bluuebunny/crossref_metadata_2025 huggingface dataset:
Uptake of Oxidized LDL by Macrophages Differs From That of Acetyl LDL and Leads to Expansion of an Acidic Endolysosomal Compartment
Authors: Lougheed Marilee , Moore Edwin D. W. , Scriven David R. L. , Steinbrecher Urs P.
Published: June 2011

Abstract
—Accumulation of cholesterol by macrophage foam cells in atherosclerotic lesions is thought to involve the uptake of modified low density lipoproteins (LDLs). Previous studies have shown that there is impaired degradation of oxidized LDL in macrophages. The present study was done to determine whether the differences in intracellular metabolism of oxidized LDL and acetyl LDL were associated with delivery to different intracellular compartments. Mouse peritoneal macrophages were incubated with 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlo- rate–labeled oxidized LDL or 3,3′-dioctadecyloxacarbocyanine perchlorate–labeled acetyl LDL and examined by fluorescence microscopy. Deconvolution image analysis showed <10% colocalization of the 2 lipoproteins at incubation times ranging from 30 minutes to 6 hours. Subcellular fractionation of macrophages after incubation with
99m
Tc-labeled oxidized LDL revealed accumulation of the tracer in a compartment with a
d
=1.042 g/mL, consistent with endosomes. Surprisingly, there was a concurrent dramatic shift of the density of lysosomal marker enzymes from
d
=1.1 g/mL to the same fractions that contained
99m
Tc, indicating that this compartment was formed after fusion with primary lysosomes. Parallel experiments in J774 cells, a murine macrophage–like cell line, did not show a similar density shift, perhaps because of the slower rate of accumulation of oxidized LDL by these cells. Fluorescence microscopy of macrophages labeled with a lysosomotropic dye revealed a marked expansion of the acidic compartment after exposure of cells to oxidized LDL. We conclude that oxidized LDL and acetyl LDL are internalized by morphologically distinct pathways. Furthermore, because of its impaired lysosomal degradation, oxidized LDL causes expansion of and a decrease in the density of the lysosomal compartment in macrophages.
Inhibition of Arteriosclerosis by T-Cell Depletion in Normocholesterolemic Rabbits Immunized With Heat Shock Protein 65
Authors: Metzler Bernhard , Mayr Manuel , Dietrich Hermann , Singh Mahauir , Wiebe Evelyn , Xu Qingbo , Wick Georg
Published: June 2011

Abstract—Previous studies in our laboratory have shown that arteriosclerotic changes can be induced in normocholesterolemic rabbits by immunization with mycobacterial heat shock protein (hsp) 65. To further investigate the immunologic mechanisms underlying such vascular lesions, 39 male New Zealand White rabbits were treated by triple immunization with fortified Freund’s complete adjuvant containing 5 mg/mLMycobacterium tuberculosisas a source of hsp65 and simultaneous immunosuppressive therapy twice per week with either anti-CD3 monoclonal antibody (1 mg/kg) and prednisolone (1 mg/kg) or prednisolone (1 mg/kg) alone. Sixteen weeks after the first immunization the animals were killed, and as expected, severe arteriosclerotic lesions in the intima of the aortic arch were found in 9 of 10 immunized rabbits. However, only 1 of 10 rabbits immunized and immunosuppressed with the combined anti-CD3 monoclonal antibody and prednisolone treatment showed a single moderate lesion in the aorta, whereas 5 of 9 rabbits immunized and immunosuppressed by prednisolone treatment alone showed lesions, albeit mild. In conclusion, the early inflammatory stages of arteriosclerotic lesions induced by immunization with hsp65 can be inhibited by immunosuppressive therapy with anti-CD3 monoclonal antibody.
Fluctuations of Radial Artery Distensibility Throughout the Menstrual Cycle
Authors: Giannattasio Cristina , Failla Monica , Grappiolo Alessandra , Stella Maria Luisa , Del Bo Alberto , Colombo Manuela , Mancia Giuseppe
Published: June 2011

Abstract
—Estrogen administration has a number of favorable cardiovascular effects, and recent evidence suggests that these include an increase in arterial distensibility. Whether this is also the case for the physiological changes in estrogen production during the menstrual cycle has never been determined, however. In 21 premenopausal healthy women, we continuously measured radial artery diameter and blood pressure by an echo-tracking device and a beat-to-beat finger device, respectively. Arterial distensibility was calculated as distensibility/blood pressure curve. The measurements were made during the follicular, ovulatory, and luteal phases of the menstrual cycle. As expected, compared with the follicular phase, plasma estradiol, follicle-stimulating hormone, luteinizing hormone, and prolactin were increased in the ovulatory phase, whereas progesterone was increased in the luteal phase, together with antidiuretic hormone. Radial artery distensibility was increased in the ovulatory and reduced in the luteal phase, the changes being independent of the small, concomitant blood pressure changes. The arterial wall stiffening seen in the luteal phase was associated with a reduction in the flow-dependent endothelial dilatation of the radial artery as assessed by the hyperemia after short-term ischemia of the hand. Thus, the natural menstrual cycle is characterized by alterations in radial artery distensibility. The mechanisms responsible for this phenomenon remain to be clarified. It is possible, however, that the greater arterial distensibility of the ovulatory phase is due to an estrogen-dependent reduction in vascular smooth muscle tone, whereas the arterial stiffening of the luteal phase depends on vascular smooth muscle contraction due to more complex hormonal phenomena, ie, an endothelial impairment due to estrogen reduction but also to an increase in progesterone and antidiuretic hormone levels.
Urinary Albumin Excretion
Authors: Borch-Johnsen Knut , Feldt-Rasmussen Bo , Strandgaard Svend , Schroll Marianne , Jensen Jan Skov
Published: June 2011

Abstract
—Cross-sectional studies suggest that an increased urinary albumin excretion rate is associated with cardiovascular disease, dyslipidemia, and hypertension. The purpose of this study was to analyze prospectively whether the urinary albumin-to -creatinine (A/C) ratio can independently predict ischemic heart disease (IHD) in a population-based cohort. In 1983, urinary albumin and creatinine levels were measured, along with the conventional atherosclerotic risk factors, in 2085 consecutive participants without IHD, renal disease, urinary tract infection, or diabetes mellitus. The participants were followed up until death, emigration, or December 31, 1993. IHD was defined as a hospital discharge diagnosis or cause of death including the diagnoses ICD-8 and 410 to 414. Seventy-nine individuals developed IHD during the 21 130 person-years of follow-up. They were characterized by a preponderance of males and higher age, body mass index, blood pressure, lipoproteins, and proportion of current smokers. Microalbuminuria was defined as an A/C ratio) >90 percentile (>0.65 mg/mmol). When adjusted for other risk factors, the relative risk of IHD associated with microalbuminuria was 2.3 (95% CI, 1.3 to 3.9,
P
=0.002), and the 10-year disease-free survival decreased from 97% to 91% (
P
<0.0001) when microalbuminuria was present. An interaction between microalbuminuria and smoking was observed, and the presence of microalbuminuria more than doubled the predictive effect of the conventional atherosclerotic risk factors for development of IHD. It is concluded that microalbuminuria is not only an independent predictor of IHD but also substantially increases the risk associated with other established risk factors.
Oral Contraceptives Highlight the Genotype-Specific Association Between Serum Phospholipids and Activated Factor VII
Authors: Mariani G. , Conard J. , Bernardi F. , Bertina R. , Garcia V. Vicente , Prydz H. , Samama M. , Sandset P. M. , Puopolo M. , Ciarla M. V. , Poso R. , Di Nucci G. D. , Ceci F. , Marchetti G.
Published: June 2011

Abstract
—The present analysis was undertaken to study the effect of oral contraceptive (OC) use on activated factor VII (FVIIa) in subjects characterized by FVII genotypes, with the further aim of evaluating the role of lipids in this pharmacological interaction. In OC users (n=42) and nonusers (n=130) of comparable age, we examined the FVII phenotypic variables (FVII coagulant activity [FVIIc], FVII antigen, and FVIIa), FVII genotypes (the 353R/Q and 5′F7 polymorphisms analyzed in combination; alleles
M1
/
M2
and
A1
/
A2
, respectively), and a number of lipid and lipoprotein parameters: serum concentrations of total cholesterol (chol), low density lipoprotein and high density lipoprotein-chol, triglycerides, phospholipids (PhLs), apolipoprotein A1, and lipoprotein(a). PhLs, triglycerides, apolipoprotein A1, chol, FVII antigen, FVIIc, and high density lipoprotein-chol levels were shown to be statistically higher in users than nonusers. FVII levels, particularly those of FVIIa and FVIIc, were much higher in homozygotes for the
A1
and
M1
alleles (
A11
M11
), especially in OC users. A strong association was found between PhL and FVIIa: in the multiple regression analysis, women taking OCs who had elevated PhL concentrations also had very high levels of FVIIa, but only if their genotype was
A11 M11
. These results indicate that the increased FVII levels in OC users depend on the FVII genotype and that high PhL concentrations predict very high levels of FVIIa and FVIIc.
LDL Size in African Americans, Hispanics, and Non-Hispanic Whites
Authors: Haffner Steven M. , D’Agostino Ralph , Goff David , Howard Barbara , Festa Andreas , Saad Mohammed F. , Mykkänen Leena
Published: June 2011

Abstract
—The prevalence of cardiovascular disease (CVD) and atherosclerosis varies among several minority ethnic groups in the United States. Recently, small, dense low density lipoprotein (LDL) particle size has been recognized as a risk factor for CVD. We examined LDL size as a possible explanation for differences in CVD rates in 1571 subjects from the Insulin Resistance Atherosclerosis Study (IRAS), a multiethnic study of insulin resistance and cardiovascular risk factors. LDL size (Å) was significantly different by ethnic group (African Americans 262.1±0.6, Hispanics 257.6±0.6, and non-Hispanic whites 259.2±0.4,
P
<0.001). Ethnic differences in LDL size continued to be statistically significant after adjustment for upper body adiposity, insulin resistance, and glucose tolerance status. However, after further adjustment for other cardiovascular risk factors, especially ethnic differences in triglyceride and high density lipoprotein (HDL) cholesterol levels, the ethnic differences in LDL size were markedly attenuated and in general no longer statistically significant. The relation of triglyceride, HDL cholesterol, insulin resistance, and adiposity to LDL size in each ethnic group was similar. LDL size differs by ethnic group, which is independent of obesity or insulin resistance. These ethnic differences appear to be due to ethnic variations in dyslipidemia (especially differences in triglyceride levels); ethnic differences in LDL size are not consistent with previously reported ethnic dissimilarities in CVD or atherosclerosis.
Cellular Effects of Heparin on the Production and Release of Tissue Factor Pathway Inhibitor in Human Endothelial Cells in Culture
Authors: Lupu Cristina , Poulsen Emma , Roquefeuil Sylvie , Westmuckett Andrew D. , Kakkar Vijay V. , Lupu Florea
Published: June 2011

Abstract
—Tissue factor pathway inhibitor (TFPI), the major downregulator of procoagulant activity of the tissue factor–factor VIIa complex (TF · FVIIa), is synthesized and constitutively secreted by endothelial cells (ECs). Here we describe the in vitro effects of heparin on the cellular localization, gene expression, and release of TFPI in human ECs in culture. Both unfractionated heparin (UFH) and low-molecular-weight heparin (LMWH; Fragmin) time-dependently induced a significant enhanced secretion of TFPI, paralleled by a redistribution and increase of TFPI on the cell surface and a decrease of intracellular TFPI. Immunogold electron microscopy showed the presence of clusters of TFPI, both on the plasmalemma proper and within cell-surface opened caveolae/enlarged caveolar profiles. Activation of FX by TF · FVIIa on ECs treated with endotoxin was inhibited by both heparins but to a higher extent by LMWH. Inhibition of protein synthesis by cycloheximide did not reduce the release of TFPI induced by heparin. Long-term incubation (48 hours) resulted in a time-dependent enhanced production of TFPI. After the first 4 to 8 hours, depletion of intracellular TFPI was observed, more significantly with UFH. Northern blot analysis of TFPI mRNA also showed a decrease of the 1.4-kb transcript after 4 hours of incubation with UFH, followed by recovery and an increase over the control level after 24 hours. Incubation of ECs with phorbol ester (PMA) significantly enhanced the secretion of TFPI and increased its activity on the cell surface, probably by preventing invagination of caveolae. Heparin-stimulated release of TFPI decreased significantly in the presence of PMA to a level that was 2.4 times lower than the expected additive value for PMA and UFH separately. Pretreatment of ECs with PMA suppressed a subsequent response to heparin. Altogether, our results suggest that the heparin-induced release of TFPI might involve a more specific mechanism(s) than the previously hypothesized simple displacement of TFPI from the cell surface glycocalyx. We assume that the increased secretion and redistribution of cellular TFPI induced by heparins in ECs in culture can play an important role in the modulation of the anticoagulant properties of the endothelium.
Antithrombotic Efficacy of the Vitamin K Antagonist Fluindione in a Human Ex Vivo Model of Arterial Thrombosis
Authors: Bossavy Jean-Pierre , Sakariassen Kjell S. , Thalamas Claire , Boneu Bernard , Cadroy Yves
Published: June 2011

Abstract
—Thrombin is a main mediator of arterial thrombus formation, and its inhibition is an important antithrombotic strategy. However, the place of vitamin K antagonists among the different therapeutic strategies for preventing arterial thrombus formation is still debated. We studied the antithrombotic efficacy of the vitamin K antagonist fluindione in a human ex vivo model of arterial thrombosis and determined whether aspirin enhances fluindione efficacy. Ten healthy male volunteers were randomly assigned to receive fluindione, alone or in combination with aspirin (325 mg/d). Fluindione was given at increasing doses to give a stable international normalized ratio (INR) between 1.5 and 2.0 and between 2.1 and 3.0. We induced arterial thrombus formation ex vivo by exposing collagen- or tissue factor (TF)–coated coverslips in a parallel-plate perfusion chamber to native blood for 3 minutes at an arterial wall shear rate of 2600 s
−1
. Platelet and fibrin deposition were measured by immunoenzymatic methods. Fluindione inhibited thrombus formation on TF-coated coverslips in a dose-dependent manner by 50% and 80% at INR 1.5 to 2.0 and INR 2.1 to 3.0, respectively (
P
<0.05). Fluindione in combination with aspirin inhibited TF-induced thrombus formation in a comparable manner. Collagen-induced thrombus formation was not reduced in subjects treated by fluindione. It was reduced by 50% to 60% in those treated with fluindione plus aspirin, regardless of the level of anticoagulation (
P
<0.05). Thus, the effectiveness of fluindione for preventing arterial thrombosis is dependent on the nature of the thrombogenic trigger. Fluindione is very effective in preventing TF- but not collagen-triggered thrombus formation. Aspirin enhances the antithrombotic effectiveness of fluindione, because combined treatment interrupts both TF- and collagen-induced thrombus formation.
Plasma apolipoprotein A-1 absence associated with a marked reduction of high density lipoproteins and premature coronary artery disease.
Authors: Schaefer E J , Heaton W H , Wetzel M G , Brewer H B
Published: June 2011

A 45-year-old woman with corneal opacification and severe coronary artery disease was noted to have the following plasma lipid levels (mg/dl, +/- SD): total cholesterol 111 +/- 13, triglyceride 62 +/- 6, very low density lipoprotein cholesterol 4 +/- 1, low density lipoprotein cholesterol 106 +/- 14, and high density lipoprotein (HDL) cholesterol 1 +/- 1 (normal, 50 +/- 14). Her two offspring and one brother were found to have HDL cholesterol values (mg/dl) of 23, 20, and 20, respectively. The percentage of cholesterol in the esterified form in the patient's plasma was normal at 70%. Lipoprotein electrophoresis showed no alpha lipoprotein band, and no HDL was detectable when plasma was subjected to analytic ultracentrifugation. Only trace amounts of lipids were noted within the HDL density region following preparative ultracentrifugation. Mean plasma apolipoprotein (apo) A-ll, apo B, and apo C-ll plasma levels were 13.8%, 130.6% and 26.6% of normal, respectively. The ratio of apo B to cholesterol within LDL was elevated. Apo A-l, the major HDL protein constituent, was immunologically undetectable in this patient's plasma. A decreased HDL cholesterol concentration has been associated with premature coronary artery disease. These data indicate that plasma apo A-l absence results in a striking reduction in HDL, is associated with premature coronary artery disease, and represents a new distinct disease entity.
Effect of interferon on plasma lipoproteins and on the activity of postheparin plasma lipases.
Authors: Ehnholm C , Aho K , Huttunen J K , Kostiainen E , Mattila K , Pakkarainen J , Cantell K
Published: June 2011

The effect of interferon administration on the concentration of plasma lipoproteins and on the activity of postheparin plasma lipoprotein lipase and hepatic lipase was studied in six healthy men. Daily injection of human leukocyte interferon for 1 week lowered the plasma level of total cholesterol, very low density lipoprotein + low density lipoprotein cholesterol, high density lipoprotein cholesterol, and apolipoprotein A-1 in all subjects. Simultaneously, the activity of postheparin plasma hepatic lipase and lipoprotein lipase decreased by 20% to 50%. These observations may be of importance in the interpretation of lipoprotein changes seen in acute and chronic infections and should be borne in mind when prolonged treatment with interferon is considered.
Cholesterol homeostasis of skin fibroblasts after incubation with postabsorptive and postprandial lipoproteins. The effect of a fatty meal.
Authors: Kenagy R D , Florén C H , Bierman E L , Kudchodkar B , Albers J J
Published: June 2011

To determine if lipoproteins formed after a fatty meal deliver more cholesterol to cultured skin fibroblasts than do lipoproteins in the basal state, very low density lipoproteins and remnants (d less than 1.019), low density lipoproteins (LDL), and high density lipoproteins (HDL) were isolated from plasma obtained before, and 3 and 6 hours after, consumption of a high fat-cholesterol formula by seven normal males. Binding of 125I-LDL to cells and cell cholesterol content were determined after incubation of normal human skin fibroblasts for 48 hours with the lipoprotein fractions at 5% or 15% of plasma concentration. Activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase was also measured after preincubation of cells with HLD for 48 hours. Despite a 40% increase in unesterified cholesterol in the d less than 1.019 fraction at 3 hours compared to the 0-hour fraction, the 3-hour d less than 1.019 fraction did not decrease LDL binding or increase cell cholesterol more than did the 0-hour fraction. Preincubation of cells with LDL, concentrations of which were unchanged by feeding, decreased LDL binding and increased cellular cholesterol. These effects also were not altered by the meal. HDL lipids and apo A-I were decreased at 3 hours, but not at 6 hours. Effects of HDL on LDL binding and cellular cholesterol were not altered by feeding, but the 3-hour and 6-hour fractions increased 3-hydroxy-3-methylglutaryl coenzyme A reductase activity, while the 0-hour fraction had little effect. These data indicate that consumption of a high fat-cholesterol meal as a bolus does not acutely alter the cholesterol delivery capacity of serum lipoproteins of normal male subjects.
Clustering of anthropometric parameters, glucose tolerance, and serum lipids in children with high and low beta- and pre-beta-lipoproteins. Bogalusa Heart Study.
Authors: Voors A W , Harsha D W , Webber L S , Radhakrishnamurthy B , Srinivasan S R , Berenson G S
Published: June 2011

Children initially aged 21/2 to 14 years living in Bogalusa, Louisiana (n = 2530) were examined twice, 3 years apart, for fasting serum pre-beta- and beta-lipoprotein cholesterol (beta-LPC) levels. Based on averages of these levels, the children were ranked for pre-beta- and beta-LPC in combinations of extreme quintiles (low-low, high-high) or quartiles (low-high, high-low), n = 388, and were reexamined for serum lipids, lipoprotein cholesterol, glucose tolerance, and anthropometry. Skinfolds were thicker in whites than in blacks except for subscapular skinfold. Children in the high-high stratum were heavier and more obese. The postglucose insulin level was positively correlated with fasting serum triglycerides and pre-beta-LPC. Compared with other strata, high-high strata showed more clustering among half-hour and 1-hour plasma insulin, serum triglycerides and pre-beta-LPC, and trunk skinfolds. We conclude that racial differences in lipid and carbohydrate metabolism occur in all four strata, and that a strong clustering occurs more in the high-high stratum, which may, in part, explain the coincidence of several high cardiovascular risk factor levels observed in the same children. These observations document in free-living children changes of obesity, plasma glucose, and insulin metabolism related to serum lipoproteins that are involved in the early natural history of atherosclerosis.
Social status, environment, and atherosclerosis in cynomolgus monkeys.
Authors: Kaplan J R , Manuck S B , Clarkson T B , Lusso F M , Taub D M
Published: June 2011

The purpose of this experiment was to examine the effects of social environment and social status on coronary artery and aortic atherosclerosis in adult male cynomolgus monkeys (Macaca fascicularis). Thirty experimental animals were assigned to six groups of five members each, and all animals were fed a moderately atherogenic diet (43% of calories as fat, 0.34 mg cholesterol/Cal) for 22 months. Group memberships were changed periodically among 15 monkeys (unstable social condition) and remained fixed throughout the experiment in the remaining animals (stable social condition). Within each condition, individual monkeys were classified as either dominant or subordinate animals, based on dyadic patterns of aggression and submission. At necropsy, the coronary arteries were subjected to pressure fixation and five sections each were taken from the left anterior descending, left circumflex, and right coronary arteries. The mean intimal area measurement, based on all arterial sections, served as a coronary index for each animal. Results indicated that dominant animals in the unstable condition had significantly greater coronary artery atherosclerosis than dominant monkeys housed in stable social groups. Coronary artery atherosclerosis in the unstable dominants was also greater than among similarly housed (i.e., unstable) subordinates. A similar pattern was observed in the abdominal aorta, but was not statistically significant. No significant differences or similar patterns were seen in the thoracic aorta. Additional analyses revealed that the coronary artery effects were not due to concomitant differences in total serum cholesterol or high density lipoprotein cholesterol concentrations, blood pressures, ponderosity, or fasting glucose concentrations among the experimental animals. Behaviorally, manipulation of group memberships intensified agonistic encounters and disrupted patterns of affiliative interaction between dominant and subordinate monkeys. Overall, these results suggest that social dominance (an individual behavioral characteristic) is associated with increased coronary artery atherosclerosis, but only under social conditions that provide recurrent threats to the status of dominant animals (i.e., under behavioral challenge).
Lipoprotein lipase in cultured pig aortic smooth muscle cells.
Authors: Vance J E , Khoo J C , Steinberg D
Published: June 2011

Acetone powder extracts prepared from cultured pig aortic smooth muscle cells and the culture medium from these cells (particularly when incubated with heparin) were shown to contain a lipolytic enzyme which was identified as lipoprotein lipase by the following criteria: 1) stimulation by apolipoprotein C-II; 2) an optimal activity at approximately pH 8.0; 3) inhibition by NaCl, and 4) binding to a heparin-Sepharose affinity column. In addition, we found that cultured arterial smooth muscle cells from guinea pig and rabbit secreted a similar lipase into the culture medium. In contrast, studies using cultured bovine aortic endothelial cells yielded no evidence for either the synthesis or secretion of lipoprotein lipase by these cells. The production of lipoprotein lipase by the smooth muscle cells of the artery may play a role in the process of atherogenesis.
Postprandial Hyperlipidemia in Streptozotocin-Induced Diabetic Rats Is Due to Abnormal Increase in Intestinal Acyl Coenzyme A:Cholesterol Acyltransferase Activity
Authors: Kusunoki Jun , Aragane Katsumi , Kitamine Tetsuya , Kozono Hideki , Kano Kyoko , Fujinami Kouji , Kojima Kazuhiro , Chiwata Tsuyoshi , Sekine Yasuo
Published: June 2011

Abstract
—Postprandial hyperlipidemia (PH) is recognized as a significant risk factor for cardiovascular disease. The present study, involving rats with streptozotocin (STZ)-induced diabetes, was performed to establish a PH model and to examine the relation between small intestinal acyl-coenzyme A:cholesterol acyltransferase (ACAT) activity and serum lipid levels in the postprandial state. The small intestinal ACAT activities in normal rats during the experimental period were 4 to 5 pmol/mg protein per minute. In contrast, in the diabetic rats, the ACAT activities were 2 to 3 times higher than activities seen in normal rats from 7 to 21 days after the STZ injection in the absence of a high fat diet and hyperplasia in the gut. In an oral fat-loading test that used diabetic rats that had been injected with STZ (60 mg/kg) intravenously 14 days previously, the postloading changes in the serum concentrations of total cholesterol (TC) and triglyceride (TG) were significantly greater in the diabetic rats than in normal rats. Single oral administration of (1
s
,2
s
)-2-[3-(2,2-dimethylpropyl)-3-nonylureido]cyclohexane-1-yl 3-[(4
R
)-
N
-(2,2,5,5-tetramethyl-1,3-dioxane-4-carbonyl)amino]propionate (F-1394, 3 to 30 mg/kg), a potent ACAT inhibitor, suppressed the post–fat-loading elevation of serum TC levels in the diabetic rats in a dose-dependent manner without affecting serum glucose levels. Furthermore, the small intestinal ACAT activity, serum TG levels, and lymphatic absorption of TC and TG in the rats that were administered F-1394 (30 mg/kg) were reduced by ≈90%, 70%, 30%, and 15%, respectively. This is the first evidence that elevated ACAT activity in the gut, unlike hyperplasia and hyperphagia, induces PH in rats. Our results strongly suggest that F-1394 may be a potential treatment for PH in humans.
A Single Amino Acid Deletion in the Carboxy Terminal of Apolipoprotein A-I Impairs Lipid Binding and Cellular Interaction
Authors: Huang Wei , Sasaki Jun , Matsunaga Akira , Han Hua , Li Wei , Koga Takafumi , Kugi Mari , Ando Setsuko , Arakawa Kikuo
Published: June 2011

Abstract
—The carboxy-terminal region of apolipoprotein (apo) A-I has been shown by mutagenesis or synthetic peptides to play an important role in lipid binding. However, the precise functional domain of the
C
-terminal remains to be defined. In this study, apoA-I Nichinan, a naturally occurring human apoA-I variant with a deletion of glutamic acid 235, was expressed in
Escherichia coli
to examine the effect of this mutation on the functional domain of apoA-I for lipid binding and related consequences. A dimyristoyl phosphatidylcholine binding study with recombinant (r-) proapoA-I Nichinan showed a significantly slow initial rate of lipid binding. On preincubation with human plasma lipoprotein fractions (
d
<1.225 g/mL) at 37°C for 1 hour,
125
I-labeled normal r-proapoA-I was chromatographed as a single peak at the high density lipoprotein (HDL) fraction, whereas
125
I-labeled r-proapoA-I Nichinan was chromatographed into the HDL fraction as well as the free r-proapoA-I fraction (23% of radioactivity). Circular dichroism measurements showed that the α-helix content of lipid-bound r-proapoA-I Nichinan was reduced, being 62% (versus 73%) of normal r-proapoA-I. Nondenaturing gradient gel electrophoresis of reconstituted HDL particles assembled with r-proapoA-I Nichinan and normal r-proapoA-I showed similar particle size. To study cholesterol efflux, human skin fibroblasts were labeled with [
3
H]cholesterol, followed by incubation with either lipid-free r-proapoA-I or DMPC/r-proapoA-I complex. Fractional cholesterol efflux from [
3
H]cholesterol-labeled fibroblasts to lipid-free r-proapoA-I Nichinan or DMPC/r-proapoA-I Nichinan complexes was significantly reduced relative to that of normal r-proapoA-I or DMPC/r-proapoA-I during the 6-hour incubation
.
Binding assays of human skin fibroblasts by lipid-free r-proapoA-I showed that r-proapoA-I Nichinan was 32% less bound to fibroblasts than was normal r-proapoA-I. Our data demonstrate that the deletion of glutamic acid 235 at the
C
-terminus substantially reduces the lipid-binding properties of r-proapoA-I Nichinan, which may cause a reduction in its capacity to interact with plasma membranes as well as to promote cholesterol efflux from cultured fibroblasts.
All-
trans
-Retinoic Acid Limits Restenosis After Balloon Angioplasty in the Focally Atherosclerotic Rabbit
Authors: Wiegman Peter J. , Barry William L. , McPherson John A. , McNamara Coleen A. , Gimple Lawrence W. , Sanders John M. , Bishop Gregory G. , Powers Eric R. , Ragosta Michael , Owens Gary K. , Sarembock Ian J.
Published: June 2011

Abstract
—All-
trans
-retinoic acid (atRA) has potent in vitro effects on a number of processes involved in vascular injury and repair, such as modulating smooth muscle cell (SMC) proliferation and inducing SMC differentiation, and may play an important role in the in vivo response to vascular injury. We hypothesized that atRA would limit restenosis after balloon angioplasty through SMC-modulated changes in plaque size and vessel geometry. Balloon angioplasty was performed on rabbits with focal femoral atherosclerosis randomized to treatment with atRA or saline. At 28 days after balloon angioplasty, minimal luminal diameter was significantly larger in the atRA group (1.24±0.17 versus 1.12±0.22 mm,
P
=0.02). Histomorphometry confirmed a larger lumen area (0.51±0.20 versus 0.34±0.13 mm
2
,
P
=0.004) in the atRA group, with no difference in absolute plaque area. Internal elastic lamina and external elastic lamina areas were significantly larger in the atRA group (0.89±0.27 versus 0.66±0.24 mm
2
,
P
=0.001, and 1.29±0.38 versus 0.98±0.32 mm
2
,
P
=0.001, respectively). Vessel sections exhibited significantly more α-actin and desmin immunostaining (
P
=0.01) in the atRA-treated group. No differences in early cellular proliferation and collagen content were detected with the use of bromodeoxyuridine. In this atherosclerotic model of vascular injury, atRA limits restenosis after balloon angioplasty by effects secondary to overall vessel segment enlargement at the angioplasty site rather than by effects on plaque size or cellular proliferation. Increased α-actin and desmin immunostaining suggest a possible role for phenotypic modulation of SMCs in this favorable remodeling effect.
Accumulation of LDL in Rat Arteries Is Associated With Activation of Tumor Necrosis Factor-α Expression
Authors: Niemann-Jönsson Audrey , Dimayuga Paul , Jovinge Stefan , Calara Federico , Ares Mikko P. S. , Fredrikson Gunilla Nordin , Nilsson Jan
Published: June 2011

Abstract
—Activation of vascular inflammation in response to hyperlipidemia is believed to play an important role during the early stages of atherogenesis. We demonstrate here that exposure of cultured, rat aortic smooth muscle cells to low density lipoprotein (LDL) stimulated tumor necrosis factor-α (TNF-α) mRNA and protein expression. Oxidative modification of LDL resulted in a reduction of this stimulatory effect. To analyze whether a similar response also occurs in vivo, we used a recently developed model in which the effects of a rapid accumulation of human LDL in rat arteries can be studied. As previously reported, epitopes specific for human apolipoprotein B began to accumulate in the aorta within 2 to 6 hours after injection of 6 mg of human LDL. This was followed by expression of oxidized LDL–specific epitopes after 12 hours. There was no vascular expression of TNF-α at baseline or in phosphate-buffered saline–injected control rats. However, 24 hours after injection of native LDL, there was a marked induction of TNF-α mRNA and immunoreactivity in the aorta and other large arteries, whereas injection of oxidized LDL was without effect in this respect. Preincubation of LDL with the antioxidant probucol before injection markedly decreased the expression of TNF-α immunoreactivity. The present findings support the notion that LDL may activate arterial expression of TNF-α and suggest 1 possible mechanism for the inflammatory response in the early stages of atherosclerosis. The role of LDL oxidation in this process remains to be fully elucidated.
Electronegative LDL From Normolipemic Subjects Induces IL-8 and Monocyte Chemotactic Protein Secretion by Human Endothelial Cells
Authors: Conxita De Castellarnau  , Sánchez-Quesada José Luis , Benítez Sonia , Rosa Roser , Caveda Luis , Vila Luis , Ordóñez-Llanos Jordi
Published: June 2011

Abstract
—The presence in plasma of an electronegative LDL subfraction [LDL(−)] cytotoxic for endothelial cells (ECs) has been reported. We studied the effect of LDL(−) on the release by ECs of molecules implicated in leukocyte recruitment [interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1)] and in the plasminogen activator inhibitor-1 (PAI-1). LDL(−), isolated by anion-exchange chromatography, differed from nonelectronegative LDL [LDL(+)] in its higher triglyceride, nonesterified fatty acid, apoprotein E and apoprotein C-III, and sialic acid contents. No evidence of extensive oxidation was found in LDL(−); its antioxidant and thiobarbituric acid–reactive substances contents were similar to those of LDL(+). However, conjugated dienes were increased in LDL(−), which suggests that mild oxidation might affect these particles. LDL(−) increased, in a concentration-dependent manner, the release of IL-8 and MCP-1 by ECs and was a stronger inductor of both chemokines than oxidized LDL (oxLDL) or LDL(+). PAI-1 release increased slightly in ECs incubated with both LDL(−) and oxLDL but not with LDL(+). However, no cytotoxic effects of LDL(−) were observed on ECs. Actinomycin D inhibited the release of IL-8 and MCP-1 induced by LDL(−) and oxLDL by up to 80%, indicating that their production is mediated by protein synthesis. Incubation of ECs with
N
-acetyl cysteine inhibited production of IL-8 and MCP-1 induced by LDL(−) and oxLDL by >50%. The free radical scavenger butylated hydroxytoluene slightly inhibited the effect of oxLDL but did not modify the effect of LDL(−). An antagonist (BN-50730) of the platelet-activating factor receptor inhibited production of both chemokines by LDL(−) and oxLDL in a concentration-dependent manner. Our results indicate that LDL(−) shows proinflammatory activity on ECs and may contribute to early atherosclerotic events.
Platelet Glycoprotein IIb/IIIa Receptor Inhibitor Preserves Coronary Flow Reserve During Progressive Coronary Arteriostenosis in Swine
Authors: Shen You-Tang , Wiedmann Richard T. , Lynch Joseph J. , Gould Robert J.
Published: June 2011

Abstract
—Thrombosis resulting from blood platelet aggregation via glycoprotein (GP) IIb/IIIa receptor activation triggers the local release of vasoactive substances. Therefore, inhibition of these receptors could affect coronary vasoactive function during thrombotic coronary arteriostenosis. Twenty pigs were instrumented with an aortic catheter and with hydraulic occluders and flow probes on both the left anterior descending (LAD) and the left circumflex (LCx) coronary arteries. One of these 2 coronary arteries was repeatedly injured by external clamping for 15-second periods at 30-minute intervals while the pigs were given either a GP IIb/IIIa receptor inhibitor (L-739,758) (n=5), heparin (n=5), aspirin (n=3), or saline (n=7). There were no baseline differences between the 4 groups in mean arterial pressure, resting coronary blood flow (CBF), or reactive hyperemic response (RHR), which was induced by brief coronary artery occlusion and expressed as flow debt repayment. After multiple injuries, resting CBF had decreased by 95±2% (ie, nearly complete coronary artery occlusion) at 15±4 minutes in the control group, whereas in the heparin-, aspirin-, and GP IIb/IIIa inhibitor–treated groups, resting CBF had decreased by only 21±7% at 18±3 minutes, 15±3% at 18±5 minutes, and 15±7% at 21±4 minutes, respectively, suggesting that heparin, aspirin, and the GP IIb/IIIa inhibitor each prevented injury-induced coronary artery occlusion. After the initial injury, the RHR was progressively reduced in the control and heparin- and aspirin-treated groups but not in the GP IIb/IIIa inhibitor–treated group. At a comparable level of resting CBF (≈15% below baseline), the RHR was reduced more in the control (−56±9%), heparin-treated (−49±9%), and aspirin-treated (−61±12) groups (
P
<0.05) than in the GP IIb/IIIa inhibitor–treated group (−26±6%). When the resting CBF had decreased by ≈35%, the RHR still was reduced significantly more (
P
<0.01) in the heparin-treated group (−64±9%) than in the GP IIb/IIIa inhibitor–treated group (−21±6%). In a separate group of control pigs (n=4) subjected to 2 injuries, coronary perfusion pressure distal to the injury site was reduced by 14±1 mm Hg from the arterial pressure, and the RHR was 20±6%. When the distal coronary perfusion pressure was reduced similarly (−14±1 mm Hg) in a separate group of GP IIb/IIIa inhibitor–treated pigs (n=4) by 2 injuries and the use of a hydraulic occluder, the RHR was 130±16% (
P
<0.01 versus control). Our data demonstrate for the first time that a platelet GP IIb/IIIa receptor inhibitor can preserve the distal coronary vasodilatory response during progressive coronary arteriostenosis.
Bezafibrate Increases Preβ1-HDL at the Expense of HDL
2b
in Hypertriglyceridemia
Authors: Miida Takashi , Sakai Katsuyuki , Ozaki Kazuyuki , Nakamura Yuichi , Yamaguchi Toshio , Tsuda Takashi , Kashiwa Takayuki , Murakami Toru , Inano Koichi , Okada Masahiko
Published: June 2011

Abstract

—Preβ1-high density lipoprotein (preβ1-HDL), the initial acceptor of cell-derived cholesterol, can be generated from HDL
2
by hepatic lipase. Because bezafibrate elevates lipase activity, it may increase preβ1-HDL at the expense of HDL
2
. To answer this question, we determined the apolipoprotein A-I (apoA-I) distribution in 20 hypertriglyceridemics (triglycerides>2.26 mmol/L) and 20 sex-matched normolipidemics by native 2-dimensional gel electrophoresis. At baseline, preβ1-HDL was 70% higher in hypertriglyceridemics than in normolipidemics (123.5±49.9 versus 72.5±34.1 mg/L apoA-I,
P
<0.01). Preβ1-HDL was positively correlated with triglyceride (
r
=0.624,
P
<0.0001). A 4-week bezafibrate treatment (400 mg daily) increased preβ1-HDL by 30% (160.2±64.5 mg/L apoA-I,
P
<0.05) but decreased HDL
2b
by 31% (from 188.8±94.9 to 129.3±78.7 mg/L apoA-I,
P
<0.05). Hepatic lipase activity increased by 24% (
P
<0.005). Preβ1-HDL was generated either from ultracentrifugally isolated HDL
2
or from plasma during incubation with triglyceride lipase. In conclusion, bezafibrate increases preβ1-HDL at the expense of HDL
2
. We speculate that such an effect might partly contribute to the antiatherogenic action of bezafibrate.
P-Selectin–Dependent Inhibition of Thrombosis During Venous Stasis
Authors: Eppihimer Michael J. , Schaub Robert G.
Published: June 2011

Abstract—Leukocyte adhesion, transendothelial migration, and stasis are important components in the pathogenesis of deep vein thrombosis. Anesthetized cats were treated with saline, a recombinant soluble form of P-selectin glycoprotein ligand-1 (rPSGL-Ig), or an E- and L-selectin antibody (EL-246) before exposure and occlusion of a jugular vein. After 2 or 6 hours of occlusion, jugular veins were perfused with buffer, fixed, and prepared for scanning electron microscopy. In cats receiving saline, 2 and 6 hours of occlusion produced moderate levels of leukocyte and platelet adhesion and endothelial cell injury. Treatment of cats with rPSGL-Ig or EL-246 had no apparent effect on the magnitude of cell adhesion and endothelial cell injury compared with no treatment. After 6 hours of occlusion, the presence of a mural thrombus in untreated veins was observed and confirmed by scanning electron microscopy. Pretreatment of cats with rPSGL-Ig completely (4.0 mg/kg) or partially (1.0 mg/kg) prevented the occurrence of thrombi in the jugular veins. The reduction in thrombosis by rPSGL-Ig treatment after 6 hours of venous stasis, in the absence of any effect on leukocyte-mediated endothelial cell injury, suggests an antithrombotic mechanism of action for this protein.
Protein Kinase A–Dependent Stimulation of Rat Type II Secreted Phospholipase A
2
Gene Transcription Involves C/EBP-β and -δ in Vascular Smooth Muscle Cells
Authors: Couturier Cyril , Antonio Valérie , Brouillet Arthur , Béréziat Gilbert , Raymondjean Michel , Andréani Marise
Published: June 2011

Abstract
—Type II secreted phospholipase A
2
(sPLA
2
) releases precursors of important inflammatory lipid mediators from phospholipids. Some observations have indicated that the sPLA
2
, which has been implicated in chronic inflammatory conditions such as arthritis, contributes to atherosclerosis in the arterial wall. sPLA
2
was not detected in control vascular smooth muscle cells (VSMC). Treatment of VSMC with agents that increase intracellular cAMP (eg, forskolin, dibutyryl [db]-cAMP) resulted in a time- and concentration-dependent increase in sPLA
2
gene expression. Semiquantitative reverse transcriptase polymerase chain reaction (RT-PCR) showed a marked dose-dependent inhibition of forskolin-induced mRNA by protein kinase A inhibitor. Electrophoretic mobility shift analysis of nuclear proteins from forskolin-treated and db–cAMP-treated VSMC with C/EBP consensus oligonucleotides and C/EBP oligonucleotides from the rat promoter revealed greater binding than in control VSMC. Incubation of VSMC with H89, a specific protein kinase inhibitor, also blocked the binding of nuclear C/EBP to the C/EBP site of the rat promoter induced by db-cAMP and forskolin. Binding was unchanged with the use of CRE consensus oligonucleotides. Antibodies revealed the specific formation of C/EBP/DNA complexes, the majority of which were supershifted by C/EBP-β and -δ antibodies. Functional activation of C/EBP was confirmed by a luciferase reporter gene assay. A construct comprising 4 tandem repeat copies of the C/EBP element from the rat sPLA
2
promoter linked to luciferase was transcriptionally activated in VSMC by cotransfection with expression vector for the protein kinase A catalytic subunit. It was also significantly activated in transfected VSMC treated by forskolin or db-cAMP. H89 inhibited this activations. We therefore conclude that the increases in sPLA
2
mRNA and enzyme activity produced by cAMP-elevating agents is controlled by a mechanism involving nuclear C/EBP-β and -δ acting through a protein kinase A signaling pathway.
Hyperhomocysteinemia Impairs Angiogenesis in Response to Hindlimb Ischemia
Authors: Duan Junli , Murohara Toyoaki , Ikeda Hisao , Sasaki Ken-ichiro , Shintani Satoshi , Akita Takako , Shimada Toshifumi , Imaizumi Tsutomu
Published: June 2011

Abstract
—Hyperhomocysteinemia (HH) is an independent risk factor for atherosclerosis, including peripheral arterial occlusive disease (PAOD). Because angiogenesis and collateral vessel formation are important self-salvage mechanisms for ischemic PAOD, we examined whether HH modulates angiogenesis in vivo in a rat model of hindlimb ischemia. Rats were divided into 3 groups: the control group was given tap water, the HH group was given water containing l-methionine (1 g · kg
−1
· d
−1
), and the HH+L-arg group was given water containing methionine (1 g · kg
−1
· d
−1
) and l-arginine (2.25 vol%). At day 14 of the dietary modifications, the left femoral artery and vein were excised, and the extent of angiogenesis and collateral vessels in the ischemic limb were examined for 4 weeks. Plasma homocysteine levels significantly increased (
P
<0.001), and plasma and tissue contents of nitrite+nitrate as well as tissue cGMP levels significantly decreased in the HH group compared with the control group (
P
<0.01). Laser Doppler blood flowmetry (LDBF) revealed a significant decrease in the ischemic/normal limb LDBF ratio in the HH group at days 7, 14, 21, and 28 (
P
<0.01 versus control). Angiography revealed a significant decrease in the angiographic score in the HH group at day 14 (
P
<0.001 versus control). Immunohistochemistry of ischemic tissue sections showed a significant reduction in the capillary density in the HH group (
P
<0.001 versus control). Oral l-arginine supplementation in rats with HH (HH+L-arg) restored the decreased plasma and tissue nitrite+nitrate and cGMP contents (
P
<0.05) as well as angiogenesis, as assessed by LDBF (
P
<0.05 versus HH), angiographic score (
P
<0.01 versus HH), and capillary density (
P
<0.001 versus HH). In summary, HH impaired ischemia-induced angiogenesis and collateral vessel formation in a rat model of hindlimb ischemia in vivo. The mechanism of the HH-induced impairment of angiogenesis might be mediated in part by a reduced bioactivity of endogenous NO in the HH state.