Arteriosclerosis and thrombosis : a journal of vascular biology最新文献

An enzyme with acyl coenzyme A:cholesterol acyltransferase (ACAT) activity was isolated from porcine liver, and sequences derived from trypsinized peptides indicated homology to liver carboxylesterase. By use of degenerate primers, human cDNA clones were identified, which were identical to human liver carboxylesterase. Expression of the full-length cDNA in Chinese hamster ovary (CHO) cells led to an approximately threefold increase in cellular ACAT activity. This was accompanied by an approximately 20-fold increase of cellular cholesteryl ester content. By light and electron microscopy, recombinant CHO cells contained numerous lipid droplets that were not present in control CHO cells. Expression of an antisense cDNA in HepG2 cells reduced cellular ACAT activity by 35% compared with control. To further investigate the role of the enzyme in cellular cholesterol homeostasis, regulation of the mRNA was investigated in 7-day cultured human mononuclear phagocytes (MNPs). When these cells were incubated in lipoprotein-deficient serum for 18 hours, the mRNA for ACAT/carboxylesterase was almost not detectable on Northern blots, whereas after incubation with acetylated low-density lipoproteins, a strong hybridization signal was obtained. This is evidence that the mRNA of ACAT/carboxylesterase is induced by cholesterol loading. It is concluded from the data presented that ACAT/carboxylesterase is relevant for cellular cholesterol esterification in vivo. The regulation in MNPs indicates that the enzyme is also involved in foam cell formation during early atherogenesis.

The effect of human leukemia inhibitory factor (hLIF) on the development of atherosclerosis was investigated in an experimental animal model. Two conditions were examined: one in which lesions could arise because of the influence of both "injury" (cuffed vessel) and diet and one in which only the effect of diet could be significant in other areas of the vasculature (aorta). At time zero, the right carotid artery of rabbits (n = 32) was ensheathed in a soft Silastic cuff, and an osmotic minipump (2-mL capacity; 2.5 microL/h; 28 days) containing either hLIF or saline was inserted into the peritoneal cavity. Rabbits were divided into four groups (n = 8): group 1 received normal diet/saline; group 2, normal diet/LIF (30 micrograms.kg-1.d-1); group 3, 1% cholesterol diet/saline; and group 4, 1% cholesterol diet/LIF (30 micrograms.kg-1.d-1). After 28 days, the cholesterol diet (group 3) resulted in a sixfold increase in plasma cholesterol level compared with group 1 rabbits on a normal diet (3.80 +/- 0.50 versus 0.55 +/- 0.01 mmol/L). This was significantly lower (P = .01) with hLIF treatment in group 4 rabbits (2.80 +/- 0.44 mmol/L). Group 2 rabbits had higher aortic tissue cholesterol levels (1.40 +/- 0.35 mg/g) compared with group 1 rabbits on a normal diet (0.10 +/- 0.06 mg/g) (P = .01), whereas hLIF treatment decreased tissue cholesterol levels by 60% in group 4 rabbits (0.60 +/- 0.05 mg/g) (P = .01). Group 3 rabbits developed lipid-filled lesions covering 63.25 +/- 17.66% of the thoracic aorta surface, whereas lesions were significantly reduced (9.88 +/- 8.79%) (P = .01) with LIF treatment (group 4).(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of dietary cholesterol and chronic administration of moderate amounts of ethanol on collagen-induced platelet responses were investigated. Three groups of rabbits were fed the following diets for 8 weeks: a normal chow diet, a cholesterol-enriched (0.25% wt/wt) chow diet, and a cholesterol-enriched chow diet plus 6% ethanol in the drinking water for the final week of the dietary period. Cholesterol feeding enhanced collagen-induced responses-aggregation, secretion of [14C]serotonin from prelabeled platelets, and thromboxane formation--of suspensions of washed platelets, and chronic ethanol treatment significantly reduced these enhanced responses. These effects are mediated by thromboxane A2 (TxA2) rather than ADP. Experiments with collagen-stimulated platelets in which feedback amplification of TxA2 was blocked with the prostaglandin H2/TxA2 receptor blocker BM 13.177 and experiments with aspirin-treated platelets stimulated with the stable TxA2 mimetic U46619 showed that cholesterol feeding enhanced platelet sensitivity to TxA2 rather than formation of TxA2 by platelets that had interacted with collagen. Without BM 13.177 or aspirin, TxA2 increased the amount of TxA2 formed by feedback amplification. In contrast, decreased responsiveness to collagen by platelets from cholesterol-fed rabbits given ethanol was due to inhibition of TxA2 formation rather than reduced sensitivity to TxA2. Platelets from cholesterol-fed rabbits given ethanol did not develop tolerance to the acute inhibitory effects of ethanol. Our results indicate that administration of moderate amounts of ethanol to cholesterol-fed rabbits inhibits enhanced collagen-induced responses of platelets by a TxA2-dependent pathway that involves reduction of TxA2 formation rather than reduction of platelet responses to TxA2.

The triglyceride content of the plasma very-low-density lipoprotein (VLDL) fraction is the most important factor affecting the size of low-density lipoprotein (LDL) in humans. Because cholesteryl ester transfer protein (CETP) can influence the size distribution of LDL particles in human plasma, the implication of lipid transfers in the formation of small-sized LDL patterns, which have been associated with elevated plasma triglyceride levels, was investigated. The size distribution of LDL particles in 15 plasma samples was determined by electrophoresis of the plasma LDL fraction on 20 to 160 g/L polyacrylamide gradient gels. The apparent diameter of the major LDL subfraction was shown to correlate negatively with triglyceride concentrations (r = -.706, P < .005) and positively with both high-density lipoprotein cholesterol levels (r = .637, P < .02) and the high-density lipoprotein/VLDL + LDL cholesterol ratio (r = .768, P < .001). In addition, LDL size correlated negatively with both the ability of plasma LDL to donate cholesteryl esters (r = -.79, P < .001) and its ability to acquire triglycerides (r = -.72, P < .005). Whereas these observations indicated that CETP-mediated alterations of the triglyceride/cholesteryl ester ratio of the LDL core would contribute to changes in LDL diameter, they suggested that the formation of small-sized gradient gel LDL patterns would require another biochemical event, such as lipolysis, in addition to neutral lipid transfers. To test this hypothesis, total plasma samples with or without added VLDL (added triglyceride concentration, 2.0 g/L) were preincubated for 24 hours at 37 degrees C. Preincubation mainly induced the replacement of cholesteryl esters by triglycerides in the LDL core, and changes in LDL composition were greater when total plasma was supplemented with VLDL. Subsequently, isolated LDL was incubated in the presence of bovine milk lipoprotein lipase as a source of triglyceride hydrolysis activity. Lipolysis tended to reduce the size of the major LDL subpopulation, and the mean change in LDL diameter was significantly greater when plasma was preincubated with VLDL supplementation than when it was not (-0.6 +/- 0.3 versus -0.2 +/- 0.2 nm, respectively; (P < .01). Moreover, sequential effects of lipid transfer and lipolysis activities induced dramatic changes in the general shape of gradient gel LDL patterns. The largest plasma LDL subpopulations tended to disappear, and the formation of new, small LDL particles could be observed. The combined effects of neutral lipid transfers and triglyceride hydrolysis could account for variations of gradient gel LDL profiles in human plasma.(ABSTRACT TRUNCATED AT 400 WORDS)

The aim of this study was to evaluate whether high-risk hypertensive patients (n = 137) had larger far-wall common carotid artery intima-media thickness than a control group (n = 37) and to study whether intima-media thickness was related to other signs of atherosclerotic disease. The results showed that intima-media thickness was significantly larger in the hypertension group than in the control group. Lumen diameter and mean cross-sectional area of the intima-media complex were larger both for hypertensive patients with a positive history of manifest clinical cardiovascular disease and for hypertensive patients with no such history than in the control group. There was a significant relationship between far-wall common carotid artery intima-media thickness and plaque status (visual scoring, no, small, moderate/large) in the carotid artery region. In univariate analyses, low diastolic blood pressure and high pulse pressure were both significantly related to plaque status. In multivariate analyses, pulse pressure was significantly and independently related both to common carotid artery intima-media thickness and to plaque status in the carotid artery region. In multivariate analyses, there was also an independent relationship between age and common carotid artery intima-media thickness, between smoking status and plaque status, and between a positive history of manifest clinical cardiovascular disease and plaque status. In conclusion, common carotid artery intima-media thickness and lumen diameter were increased in elderly high-risk hypertensive patients, in whom more than one third of the patients also had a moderate to large plaque in the carotid artery region.(ABSTRACT TRUNCATED AT 250 WORDS)

It recently has been hypothesized that increased plasma plasminogen activator inhibitor-1 (PAI-1) activity could be a possible link between insulin resistance and coronary heart disease. However, it is not known whether insulin sensitivity per se is a determinant of plasma PAI-1 activity or whether other intermediates could explain this association. We investigated the relationship between plasma PAI-1 activity and insulin sensitivity, obesity, distribution of body fat, blood pressure, plasma insulin concentration, and serum lipid levels in normoglycemic men (n = 61) and women (n = 77) 53 to 61 years old who participated in a previous population-based study. Insulin sensitivity was estimated by the minimal model from a frequently sampled intravenous glucose tolerance test. In univariate analyses, PAI-1 correlated positively with body mass index, waist-to-hip ratio (WHR), fasting and 2-hour insulin levels, and triglyceride level in both men and women. Furthermore, in women PAI-1 correlated inversely with high-density lipoprotein (HDL) cholesterol level. There was an inverse relationship between PAI-1 and insulin sensitivity (r = -.39, P < .01 in men; r = -.38, P < .001 in women). In multivariate analyses in men, insulin sensitivity failed to show any significant association with PAI-1. In contrast, triglyceride level and body mass index were independently associated with PAI-1. Also in women, insulin sensitivity was not independently associated with PAI-1. In women, WHR and HDL cholesterol concentration or WHR and 2-hour insulin concentration were independently related to PAI-1.(ABSTRACT TRUNCATED AT 250 WORDS)

The 10-kb deletion ("French Canadian mutation") of the low-density lipoprotein (LDL) receptor gene is the most common mutation causing familial hypercholesterolemia among subjects of French Canadian descent. In affected subjects, it results in a null allele of the LDL receptor gene and provides a unique opportunity to examine single-allele regulation of this gene in humans. We sought to ascertain the response of inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A (HMG CoA) reductase in subjects with the French Canadian mutation of the LDL receptor gene and to correlate this response with biochemical variables and the haplotype of the nondeletion LDL receptor allele. The prevalence of non-responders to high doses of HMG CoA reductase inhibitors (defined as < 15% decrease in LDL cholesterol [LDL-C] from baseline values after dietary intervention) was ascertained in 105 patients heterozygous for the 10-kb deletion after excluding first-degree relatives and those on combined lipid-lowering therapy or other lipid-lowering agents. Lipoprotein cholesterol levels were examined after a diet period (30% calories as fat) and after receiving HMG CoA reductase inhibitors as mono-therapy for a minimum of 3 months. The mean reduction in total cholesterol was 45 +/- 23%, in LDL-C 33 +/- 15%, and in triglycerides 32 +/- 49% (all P < .005). There was a slight increase in high-density lipoprotein cholesterol of 8.5 +/- 18% (P > .05).(ABSTRACT TRUNCATED AT 250 WORDS)

We periodically obtained blood samples from mildly hypercholesterolemic, but otherwise healthy, premenopausal women who were recruited to participate in a study of a long-term, cholesterol-lowering diet. All meals were prepared and most meals were consumed in the study center dining facility. Tests performed on blood samples included fibrinogen, cholesterol, factor VII coagulant activity (VIIc), and other measures of factor VII. We found that when women switched from a typical American diet (37% fat, polyunsaturated fatty acid to saturated fatty acid [P/S] ratio 0.5, 300 mg cholesterol/d) to a diet lower in fat and cholesterol (American Heart Association phase 2 diet: 30% fat, P/S ratio of 1, 150 to 200 mg cholesterol/d) and maintained that diet for 20 weeks, their plasma cholesterol levels decreased by approximately 6% after 4 weeks and remained at that level until study termination. Likewise, VIIc decreased by approximately 11% while factor VII antigen, total factor VII activity, and fibrinogen concentration did not change appreciably from baseline values. Our results show that premenopausal women benefit from a diet lower in total and saturated fat by a reduction in blood cholesterol and VIIc. Extrapolation from data on men in the Northwick Park Heart Study indicates that the 11% decrease in VIIc activity would correspond to an approximately 30% decrease in risk of mortality from coronary heart disease.

Lipoprotein(a) [Lp(a)] concentration and apolipoprotein(a) [apo(a)] isoforms (identified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis [SDS-PAGE] and Western blotting) were determined in a group of 508 asymptomatic Caucasian members of the community and in 318 Caucasian patients with angiographically defined coronary artery disease (CAD). Conventional risk factors for CAD were also measured. Lp(a) concentration was almost twice as high in subjects with CAD (geometric mean, 152 mg/L [geometric SD, 10 to 1398 mg/L]) as in asymptomatic control subjects (geometric mean, 84 mg/L [geometric SD, 21 to 334 mg/L]). Asymptomatic women had higher concentrations of Lp(a) than asymptomatic men. Patients with CAD were older and were more likely to have smoked and to have a first-degree relative with premature CAD (< 55 years of age), and a higher proportion were male. Patients with CAD had higher concentrations of Lp(a) independently of the number of isoform bands expressed. When apo(a) isoforms were allocated to 1 of 10 classes on the basis of their molecular size (Rf versus apoB in SDS-PAGE), patients with CAD did not express an excess of low-molecular-mass (higher concentration) isoforms but did express a higher proportion of double-band phenotypes with fewer "null" phenotypes. The relationship between the two isoform bands in a double-band phenotype was the same in both populations. Isoform mobility was defined as a continuous variable equal to the mobility of a single isoform band (single-band phenotypes) or the mean of the two isoforms in a double-band phenotype. Two variables, isoform mobility and the number of isoform bands expressed, were used to summarize the large range of isoform patterns (at least 45) that could be identified. Isoform mobility, the number of isoform bands expressed, and the presence of CAD were the three most important independent predictors of Lp(a) concentration (descending order). Only sex and LDL cholesterol were additional independent predictors of Lp(a) concentration in step-wise regression models including a wide range of demographic factors and lipid and glycemic risk factors. We conclude that Lp(a) concentration is associated with CAD independently of the isoform pattern expressed. The apo(a) gene locus exerts a strong control over circulating Lp(a) concentration, and a better understanding of the control of expression of the apo(a) gene will be essential to understand the relationship between Lp(a) and CAD.

Lipid deposits in human atherosclerotic fibrous plaques exhibit marked differences in chemistry and ultrastructure from lipid deposits in fatty streaks, leading some investigators to question whether fibrous plaques originate from fatty streaks. To examine lesion transition, we employed lipid microanalysis, electron microscopy, and immunohistochemistry on fatty streaks, fibrolipid lesions (small raised lesions), and fibrous plaques from human aorta. Both fatty streaks and caps of fibrolipid lesions were high in esterified cholesterol content (mean, 62% of total cholesterol) and high in cholesteryl oleate content compared with cholesteryl linoleate content. Fatty streaks and fibrolipid lesion caps also showed similar morphology, characterized mostly by macrophage-derived foam cells in the superficial intima. Core lipids in both small and large raised lesions differed markedly from this pattern. Fibrolipid lesion cores showed mostly vesicular extracellular deposits, sometimes accompanied by cholesterol clefts, while fibrous plaque core deposits were also extracellular but had a variable appearance. Compared with fatty streaks, fibrolipid lesion cores showed significantly increased free/total cholesterol fractions (63%) and decreased fractional contents of cholesteryl oleate. Fibrous plaque cores had variable distributions of free and esterified cholesterol but significantly decreased cholesteryl oleate fractions compared with fatty streaks. The results support the concept of lesion transition, which is marked by deep intimal, extracellular deposition of cholesterol-rich, vesicular lipid deposits in small raised lesions. In the core region of larger raised lesions, both cholesterol-rich and cholesteryl ester-rich lipid deposits appear to form in the extracellular space.