bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of Signal Properties in Atherosclerotic Plaque Components by Intravascular MRI
Authors: Rogers Walter J. , Prichard Jeffrey W. , Hu Yong-Lin , Olson Peter R. , Benckart Daniel H. , Kramer Christopher M. , Vido Diane A. , Reichek Nathaniel
Published: June 2011

Abstract
—Magnetic resonance imaging (MRI) is capable of distinguishing between atherosclerotic plaque components solely on the basis of biochemical differences. However, to date, the majority of plaque characterization has been performed by using high-field strength units or special coils, which are not clinically applicable. Thus, the purpose of the present study was to evaluate MRI properties in histologically verified plaque components in excised human carotid endarterectomy specimens with the use of a 5F catheter–based imaging coil, standard acquisition software, and a clinical scanner operating at 0.5 T. Human carotid endarterectomy specimens from 17 patients were imaged at 37°C by use of an opposed solenoid intravascular radiofrequency coil integrated into a 5F double-lumen catheter interfaced to a 0.5-T General Electric interventional scanner. Cross-sectional intravascular MRI (156×250 μm in-plane resolution) that used different imaging parameters permitted the calculation of absolute T1and T2, the magnetization transfer contrast ratio, the magnitude of regional signal loss associated with an inversion recovery sequence (inversion ratio), and regional signal loss in gradient echo (gradient echo–to–spin echo ratio) in plaque components. Histological staining included hematoxylin and eosin, Masson’s trichrome, Kossa, oil red O, and Gomori’s iron stain. X-ray micrographs were also used to identify regions of calcium. Seven plaque components were evaluated: fibrous cap, smooth muscle cells, organizing thrombus, fresh thrombus, lipid, edema, and calcium. The magnetization transfer contrast ratio was significantly less in the fibrous cap (0.62±13) than in all other components (
P
<0.05) The inversion ratio was greater in lipid (0.91±0.09) than all other components (
P
<0.05). Calcium was best distinguished by using the gradient echo–to–spin echo ratio, which was lower in calcium (0.36±0.2) than in all plaque components, except for the organizing thrombus (
P
<0.04). Absolute T1 (range 300±140 ms for lipid to 630±321 ms for calcium) and T2 (range 40±12 ms for fresh thrombus to 59±21 ms for smooth muscle cells) were not significantly different between groups. In vitro intravascular MRI with catheter-based coils and standard software permits sufficient spatial resolution to visualize major plaque components. Pulse sequences that take advantage of differences in biochemical structure of individual plaque components show quantitative differences in signal properties between fibrous cap, lipid, and calcium. Therefore, catheter-based imaging coils may have the potential to identify and characterize those intraplaque components associated with plaque stability by use of existing whole-body scanners.
Nov
Gene Encodes Adhesion Factor for Vascular Smooth Muscle Cells and Is Dynamically Regulated in Response to Vascular Injury
Authors: Ellis Peter D. , Chen Qin , Barker Patrick J. , Metcalfe James C. , Kemp Paul R.
Published: June 2011

Abstract
—Nephroblastoma overexpressed (NOV) is a member of the CCN family (connective tissue growth factor, CYR61, and NOV) of proteins that are involved in regulating the proliferation, differentiation, and adhesion of a variety of cell types. We have examined the expression of the
Nov
gene and NOV protein by vascular smooth muscle cells (VSMCs), in vitro and in vivo, and the effects of recombinant NOV on VSMCs. Rat aortic VSMCs were found to express
Nov
mRNA and NOV protein in vitro and in vivo.
Nov
expression in adult rat tissues was very high in the aorta and was detected only weakly in the brain and lung by Northern analysis (relative levels 33:3:1). During postnatal development (3 days to 12 weeks), the expression of
Nov
was correlated with markers of the differentiated smooth muscle cell phenotype (smooth muscle myosin heavy chain and SM22α). In the rat carotid artery balloon injury model,
Nov
was detectable by in situ hybridization and was downregulated in the media of the injured artery compared with the uninjured artery at 7 and 14 days after injury. Expression in the developing intima was barely detectable at 7 days after injury except for strong expression at the luminal surface. At 14 days after injury,
Nov
expression was substantially increased throughout the intima. In vitro studies of the function of NOV protein showed that it promoted VSMC adhesion via a mechanism that was divalent cation and Arg-Gly-Asp independent but that it did not modulate VSMC proliferation or phenotype. The strong expression and dynamic regulation of
Nov
in the arterial wall, together with its ability to promote VSMC adhesion, suggest that it may be involved in homeostasis and repair.
Polymerase Chain Reaction–Based Method for Quantifying Recruitment of Monocytes to Mouse Atherosclerotic Lesions In Vivo
Authors: Kim Chee-Jeong , Khoo John C. , Gillotte-Taylor Kristin , Li Andrew , Palinski Wulf , Glass Christopher K. , Steinberg Daniel
Published: June 2011

Abstract
—The critical role of monocyte recruitment in atherogenesis has been appreciated for some time. However, until recently, there have been no sufficiently sensitive methods for measuring rates of monocyte recruitment to the arterial wall in vivo. We have developed a novel highly sensitive method, based on the polymerase chain reaction, for quantitatively tracking DNA-marked monocytes and have adapted it for use in mice. We use the uniquely male gene,
Sry
, on the Y chromosome as a gene marker. We transfuse monocytes from a male donor into a congenic female mouse, euthanize the mouse after 24 to 48 hours, and then quantify the arterial uptake of monocytes by quantitative polymerase chain reaction. This study describes the techniques used and their sensitivity and reproducibility and demonstrates the approach by assessing the effects of cytokines. In control low density lipoprotein receptor–negative mice, monocyte recruitment decreased slightly but significantly as lesions progressed. Intraperitoneal injection of a combination of tumor necrosis factor-α and interleukin-1β more than doubled the rate of monocyte recruitment into developing lesions. However, the response to the cytokines was much greater in younger mice with less advanced lesions than in older animals with more advanced lesions.
Blood Pressure, LDL Cholesterol, and Intima-Media Thickness
Authors: Sun Ping , Dwyer Kathleen M. , Merz C. Noel Bairey , Sun Wei , Johnson C. Anderson , Shircore Anne M. , Dwyer James H.
Published: June 2011

Abstract
—The “response to injury” hypothesis is a plausible model of the development of atherosclerosis supported by observations from animal models. The present study uses epidemiological data to investigate the hypothesis that wall damage due to hypertension is a precursor of low density lipoprotein cholesterol (LDL-C)–mediated atherosclerosis. The Los Angeles Atherosclerosis Study is following a cohort of 576 participants who were aged 40 to 60 years and were free of symptomatic cardiovascular disease at recruitment. Common carotid artery intima-media thickness (IMT) was assessed by B-mode ultrasonography. After exclusion for nonfasting blood draw and other missing data, 511 subjects were available for analysis. IMT was regressed on LDL-C within tertiles of systolic blood pressure (SBP): low (93 to 122 mm Hg), middle (123 to 132 mm Hg), and high (133 to 175 mm Hg). Covariates were age, sex, body height, body mass index, ethnicity, smoking status, diabetes, and pharmacological treatment for hypertension or hypercholesterolemia. IMT was significantly related to LDL-C in the high SBP group (β=0.025±0.008, where β values are IMT [mm]/LDL-C [mmol/L];
P
=0.002) but not in the middle (β=−0.006±0.008,
P
=0.39) or low (β=−0.004±0.009,
P
=0.64) SBP group. The slope in the high SBP group was significantly greater than in the middle (
P
=0.004) or low (
P
=0.014) SBP group. Results were similar for women and men, and after the exclusion of diabetics and persons using antihypertensive or lipid-lowering medications. Elevated LDL-C was associated with increased IMT in the upper tertile of SBP but not in the lower tertiles. These findings are consistent with the hypothesis that wall injury due to elevated SBP increases the susceptibility of the artery wall to LDL-C–mediated atherogenesis.
The Metabolic Syndrome, LDL Particle Size, and Atherosclerosis
Authors: Hulthe Johannes , Bokemark Lena , Wikstrand John , Fagerberg Björn
Published: June 2011

Abstract
—An operative definition of the metabolic syndrome has been suggested by a working group associated with the World Health Organization in 1998. The aim of this study was to examine whether small, low density lipoprotein (LDL) particle size was associated with the metabolic syndrome and with subclinical atherosclerosis as measured by ultrasound in the carotid and femoral arteries. The study was performed in a population-based sample of clinically healthy men (N=391), all 58 years old and not undergoing any treatment with cardiovascular drugs. Exclusion criteria were cardiovascular or other clinically overt diseases or continuous medication with cardiovascular drugs. The results showed that subjects characterized by the metabolic syndrome (n=62) had a thicker mean intima-media complex (IMT) in both the carotid and femoral arteries (0.86 versus 0.77 mm,
P
<0.001, and 1.03 versus 1.00 mm,
P
=0.022, respectively) and also lower mean values for LDL particle size (25.78 versus 26.80 nm, respectively,
P
<0.001) compared with subjects with no risk factors (n=77). The group with the metabolic syndrome (n=62) also had higher mean values for serum cholesterol and heart rate. In the whole study group (N=391), there were significant but weak negative relationships between small LDL particle size, increasing IMT, and increasing cross-sectional intima-media area of the carotid and femoral arteries and also negative relationships between LDL particle size and plaque occurrence and size in the carotid and femoral arteries. In summary, this is the first large-scale study to demonstrate a relationship between the clustering of risk factors that constitute the metabolic syndrome and a small LDL particle size pattern and the occurrence of preclinical atherosclerosis in the carotid and femoral arteries, as assessed by the ultrasound technique, in healthy 58-year-old men recruited from the general population.
Overexpression of Inducible Heat Shock Protein 70 in COS-1 Cells Fails to Protect From Cytotoxicity of Oxidized LDLs
Authors: Pirillo A. , Norata G. D. , Zanelli T. , Catapano A. L.
Published: June 2011

Abstract
—Oxidized low density lipoproteins (OxLDLs) are believed to play a central role in atherogenesis and to possess a wide variety of biological properties; among them, OxLDLs are cytotoxic to cultured vascular cells in that they induce necrosis and apoptosis. Moreover, OxLDLs are known to induce the expression of heat shock protein 70 (Hsp70), a protein that protects cells from several cytotoxic stimuli. To determine whether Hsp70 can protect cells against OxLDL-induced cytotoxicity, COS-1 cells were transfected with a construct containing human Hsp70. A number of cell lines permanently expressing Hsp70 were obtained, 1 of which (
cos-Hsp70/10
, with high Hsp70 expression) was selected for further studies. Hsp70 overexpression protected cells from toxic stimuli, such as H
2
O
2
, UV irradiation, and heat shock, suggesting that the overexpressed protein was functional. When incubated with OxLDLs, however, the clone overexpressing Hsp70 showed a significant decrease in viability, as determined by the [
3
H]adenine release assay (319.8±3.16% of control for transfected cells versus 217.6±6.08% for control cells exposed to 100 μg protein/mL of OxLDL), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (12.5±0.9% versus 28.9±1.99% of control, respectively), and LDH release (48.4±0.04% versus 15.2±0.06% of control cells). The increased expression of
Bax
and the decreased expression of
Bcl-2
(a proapoptotic and an antiapoptotic protein, respectively) in
cos-Hsp70/10
cells and in control cells on incubation with OxLDLs suggested that overexpression of Hsp70 did not confer protection against apoptosis induced by OxLDLs. The analysis of nucleosome content and the nuclear staining with Hoechst 33258 confirmed this finding. These data suggest that overexpression of Hsp70 not only fails to protect COS-1 cells against OxLDL-induced apoptosis but rather confers a higher sensitivity to the cytotoxic action of these lipoproteins. Thus, the Hsp70 response, although induced by OxLDLs, cannot protect cells from lipoprotein toxicity.
VEGF Protects Against Oxidized LDL Toxicity to Endothelial Cells by an Intracellular Glutathione-Dependent Mechanism Through the KDR Receptor
Authors: Kuzuya Masafumi , Ramos Miguel A. , Kanda Shigeru , Koike Teruhiko , Asai Toshinobu , Maeda Keiko , Shitara Kenya , Shibuya Masabumi , Iguchi Akihisa
Published: June 2011

Abstract
—Although the accumulation of vascular endothelial growth factor (VEGF) has been observed in human atherosclerotic lesions, the exact role of this growth factor in atherogenesis remains unknown. We hypothesized that VEGF in the vascular wall might have a preventive effect on endothelial cell damage during atherosclerosis. To test our hypothesis, we examined whether VEGF protects against the toxicity of oxidized low density lipoprotein (Ox-LDL) in cultured endothelial cells derived from bovine aortas (BAECs). Preincubation of BAECs with VEGF prevented Ox-LDL–induced toxicity in a preincubation time– and VEGF concentration–dependent manner. Addition of
N

ω

-nitro-
l
-arginine methyl ester, a nitric oxide synthase inhibitor, did not reverse the protective effect of VEGF on Ox-LDL toxicity. Incubation of BAECs with VEGF increased intracellular glutathione (GSH) content in a time-dependent manner. Combined addition of VEGF and
l
-buthionine sulfoximine, a GSH synthesis inhibitor, reversed both GSH levels and the protective effect of VEGF on Ox-LDL–induced cytotoxicity. Placenta growth factor, which ligates to the VEGF Flt-1 receptor but not KDR/Flk-1, failed to prevent Ox-LDL toxicity and had no effect on intracellular GSH levels. An anti-KDR antibody completely blocked these beneficial activities of VEGF. These results suggest that VEGF prevents Ox-LDL–induced endothelial cell damage via an intracellular GSH-dependent mechanism through the KDR/Flk-1 receptor.
Identification of a γ-Interferon–Responsive Element in the Promoter of the Human Macrophage Scavenger Receptor A Gene
Authors: Grewal Thomas , Priceputu Elena , Davignon Jean , Bernier Lise
Published: June 2011

Abstract
—In the present study, we demonstrate γ-interferon (γ-IFN)–inducible scavenger receptor A (SR-A) mRNA expression during the early stages of THP-1 and blood monocyte differentiation. Predominant induction of SR-A type II mRNA parallels the increased accumulation of cholesteryl esters under these conditions. A potential signal transducer and activator of transcription (STAT1) binding site (γ-interferon activation site) in the SR-A promoter demonstrates γ-IFN–inducible DNA binding activity and is most likely responsible for the γ-IFN–dependent expression of an SR-A promoter–luciferase fusion construct. In contrast, γ-IFN inhibits SR-A expression in mature macrophages as well as after prolonged γ-IFN incubation of THP-1 monocytes. Taken together, these results demonstrate opposite effects of γ-IFN on SR-A expression and activity during the early versus late stages of monocyte maturation. γ-IFN–induced STAT1 activation, leading to increased SR-A expression, could therefore play an important role in the initial steps of foam cell formation and xanthomatosis.
Serum C3 but Not Plasma Acylation-Stimulating Protein Is Elevated in Finnish Patients With Familial Combined Hyperlipidemia
Authors: Ylitalo Kati , Pajukanta Päivi , Meri Seppo , Cantor Rita M. , Mero-Matikainen Niina , Vakkilainen Juha , Nuotio Ilpo , Taskinen Marja-Riitta
Published: June 2011

Abstract
—A trapping defect of fatty acids due to impaired function of acylation-stimulating protein (ASP) has been suggested as one mechanism underlying the metabolic abnormalities in familial combined hyperlipidemia (FCHL). The study aimed at defining the role of ASP and complement C3 in 35 Finnish FCHL families. There was no difference in plasma ASP levels between the 66 hypertriglyceridemic FCHL patients and their 84 normotriglyceridemic relatives. No response in plasma ASP could be observed after a fatty meal in 10 FCHL patients or in 10 control subjects. In familial correlation analyses, C3 exhibited a significant sibling-sibling correlation. The FCHL patients had higher serum C3 levels than their unaffected relatives (
P
<0.001). Furthermore, serum C3 levels correlated significantly with several lipid parameters. The correlations between ASP and lipid variables were weaker than those of C3. These analyses suggest that common genes might contribute to the regulation of serum C3, triglycerides, HDL-C, free fatty acids, and insulin. The present data do not support the hypothesis that defects of the ASP pathway are reflected in plasma lipoproteins or in impaired plasma lipid clearance postprandially.
Critical Role of Rho-Kinase and MEK/ERK Pathways for Angiotensin II–Induced Plasminogen Activator Inhibitor Type-1 Gene Expression
Authors: Takeda Kotaro , Ichiki Toshihiro , Tokunou Tomotake , Iino Naoko , Fujii Satoshi , Kitabatake Akira , Shimokawa Hiroaki , Takeshita Akira
Published: June 2011

Abstract
—Plasminogen activator inhibitor type-1 (PAI-1) plays an integral role not only in the regulation of fibrinolytic activity but also in the pathogenesis of atherosclerosis and hypertension. We investigated the signaling pathways of angiotensin II (Ang II) leading to PAI-1 gene expression. Ang II increased the PAI-1 mRNA and protein levels in a time- and dose-dependent manner through the Ang II type 1 receptor in vascular smooth muscle cells. PAI-1 gene promoter activity measured by luciferase assay was significantly increased by Ang II. PAI-1 mRNA stability was also increased by Ang II. Ang II–induced PAI-1 mRNA upregulation was inhibited by BAPTA-AM, genistein, and AG1478, suggesting that intracellular calcium, tyrosine kinase, and epidermal growth factor receptor transactivation are involved. Furthermore, PD98059, an inhibitor of extracellular signal–regulated kinase (ERK) kinase (MEK), almost completely suppressed Ang II–induced PAI-1 upregulation. Adenovirus-mediated overexpression of the dominant-negative form of Rho-kinase or Y27632, a Rho-kinase inhibitor, also completely prevented PAI-1 induction by Ang II without affecting Ang II–induced ERK activation. These data suggest that activation of MEK/ERK and Rho-kinase pathways plays a pivotal role in PAI-1 gene upregulation by Ang II. The Rho-kinase pathway may be a novel target to inhibit Ang II signaling, and its inhibition may be useful in the treatment of hypertension as well as atherosclerosis.
Plasma Fibrinogen as a Predictor of Total and Cause-Specific Mortality in Elderly Japanese-American Men
Authors: Yano Katsuhiko , Grove John S. , Chen Randi , Rodriguez Beatriz L. , Curb J. David , Tracy Russell P.
Published: June 2011

Abstract
—The relation between plasma fibrinogen and total and cause-specific mortality was investigated in a cohort of 3571 Japanese-American men aged 71 to 93 years during a median follow-up of 4.4 years. There were a total of 728 deaths, of which 37% were accounted for by cardiovascular disease and 27% by cancer. The age-adjusted relative risk (RR) for total mortality in the top quintile of fibrinogen (>3.51 g/L) compared with the bottom quintile (<2.57 g/L) was 4.3 (
P
<0.0001) in the first year of follow-up. RR was reduced to 1.7 in the second year but remained significantly and slightly increased in subsequent years. After adjustment for age and confounding risk factors, the RRs (and 95% confidence intervals) associated with a 1-SD increment of fibrinogen (0.64 g/L) for all-cause, cardiovascular disease, cancer, and other-cause mortality were 1.3 (1.2 to 1.4), 1.2 (1.1 to 1.4), 1.3 (1.2 to 1.5), and 1.3 (1.2 to 1.5), respectively. Preexisting diseases did not influence the significant association of fibrinogen with mortality. There was a significant interaction of fibrinogen with white blood cell count but not with cigarette smoking. We conclude that plasma fibrinogen is an independent risk factor for mortality from a broad spectrum of diseases in elderly men and that this universal effect of fibrinogen on mortality may be mediated partly through inflammation.
Adrenomedullin Inhibits Angiotensin II–Induced Expression of Tissue Factor and Plasminogen Activator Inhibitor-1 in Cultured Rat Aortic Endothelial Cells
Authors: Sugano T. , Tsuji H. , Masuda H. , Nishimura H. , Yoshizumi M. , Kawano H. , Kimura S. , Ukimura N. , Yano S. , Kunieda Y. , Nakagawa K. , Nakagawa M.
Published: June 2011

Abstract
—Adrenomedullin (AM) is a potent vasodilating peptide having a variety of pharmacological properties mainly in respect to vascular pathophysiology. We have previously demonstrated that angiotensin II (Ang II) or natriuretic peptides have influence on the expression of tissue factor (TF) and plasminogen activator inhibitor-1 (PAI-1) in vascular endothelial cells. The aim of this study was to elucidate the effects of AM on TF and PAI-1 mRNA and protein expression in endothelial cells. As a result, AM inhibited Ang II–induced TF and PAI-1 mRNA expression in a dose- and time-dependent manner. Because the expression of TF and PAI-1 mRNA induced by Ang II was attenuated by the increase of intracellular concentrations of cAMP by forskolin and 8-bromo-cAMP and because AM increased the intracellular level of cAMP in rat aortic endothelial cells, it was indicated that the inhibitory effect of AM on the expressions of TF and PAI-1 was mainly mediated by the cAMP-dependent signal transduction. Furthermore, the inhibitory effect of AM on TF and PAI-1 expression was partly attenuated by an NO synthase inhibitor,
N
G
-nitro-
l
-arginine methyl ester. In conclusion, AM is shown to contribute to the regulation of blood coagulation and fibrinolysis by vascular endothelial cells mainly via the cAMP pathway.
2000 George Lyman Duff Memorial Lecture
Authors: Davis Roger A. , Hui To Y.
Published: June 2011

Abstract
—The production of apolipoprotein B (apoB)–containing lipoproteins by the liver is regulated by a complex series of processes involving apoB being cotranslationally translocated across the endoplasmic reticulum and assembled into a lipoprotein particle. The translocation of apoB across the endoplasmic reticulum is facilitated by the intraluminal chaperone, microsomal triglyceride transfer protein (MTP). MTP facilitates the translocation and folding of apoB, as well as the addition of lipid to lipid-binding domains (which consist of amphipathic β sheets and α helices). In the absence of MTP or sufficient lipid, apoB exhibits translocation arrest. Thus, apoB translation, translocation, and assembly with lipids to form a core-containing lipoprotein particle occur as concerted processes. Abrogation of ≥1 of these processes diverts apoB into a degradation pathway that is dependent on conjugation with ubiquitin and proteolysis by the proteasome. The nascent core-containing lipoprotein particle that forms within the lumen of the endoplasmic reticulum can be “enlarged” to form a mature very low density lipoprotein particle. Additional studies show that the assembly and secretion of apoB-containing lipoproteins are linked to the cholesterol/bile acid synthetic pathway controlled by cholesterol 7α-hydroxylase. Studies in cultured cells and transgenic mice indicate that the expression of cholesterol 7α-hydroxylase indirectly regulates the expression of lipogenic enzymes through changes in the cellular content of mature sterol response element binding proteins. Oxysterols and bile acids may also act via the ligand-activated nuclear receptors LXR and FXR to link the metabolic pathways controlling energy balance and lipid metabolism to nutritional state.
Genome-Wide Linkage Analysis Reveals Evidence of Multiple Regions That Influence Variation in Plasma Lipid and Apolipoprotein Levels Associated With Risk of Coronary Heart Disease
Authors: Klos Kathy L. , Kardia Sharon L. R. , Ferrell Robert E. , Turner Stephen T. , Boerwinkle Eric , Sing Charles F.
Published: June 2011

Abstract
—Results of genome-wide linkage analyses to identify chromosomal regions that influence interindividual variation in plasma lipid and apolipoprotein levels in the Rochester, Minn, population are reported. Analyses were conducted for total cholesterol (total-C), triglycerides (TGs), high density lipoprotein cholesterol (HDL-C), apolipoprotein A-I, apolipoprotein A-II, apolipoprotein B, apolipoprotein C-II, apolipoprotein C-III, apolipoprotein E, the total-C/HDL-C ratio, and the TG/HDL-C ratio. Genotypes were measured for 373 genome-wide marker loci on 1484 individuals distributed among 232 multigeneration pedigrees sampled without regard to health status. LOD scores and estimates of additive genetic variance associated with map locations were obtained by using the variance-component method of linkage analysis. No evidence of linkage with genes influencing variation in age served as a negative control. Plasma apolipoprotein E levels and the
apolipoprotein E
gene served as a positive control (LOD score 4.20). Evidence (LOD score >2.00) was provided that was suggestive of a gene or genes on chromosomes 4 and 5 influencing variation in the apolipoprotein A-II level, on chromosome 12 influencing variation in the apolipoprotein A-I level, and on chromosome 17 influencing variation of total-C/HDL-C. These analyses provide new information about genomic regions in humans that influence interindividual variation in plasma lipid and apolipoprotein levels and serve as a basis for further fine-mapping studies to identify new genes involved in lipid metabolism.
Cellular pathology of progressive atherosclerosis in the WHHL rabbit. An animal model of familial hypercholesterolemia.
Authors: Buja L M , Kita T , Goldstein J L , Watanabe Y , Brown M S
Published: June 2011

This report describes the features of developing atherosclerosis in the Watanabe heritable hyperlipidemic (WHHL) rabbit, an animal model of human familial hypercholesterolemia. Observations were made in 18 homozygous WHHL rabbits, aged 4 days to 15 months, fed standard rabbit chow; seven control New Zealand white rabbits fed a similar diet, and four New Zealand white rabbits fed rabbit chow containing 2% cholesterol and 10% corn oil for 2 weeks. The WHHL rabbits showed evidence of progressive disease of the aorta with accumulation of strongly birefringent lipid in intimal lesions, including fatty streaks, raised foam cell lesions, and plaques (atheromas), as well as in the media. As seen by electron microscopy, the cellular population of the intimal lesions consisted predominantly of smooth muscle cells with lipid deposits and lipid-laden foam cells. Lipid deposits occurred as cytoplasmic neutral lipid droplets and as multilamellar bodies. In addition to advanced atherosclerosis of the aorta, a 15-month-old WHHL rabbit also had focal coronary atherosclerosis and subcutaneous xanthomas. The New Zealand white rabbits fed a high cholesterol and fat diet for 2 weeks showed early intimal lipid accumulation in the aorta and prominent lipid accumulation in hepatocytes and macrophages of the liver and spleen. New Zealand white rabbits fed the standard rabbit chow had no abnormal lipid deposits. In contrast to the cholesterol-fed rabbits, WHHL rabbits had only mild lipid accumulation in other tissues. Thus, many similarities exist between atherosclerotic disease in the WHHL rabbit and in man. This study shows that the WHHL rabbit is a good model of familial hypercholesterolemia.
Lipoprotein profiles in rhesus monkeys with divergent responses to dietary cholesterol.
Authors: Baker H N , Eggen D A , Melchior G W , Roheim P S , Malcom G T , Strong J P
Published: June 2011

From a group of 53 rhesus monkeys, we selected 12 animals, the six with the highest and the six with the lowest response to a high cholesterol diet, and we made detailed analyses of their cholesterol and apolipoprotein profile. The high responders differed from the low responders in several ways. During the high cholesterol diet period, the high responders had much higher plasma apolipoprotein B and E concentrations and much lower plasma apolipoprotein A-I concentrations than did the low responders. Nearly all the increase in plasma cholesterol and apolipoproteins B and E concentrations in the high responders occurred in the lower density fractions (d = 1.006-1.030 g/ml), while the decrease in plasma apolipoprotein A-I concentrations in the high responders was confined to the lower density fraction of the high density lipoproteins (HDL), i.e., HDL2 (d = 1.063-1.125 g/ml). In the low responders, on the other hand, the slight increase in cholesterol concentrations was evenly distributed between the lower density fractions and HDL, and the increase in apolipoprotein A-I concentration of the lower density fractions is related to the decrease in the concentration in the HDL2 in the high responders.
Factors associated with lipoprotein cholesterol levels. The Framingham study.
Authors: Wilson P W , Garrison R J , Abbott R D , Castelli W P
Published: June 2011

Lipoprotein cholesterol determinations in 6328 individuals aged 20 to 79 years in the Framingham Heart Study Cohort and Offspring were related by multiple regression analysis to reported cigarette smoking, alcohol intake, and Quetelet Index (wt/ht2). Cigarette smoking was found to be strongly associated with "atherogenic" lipoprotein cholesterol profiles in young adults, and particularly in women. The associations for alcohol intake were mostly uniform across age groups and lipoprotein cholesterol fractions, while coefficients for Quetelet Index varied considerably. Blood chemistry associations were studied in participants under 50 years, after controlling for smoking, Quetelet Index, and alcohol intake. Significant associations for high density lipoprotein cholesterol (HDL-C) were seen for both sexes with alkaline phosphatase, serum calcium, serum u ric acid, and leucocyte count. Low density lipoprotein cholesterol (LDL-C) associations were observed with calcium, hematocrit, lactate dehydrogenase, and leucocyte count in men and women, while very low density lipoprotein cholesterol (VLDL-C) associations occurred with alkaline phosphatase, glucose, uric acid, and leucocyte count in both sexes. A 1 mg/dl higher calcium corresponded to an HDL-C approximately 4 mg/dl greater and a LDL-C typically 6 mg/dl greater after controlling for 12 other variables. A 1000/ml increase in leucocyte count was typically associated with a decrease in HDL-C by 1 mg/dl and an increase in LDL-C and VLDL-C of 1 mg/dl each.
Insulin-mediated increases in the HDL cholesterol/cholesterol ratio in humans.
Authors: Sadur C N , Eckel R H
Published: June 2011

The role of insulin in the regulation of lipoprotein cholesterol distribution was studied in 18 human volunteers who were of normal weight, normolipemic, and glucose-tolerant. We used the euglycemic clamp technique and made comparisons with six saline-infused controls. While total cholesterol and low density lipoprotein cholesterol levels fell similarly by 6 hours in both groups (p = NS), there was a greater decrease in triglyceride levels (p less than 0.02) but less decrease in high density lipoprotein cholesterol levels (p less than 0.02) in the insulin-infused group. These changes resulted in both a higher high density lipoprotein cholesterol/total cholesterol ratio (p = 0.01) and a higher high density lipoprotein cholesterol/low density cholesterol ratio (p = 0.02) in the insulin-infused group. The timing of this effect of insulin implies a mechanism unrelated to high density lipoprotein turnover. Thus, insulin infusion during euglycemia appears to alter cholesterol distribution favorably in normal subjects.
Monoclonal antibodies to E92, an endothelial cell surface antigen.
Authors: Kaplan K L , Weber D , Cook P , Dalecki M , Rogozinski L , Sepe O , Knowles D , Butler V P
Published: June 2011

Two hybridoma-derived monoclonal antibodies have been developed that react with an antigen of molecular weight 92,000 daltons on the surface of human endothelial cells. Cultured human umbilical vein endothelial cells were used for immunization, but the antigen is present on arterial, venous and capillary endothelium, as determined by biotin-avidin immunoperoxidase staining of tissue sections. With this technique, other cell types in the tissues which were examined were not reactive, except for scattered fibroblasts and histiomonocytic cells, trophoblastic cells of the placenta, and benign immature mesenchymal cells in a renal cystadenocarcinoma. By cytofluorography, the antibodies were found to be unreactive with granulocytes, T lymphocytes, B lymphocytes, and the majority of monocytes. Fibroblasts were reactive with the antibodies, but the fluorescence tracings indicated a lower density of antigen on these cells than on endothelial cells. Immunoreactivity of fibroblasts could be decreased by treatment of the cells with thrombin, trypsin, or neuraminidase, whereas these enzymes did not affect the immunoreactivity of endothelial cells. The reactive antigen (E92) does not appear to be any of several previously described endothelial cell proteins, because of its molecular weight and its absence on other cell types. The presence of E92 on trophoblastic cells of the placenta and immature mesenchymal cells, as well as fibroblasts and endothelial cells, may indicate that it is a primitive antigen of mesodermal tissue that is lost by most cell types during differentiation.
Compartmentalization of water in human atherosclerotic lesions. Changes in distribution and exclusion volumes for plasma macromolecules.
Authors: Smith E B , Ashall C
Published: June 2011

High concentrations of LDL and other plasma macromolecules are present in normal aortic intima and early proliferative atherosclerotic lesions that have not yet accumulated lipid. Their compartmentalization within the tissue water was estimated from the concentrations of low density lipoprotein (LDL), alpha 2-macroglobulin (alpha 2-M), and albumin in interstitial fluid and adjacent intimal tissue. Interstitial fluid was collected on filter paper inserted into natural strip planes in the intima; macromolecules in interstitial fluid and tissue were assayed by rocket immunoelectrophoresis directly from the papers or tissue. From their concentrations in interstitial fluid and total tissue water, the fractional and absolute volumes in which they were distributed (distribution volume) or from which they were excluded (exclusion volume) were calculated for each macromolecule. Compared with normal intima, the distribution volume increased by 60% and the exclusion volume by 95% in early gelatinous lesions; exclusion volume fraction was linearly related to molecular mass. In more advanced lesions, there was disproportionate exclusion of LDL and alpha 2-M. Their distribution volumes decreased and their concentrations in interstitial fluid increased; the concentration showed a significant inverse relation with distribution volume (regression coefficient, b = -0.53, p less than 0.01). Redistribution of tissue water may concentrate LDL in interstitial fluid, and contribute to its extracellular deposition.
Properties of cultured endothelium from adult human vessels.
Authors: Fry G , Parsons T , Hoak J , Sage H , Gingrich R D , Ercolani L , Nghiem D , Czervionke R
Published: June 2011

Endothelium was isolated from samples of aorta and vena cava obtained from cadaver donors at the time kidneys were harvested for transplantation. Digestion with collagenase and gentle swabbing were used to free the cells from the intimal surface. Low density seeding permitted isolation of individual colonies with typical endothelial morphology. Modified Medium 199 supplemented with 10%-20% human plasma-derived serum and an extract from the bovine hypothalamus (500 micrograms/ml) enabled subcultured colonies to grow to confluency when culture surfaces were coated with fibronectin (1 micrograms/cm2). The presence of Factor VIII antigen was demonstrated using an indirect immunofluorescence technique. A monoclonal antibody to cultured umbilical vein endothelium, specific for endothelium, reacted with the subcultured cells from the aorta and vena cava. Type IV procollagen, fibronectin, and thrombospondin were identified as labeled proteins secreted by cultures of adult endothelium that had been incubated with 3H-proline and 3H-glycine. When the cultured endothelium was used in a sodium-m-periodate stimulated T lymphocyte mitogenic culture system, the endothelium exhibited accessory cell function. Prostacyclin production stimulated by incubation with arachidonic acid and PGH2 was variable from vessel to vessel. However, average values were lower than normally seen with cultured primary umbilical vein endothelium.
Expression of apolipoprotein B epitopes in very low density lipoprotein subfractions. Studies with monoclonal antibodies.
Authors: Tikkanen M J , Cole T G , Hahm K S , Krul E S , Schonfeld G
Published: June 2011

Epitope expression was studied in both denatured apolipoprotein B (apo B) on Western blots and in intact low density lipoprotein (LDL) and very low density lipoprotein subfractions VLDL1 (Sf120-400), VLDL2 (Sf60-120), and VLDL3 (Sf20-60) in competitive binding immunoassays with the aid of six monoclonal anti-LDL antibodies. The apo B in all lipoprotein fractions was shown to bind to all antibodies, but significant differences in apo B epitope expression were observed. On the average, the immunoreactivity of VLDL subfractions (expressed as binding affinity and as relative 125I-LDL displacing potency) decreased with increasing flotation rate. Similarly, VLDL1 was less immunoreactive than lipolyzed "remnants" of VLDL1 after treatment with bovine milk lipoprotein lipase. The results indicate that, even when lipoprotein fractions obtained from the same individual and having the same kind of apo B subspecies were compared, significant differences in immunoreactivity occurred due to the modulating effect of other lipoprotein components on apo B epitope expression.
Aortic intimal response to endothelial removal in cebus and squirrel monkeys.
Authors: Hoover G A , Hayes K C
Published: June 2011

Whereas squirrel monkeys have an inherent susceptibility to atherosclerosis, cebus monkeys are relatively resistant. To assess whether this difference might lie in their response to endothelial injury, the acute morphologic changes in the aortic intima after endothelial removal were examined in the two species. The endothelium of the lower thoracic aorta was removed with an embolectomy catheter, and the intimal response was compared with the uninjured upper thoracic aorta in each monkey. By 21 days after aortic denudation, regrowth of the endothelium (assessed by in vivo Evans' blue dye staining) was significantly greater in squirrel monkeys (90% +/- 5% of aortic surface) than in cebus (56% +/- 11%). Squirrel monkeys had comparable sudanophilic surface in the nonballooned, control aorta (25% +/- 7%) and the ballooned, lower thoracic segment (17% +/- 6%). Cebus monkey aortas had no sudanophilia in either segment. The intima/media ratios (IMR) in all regions of the aorta were significantly greater in squirrel monkeys than in cebus, but in both species the IMR of the ventral ballooned segment was two to three times the IMR of the nonballooned control segment. In the dorsal aorta, where endothelial regrowth was more rapid, the IMR was similar to the control aortic segment. By electron microscopy the thickened aortic intima in both species contained a marked increase in modified smooth muscle cells, but lipid accumulation did not result from endothelial removal or regrowth in either species. Thus, although the squirrel monkey aorta had atherosclerotic lesions before endothelial removal, the acute intimal response to endothelial injury was similar in degree and kind in both cebus and squirrel monkeys. This suggests that factors other than those controlling the initial intimal thickening following endothelial injury are responsible for the observed difference in arterial lipid accumulation between cebus and squirrel monkeys.
In vitro reendothelialization. Microfilament bundle reorganization in migrating porcine endothelial cells.
Authors: Gotlieb A I , Spector W , Wong M K , Lacey C
Published: June 2011

An experimentally induced wound made in a confluent monolayer culture of porcine thoracic aortic endothelial cells (ECs) was studied 22 hours after wounding using 7-nitrobenz-2-oxa-1,3-diazole (NBD) phallacidin and immunofluorescence microscopy to localize actin and myosin containing microfilament (MF) bundles. ECs extending from the wound edge back toward the confluent monolayer showed a specific change in cell shape and in MF bundle distribution and orientation, which correlated with the cell migration behavior observed using time-lapse cinemicrophotography. The migrating ECs in the first zone, the leading zone, were polygonal to partially elongated in shape, and contained distinct central MF bundles oriented both parallel and perpendicular to the wound edge. The second zone, the elongated zone, was characterized by elongated cells with central MF bundles oriented parallel to the direction of migration. A third zone, the transitional zone, showed nonmigrating polygonal ECs containing prominent central and dense peripheral bands (DPB) of MF bundles. The central MF bundles were oriented randomly with respect to the wound edge. The MF bundles of the confluent resting monolayer were both centrally and peripherally located with the latter being more prominent. The results indicate that the reorientation of central MF bundles and reduction in the peripheral MF bundles are probably important in the reorganization of the cytoskeletal system during the conversion of stationary cells to migrating cells.
Intimal injury in vivo activates vascular smooth muscle cell migration and explant outgrowth in vitro.
Authors: Grünwald J , Haudenschild C C
Published: June 2011

In vivo induced endothelial injury results in an intimal thickening, mainly caused by stimulation of smooth muscle cells (SMC), which migrate into, and proliferate in, the denuded intima. We evaluated the manifestation and persistence of this stimulus in several in vitro explant and growth assays using aortic explants from either balloon catheter-injured rats or from sham-operated controls. SMC outgrowth from control explants started 48 hours after explantation, became half-maximal after 96 hours, with 74% of the explants showing outgrowth. Explants from balloon-injured aortas showed SMC outgrowth within 24 hours, became half-maximal after 48 hours, with 92% of the explants showing outgrowth. An interval of 4 to 7 days between balloon injury and sacrifice was optimal for obtaining accelerated outgrowth, while a 1-day interval showed a weak stimulation and a 21-day interval had no such effect. The in vivo stimulated cells grew more rapidly, resulting in a greater colony size per explant, and they became temporarily serum-independent. Increased growth rate persisted up to the second subculture but returned to normal in subsequent passages. Mechanisms of SMC migration can now be quantitatively assessed by replacing the intima with the tissue culture dish, allowing studies on the extent and persistence of atherogenic stimuli.
Plasma triglyceride determines structure-composition in low and high density lipoproteins.
Authors: Deckelbaum R J , Granot E , Oschry Y , Rose L , Eisenberg S
Published: June 2011

Because the association of hypertriglyceridemia and premature atherosclerosis is not due to the direct effects of the triglyceride molecule itself, we studied the effects of increased plasma triglyceride-rich lipoproteins on the composition and structure of low density lipoprotein (LDL) and high density lipoprotein (HDL). We found profound changes in the core and surface domains of both lipoproteins with increasing triglyceridemia. Core cholesterol esters were progressively depleted and replaced by triglyceride molecules. Highly significant negative correlations were found between cholesterol ester/protein ratios (r = -0.64 for LDL and -0.58 for HDL (p less than 0.001); positive correlations were found for triglyceride/protein ratios (r = 0.62 for LDL and 0.58 for HDL) and for triglyceride/cholesterol ester ratios (r = 0.70 for LDL and 0.83 for HDL) when these variables were assayed as a function of plasma triglyceride concentrations. With severe hypertriglyceridemia, triglyceride/cholesterol ester ratios of more than 1.0 were consistently observed (normal, less than 0.02). This leads to an underestimation of LDL and HDL levels when cholesterol alone is measured. At the surface, LDL and HDL were depleted of phospholipid and free cholesterol, with a relative enrichment of protein. These changes can be explained on the basis of high levels of plasma triglyceride-rich lipoproteins serving as acceptors for cholesterol esters and other constituents from LDL and HDL. Concomitantly, triglycerides are transferred to LDL and HDL. These transfer processes are likely to be mediated by the activity of lipid transfer proteins present in human plasma.
Transport of 125I-albumin across normal and deendothelialized rabbit thoracic aorta in vivo.
Authors: Ramirez C A , Colton C K , Smith K A , Stemerman M B , Lees R S
Published: June 2011

Transmural concentration profiles of 125I-albumin in vivo were measured across the normal and balloon catheter-deendothelialized rabbit descending thoracic aorta as a function of time following intravenous injection. A tracer was injected 5 or 60 minutes after deendothelialization, and the animals were sacrificed after circulation times of 10, 30 or 60 minutes. The aorta was immediately excised and frozen flat between glass slides. Samples were serially sectioned parallel to the intimal surface in a refrigerated microtome, washed with trichloroacetic acid (TCA), and counted. Relative tissue concentration profiles of TCA-precipitable radioactivity from the media of control animals showed entry from both luminal and adventitial sides, as previously found with conscious normal rabbits, but spatial gradients at both luminal and medial-adventitial borders were less steep. Relative concentration levels in ballooned animals were 10- to 40-fold higher than in controls, and the profiles were flatter. Uptake rates at equivalent circulation times were greater in experiments initiated 60 minutes, as compared with 5 minutes, after deendothelialization, suggesting that progressive medial edema may have occurred following balloon injury. These results show that the intact endothelium is the dominant mass transfer resistance for 125I-albumin transport across the aortic wall. The data also suggest that the incomplete monolayer of platelets adherent to the subendothelium after balloon deendothelialization is not a substantial resistance to transport, as compared to that of the media, and that convection plays a more important role than diffusion for 125I-albumin transport across the deendothelialized aortic wall.