blood pressure levels. To investigate this hypothesis, we analyzed data obtained from individuals in 41 families enrolled in the San Antonio Family Heart Study. Statistical methods that take advantage of the relatedness among individuals were used to differentiate between genetic and nongenetic (ie, environmental) contributions to phenotypic variation between traits. Serum levels of fasting and 2-hour insulin (measured in 767 and 743 nondiabetic family members, respectively) were used as a measure of insulin resistance. The genetic correlations were high between insulin levels (both fasting and 2-hour) and each of the following: BMI, HDL level, waist-to-hip ratio, and subscapular-to-triceps ratio, indicating that the same gene, or set of genes, influences each pair of traits. In contrast, the genetic correlations of insulin levels with systolic and diastolic blood pressures were low. We have previously shown that a single diallelic locus accounts for 31% of the phenotypic variation in 2-hour insulin levels in this population. We conducted a bivariate segregation analysis to see if the common genetic effects on insulin and these other traits could be attributable to this single locus. These results indicated a significant effect of the 2-hour insulin locus on fasting insulin levels (
P
=.02) and BMI (
P
=.05), with the “high” insulin allele associated with higher levels of fasting insulin but lower levels of BMI. There was no detectable effect of this locus on HDL level, TG level, subscapular-to-triceps ratio, or blood pressure. Overall, these results suggest that a common set of genes influencing insulin levels also influences other insulin resistance syndrome–related traits, although for the most part this pleiotropy is not attributable to the 2-hour insulin level major locus.