Oxidation affects the flow-induced aggregation of low density lipoprotein and its inhibition by albumin

We investigated whether oxidation alters the self-aggregation of low density lipoprotein (LDL) and the inhibition of such aggregation by albumin. Incubation with copper for different durations produced mildly, moderately, and highly oxidised LDL (having, respectively, ca. 60, 300 and 160 nmol lipid hydroperoxides/mg protein, and electrophoretic mobilities 1.2, 2.6 and 4.4 times that of native LDL). The rate of flow-induced aggregation was the same for native, mildly oxidised and moderately oxidised LDL, but decreased for highly oxidised LDL. The inhibitory effect of albumin (40 mg/ml) on aggregation was reduced by mild oxidation and further reduced by moderate or severe oxidation. The net result of the two effects was that in the presence of albumin, moderately oxidised LDL had the highest rate of aggregation and native the lowest. The reduction in the anti-aggregatory effect of albumin provides a new mechanism by which LDL oxidation might enhance net aggregation in vivo. (C) 2003 Elsevier B.V. All rights reserved.

Top-down systems biology modeling of host metabotype-microbiome associations in obese rodents

Covariation in the structural composition of the gut microbiome and the spectroscopically derived metabolic phenotype (metabotype) of a rodent model for obesity were investigated using a range of multivariate statistical tools. Urine and plasma samples from three strains of 10-week-old male Zucker rats (obese (fa/fa, n = 8), lean (fal-, n = 8) and lean (-/-, n = 8)) were characterized via high-resolution H-1 NMR spectroscopy, and in parallel, the fecal microbial composition was investigated using fluorescence in situ hydridization (FISH) and denaturing gradient gel electrophoresis (DGGE) methods. All three Zucker strains had different relative abundances of the dominant members of their intestinal microbiota (FISH), with the novel observation of a Halomonas and a Sphingomonas species being present in the (fa/fa) obese strain on the basis of DGGE data. The two functionally and phenotypically normal Zucker strains (fal- and -/-) were readily distinguished from the (fa/fa) obese rats on the basis of their metabotypes with relatively lower urinary hippurate and creatinine, relatively higher levels of urinary isoleucine, leucine and acetate and higher plasma LDL and VLDL levels typifying the (fa/fa) obese strain. Collectively, these data suggest a conditional host genetic involvement in selection of the microbial species in each host strain, and that both lean and obese animals could have specific metabolic phenotypes that are linked to their individual microbiomes.

Effect of oxidized low-density lipoprotein on differential gene expression in primary human endothelial cells

Oxidative modification of low-density lipoprotein (LDL) plays an important role in the initiation and progression of atherosclerosis. It has been proposed that the biological action of oxidized LDL (ox-LDL) may be partially attributed to its effect on a shift of the pattern of gene expression in endothelial cells. To examine the transcriptional response to ox-LDL, we applied cDNA array technology to cultured primary human endothelial cells challenged with oxidized human LDL. A twofold or greater difference in the expression of a particular gene was considered a significant difference in transcript abundance. Seventy-eight of the 588 genes analyzed were differentially expressed in response to the treatment. Ox-LDL significantly affected the expression of genes encoding for transcription factors, cell receptors, growth factors, adhesion molecules, extracellular matrix proteins, and enzymes involved in cholesterol metabolism. The alteration of the expression pattern of several genes was substantiated post hoc using RT-PCR. The experimental strategy identified several novel ox-LDL-sensitive genes associated with a "response to injury" providing a conceptual background to be utilized for future studies addressing the molecular basis of the early stages of atherogenesis.

Antioxidant and anti-atherogenic activities of olive oil phenolics

The aim of the current study was to investigate the antioxidant and cellular activity of the olive oil phenolics oleuropein, tyrosol, hydroxytyrosol, and homovanillic alcohol (which is also a major metabolite of hydroxytyrosol). Well-characterized chemical and biochemical assays were used to assess the antioxidant potential of the compounds. Further experiments investigated their influence in cell culture on cytotoxic effects of hydrogen peroxide and oxidized low-density lipoprotein (LDL), nitric oxide production by activated macrophages, and secretion of chemoattractant and cell adhesion molecules by the endothelium. Inhibitory influences on in vitro platelet aggregation were also measured. The antioxidant assays indicated that homovanillic alcohol was a significantly more potent antioxidant than the other phenolics, both in chemical assays and in prolonging the lag phase of LDL oxidation. Cell culture experiments suggested that the olive oil phenolics induce a significant reduction in the secretion of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 (and a trend towards a reduced secretion of monocyte chemoattractant protein-1), and protect against cytotoxic effects of hydrogen peroxide and oxidized LDL. However, no influence on nitric oxide production or platelet aggregation was evident. The data show that olive oil phenolics have biochemical and cellular actions, which, if also apparent in vivo, could exert cardioprotective effects.

Effect of circulating forms of soy isoflavones on the oxidation of low density lipoprotein

Soy isoflavones are thought to have a cardioprotective effect that is partly mediated by an inhibitory influence on the oxidation of low density lipoprotein (LDL). However, the aglycone forms investigated in many previous studies do not circulate in appreciable quantities because they are metabolised in the gut and liver. We investigated effects of various isoflavone metabolites, including for the first time the sulphated conjugates formed in the liver and the mucosa of the small intestine, on copper-induced LDL oxidation. The parent aglycones inhibited oxidation, although only 5% as well as quercetin. Metabolism increased or decreased their effectiveness. Equol inhibited 2.65-fold better than its parent compound daidzein and 8-hydroxydaidzein, not previously assessed, was 12.5-fold better than daidzein. However, monosulphated conjugates of genistein, daidzein and equol were much less effective and disulphates completely ineffective. Since almost all isoflavones circulate as conjugates, these data suggest that despite the increased potency produced by some metabolic changes, isoflavones may not be effective antioxidants in vivo unless they are deconjugated again.

Opposing effects of cis-9,trans-11 and trans-10,cis-12 conjugated linoleic acid on blood lipids in healthy humans

Background: Conjugated linoleic acid (CLA) is reported to have weight-reducing and antiatherogenic properties when fed to laboratory animals. However, the effects of CLA on human health and, in particular, the effects of individual CLA isomers are unclear. Objective: This study investigated the effects of 3 doses of highly enriched cis-9,trans-11 (0.59, 1.19, and 2.38 g/d) or trans-10,cis-12 (0.63, 1.26, and 2.52 g/d) CLA preparations on body composition, blood lipid profile, and markers of insulin resistance in healthy men. Design: Healthy men consumed 1, 2, and 4 capsules sequentially, containing either 80% cis-9,trans-11 CLA or 80% trans-10,cis-12 CLA for consecutive 8-wk periods. This phase was followed by a 6-wk washout and a crossover to the other isomer. Results: Body composition was not significantly affected by either isomer of CLA. Mean plasma triacylglycerol concentration was higher during supplementation with trans-10,cis-12 CLA than during that with cis-9,trans-11 CLA, although there was no influence of dose. There were significant effects of both isomer and dose on plasma total cholesterol and LDL-cholesterol concentrations but not on HDL-cholesterol concentration. The ratios of LDL to HDL cholesterol and of total to HDL cholesterol were higher during supplementation with trans-10,cis-12 CLA than during that with cis-9,trans-11 CLA. CLA supplementation had no significant effect on plasma insulin concentration, homeostasis model for insulin resistance, or revised quantitative insulin sensitivity check index. Conclusion: Divergent effects of cis-9,trans-11 CLA and trans10,cis-12 CLA appear on the blood lipid profile in healthy humans: trans-10,cis-12 CLA increases LDL:HDL cholesterol and total:HDL cholesterol, whereas cis-9,trans-11 CLA decreases them.

Effects of dairy products naturally enriched with cis-9,trans-11 conjugated linoleic acid on the blood lipid profile in healthy middle-aged men

Background: Interest in the development of dairy products naturally enriched in conjugated linoleic acid (CLA) exists. However, feeding regimens that enhance the CLA content of milk also increase concentrations of trans-18:1 fatty acids. The implications for human health are not yet known. Objective: This study investigated the effects of consuming dairy products naturally enriched in cis-9,trans-11 CLA (and trans-11 18:1) on the blood lipid profile, the atherogenicity of LDL, and markers of inflammation and insulin resistance in healthy middle-aged men. Design: Healthy middle-aged men (n = 32) consumed ultra-heat-treated milk, butter, and cheese that provided 0.151 g/d (control) or 1.421 g/d (modified) cis-9,trans-11 CLA for 6 wk. This was followed by a 7-wk washout and a crossover to the other treatment. Results: Consumption of dairy products enriched with cis-9,trans-11 CLA and trans-11 18:1 did not significantly affect body weight, inflammatory markers, insulin, glucose, triacylglycerols, or total, LDL, and HDL cholesterol but resulted in a small increase in the ratio of LDL to HDL cholesterol. The modified dairy products changed LDL fatty acid composition but had no significant effect on LDL particle size or the susceptibility of LDL to oxidation. Overall, increased consumption of full-fat dairy products and naturally derived trans fatty acids did not cause significant changes in cardiovascular disease risk variables, as may be expected on the basis of current health recommendations. Conclusion: Dairy products naturally enriched with cis-9,trans-11 CLA and trans-11 18: 1 do not appear to have a significant effect on the blood lipid profile.

Long-term monounsaturated fatty acid diets reduce platelet aggregation in healthy young subjects

The aim of the present study was to compare the response of a range of atherogenic and thrombogenic risk markers to two dietary levels of saturated fatty acid (SFA) substitution with monounsaturated fatty acids (MUFA) in students living in a university hall of residence. Although the benefits of such diets have been reported for plasma lipoproteins in high-risk groups, more needs to be known about effects of more modest SFA-MUFA substitutions over the long term and in young healthy adults. In a parallel design over 16 weeks, fifty-one healthy young subjects were randomised to one of two diets: (1) a moderate-MUFA diet in which 16 g dietary SFA/100 g total fatty acids were substituted with MUFA (n 25); (2) a high-MUFA diet in which 33 g dietary SFA/100 g total fatty acids were substituted with MUFA (n 26). All subjects followed an 8-week run-in diet (reference diet), with a fatty acid composition close to the UK average values. There were no differences in plasma lipid responses between the two diets over 16 weeks of the study with similar reductions in total cholesterol (P<0.001) and LDL-cholesterol (P<0.01) in both groups; a small but significant reduction in HDL-cholesterol was also observed in both groups (P<0.01). Platelet responses to ADP (P<0.01) and arachidonic acid (P<0.05) differed with time on the two diets; at 16 weeks, platelet aggregatory response to ADP was significantly lower on the high-MUFA than the moderate-MUFA (P<0.01) diet; ADP responses were also significantly lower within this group at 8 (P< 0.05) and 16 (P< 0.01) weeks compared with baseline. There were no differences in fasting factor VII activity (factors VIII and VIIag), fibrinogen concentration or tissue-type plasminogen activator activity between the diets. There were no differences in postprandial factor VIII responses to a standard meal (area under the curve) between the diets after 16 weeks, but postprandial factor VIII response was lower than on the high-MUFA diet compared with baseline (P<0.01). In conclusion, a high-MUFA diet sustains potentially beneficial effects on platelet aggregation and postprandial activation of factor VII. Moderate or high substitution of MUFA for SFA achieves similar reductions in fasting blood lipids in young healthy subjects.

ACTH reduces the rise in ApoB-48 levels after fat intake

It has been repeatedly demonstrated that ACTH administration lowers plasma lipid concentrations in man. The present study was designed to test the hypothesis, based on observations of decreased apolipoprotein B (ApoB) synthesis and secretion in vitro, that ACTH administration inhibits the postprandial output of ApoB in man. Therefore, we studied the response to a fat-rich meal supplemented with Vitamin A in eight healthy volunteers, who underwent this test without premedication, after 4 days administration of ACTH, and after 4 days administration of a glucocorticoid (betamethasone). As expected, fasting plasma levels of low-density lipoproteins (LDL)-cholesterot (-25%) and ApoB (-17%) decreased after ACTH, but not after betamethasone administration. Also, the elevation of plasma ApoB-48 in response to fat intake (to twice the basal levels) was markedly reduced after ACTH administration. However, the postprandial rise in plasma triglycerides and retinyl palmitate was unimpaired, suggesting that ACTH administration induced the secretion of fewer but larger chylomicrons. The effect of betamethasone on the postprandial response was similar but less pronounced. This study confirms earlier reports on the lipid-lowering effects of ACTH and supports our theory, based on in vitro studies, that the lipid-lowering effects of ACTH administration in man involves an inhibition of ApoB production. (c) 2006 Elsevier Ireland Ltd. All rights reserved.

Effects of consumption of probiotics and prebiotics on serum lipid levels in humans

The objective of this article is to review existing studies concerning the effects of probiotics and prebiotics on serum cholesterol concentrations, with particular attention on the possible mechanisms of their action. Although not without exception, results from animal and human studies suggest a moderate cholesterol-lowering action of dairy products fermented with appropriate strain(s) of lactic acid bacteria and bifidobacteria. Mechanistically, probiotic bacteria ferment food-derived indigestible carbohydrates to produce short-chain fatty acids in the gut, which can then cause a decrease in the systemic levels of blood lipids by inhibiting hepatic cholesterol synthesis and/or redistributing cholesterol from plasma to the liver. Furthermore, some bacteria may interfere with cholesterol absorption from the gut by deconjugating bile salts and therefore affecting the metabolism of cholesterol, or by directly assimilating cholesterol. For prebiotic substances, the majority of studies have been done with the fructooligosaccharides inulin and oligofructose, and although convincing lipid-lowering effects have been observed in animals, high dose levels had to be used. Reports in humans are few in number. In studies conducted in normal-lipidemic subjects, two reported no effect of inulin or oligofructose on serum lipids, whereas two others reported a significant reduction in serum triglycerides (19 and 27%, respectively) with more modest changes in serum total and LDL cholesterol. At present, data suggest that in hyperlipidemic subjects, any effects that do occur result primarily in reductions in cholesterol, whereas in normal lipidemic subjects, effects on serum triglycerides are the dominant feature.

Differential uptake of subfractions of triglyceride-rich lipoproteins by THP-1 macrophages

It is well known that raised plasma triglycerides (TG) are positively linked to the development of coronary heart disease. However, triglycerides circulate in a range of distinct lipoprotein subtractions and the relative atherogenicity of these subtractions is not clear. In this study, three fractions of triglyceride rich lipoprotein (TRL) were isolated from normolipidaemic males according to their differing Svedberg flotation (S-f) rates: chylomicron (CM, S-f > 400), very low-density lipoprotein (VLDL)-1 (S-f 60-400) and VLDL-2 (S-f 20-60). These fractions were incubated with THP-1 monocyte-derived macrophages for determination of cholesterol and TG accumulation, in the presence and absence of the lipoprotein lipase (LPL) inhibitor orlistat. Expression of LDL receptor related protein (LRP) and apolipoprotein B48 receptor (apoB48R) was also examined in both differentiating monocytes, and monocyte-derived macrophages, incubated with TRL. VLDL-I caused a significantly greater accumulation of TG within macrophages compared to VLDL-2. Binding studies also tended to show a greater preference for VLDL-1. No change in expression of LRP or apoB48R was observed in fully differentiated macrophages incubated with VLDL-1, VLDL-2 or CM, although a greater expression of LRP mRNA was observed in differentiating monocytes exposed to VLDL-1, compared to those incubated with CM or VLDL-2. TG loading in response to all three TRL fractions was blocked by orlistat, suggesting that it is likely that the major pathway for uptake of TG was hydrolysis by LPL. Calculations suggested that direct uptake of particles accounts for between 12 and 25% of total TAG uptake. In conclusion, THP monocyte-derived macrophages demonstrate a preference for VLDL-1, both through the LPL pathway and by direct uptake of whole particles. (c) 2005 Elsevier Ireland Ltd. All rights reserved.

Lack of effect of dietary n-6 : n-3 PUFA ratio on plasma lipids and markers of insulin responses in Indian Asians living in the UK

Background: Indian Asians living in Western Countries have an over 50% increased risk of coronary heart disease (CHD) relative to their Caucasians counterparts. The atherogenic lipoprotein phenotype (ALP), which is more prevalent in this ethnic group, may in part explain the increased risk. A low dietary long chain n-3 fatty acid (LC n-3 PUFA) intake and a high dietary n-6 PUFA intake and n-6:n-3 PUFA ratio in Indian Asians have been proposed as contributors to the increased ALP incidence and CHD risk in this subgroup. Aim: To examine the impact of dietary n-6:n-3 PUFA ratio on membrane fatty acid composition, blood lipid levels and markers of insulin sensitivity in Indian Asians living in the UK. Methods: Twenty-nine males were assigned to either a moderate or high n-6:n-3 PUFA (9 or 16) diet for 6 weeks. Fasting blood samples were collected at baseline and 6 weeks for analysis of triglycerides, total-, LDL- and HDL- cholesterol, non-esterified fatty acids, glucose, insulin, markers of insulin sensitivity and C-reactive protein. Results: Group mean saturated fatty acid, MUFA, n-6 PUFA and n-3 PUFA on the moderate and high n-6:n-3 PUFA diets were 26 g/d, 43 g/d, 15 g/d, 2 g/d and 25 g/d, 25 g/d, 28 g/d, 2 g/d respectively. A significantly lower total membrane n-3 PUFA and a trend towards lower EPA and DHA levels were observed following the high n-6:n-3 PUFA diet. However no significant effect of treatment on plasma lipids was evident. There was a trend towards a loss of insulin sensitivity on the high n-6:n-3 PUFA diet, with the increase in fasting insulin (P = 0.04) and HOMA IR [(insulin x glucose)/22.5] (P = 0.02) reaching significance. Conclusion: The results of the current study suggest that, within the context of a western diet, it is unlikely that dietary n-6:n-3 PUFA ratio has any major impact on the levels of LC n-3 PUFA in membrane phospholipids or have any major clinically relevant impact on insulin sensitivity and its associated dyslipidaemia.

Age-related increases in circulating inflammatory markers in men are independent of BMI, blood pressure and blood lipid concentrations

Objective: To examine whether age-related increase in concentrations of circulating inflammatory mediators is due to concurrent increases in cardiovascular risk factors or is independent of these. Methods and results: Cytokines (IL-6, IL-18), chemokines (6Ckine, MCP-1, IP-10), soluble adhesion molecules (sICAM-1, sVCAM-1, sE-selectin) and adipokines (adiponectin) were measured in the plasma of healthy male subjects aged 18-84 years (n = 162). These were related to known cardiovascular risk factors (age, BMI, systolic and diastolic blood pressure, plasma total cholesterol, LDL cholesterol, HDL cholesterol and triacylglycerol concentrations) in order to identify significant associations. Plasma concentrations of sVCAM-1, sE-selectin, IL-6, IL-18, MCP-1, 6Ckine, IP-10 and adiponectin, but not sICAM-1, were significantly positively correlated with age, as well as with several other cardiovascular risk factors. The correlations with other risk factors disappeared when age was controlled for. In contrast, the correlations with age remained significant for sVCAM-1, IL-6, MCP-1, 6Ckine and IP-10 when other cardiovascular risk factors were controlled for. Conclusions: Plasma concentrations of some inflammatory markers (sVCAM-1, IL-6, MCP-L 6Ckine, IP-10) are positively correlated with age, independent of other cardiovascular risk factors. This suggests that age-related inflammation may not be driven by recognised risk factors. (C) 2006 Elsevier Ireland Ltd. All rights reserved.

Effects of oils rich in eicosapentaenoic and docosahexaenoic acids on the oxidizability and thrombogenicity of low-density lipoprotein

Consumption of oily fish and fish oils is associated with protection against cardiovascular disease. Paradoxically, long-chain polyunsaturated fatty acids present in low-density lipoprotein (LDL) are suggested to be susceptible to oxidation. It is not clear whether eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have similar effects on the susceptibility of LDL to oxidation or whether they affect the thrombogenicity of oxidized LDL. This study examined the influence of highly purified preparations of EPA and DHA on LDL oxidizability and LDL-supported thrombin generation in healthy human volunteers. Forty-two healthy volunteers were randomly assigned to receive olive oil (placebo), an EPA-rich oil or a DHA-rich oil for 4 weeks at a dose of 9 g oil/day. EPA and DHA were incorporated into LDL phospholipids and cholesteryl esters during the supplementation period, but were progressively lost during ex vivo copper-mediated oxidation. Following supplementation, the EPA treatment significantly increased the formation of conjugated dienes during LDL oxidation compared with baseline, whereas the DHA treatment had no effect. Neither treatment significantly affected the lag time for oxidation, oxidation rate during the propagation phase or maximum diene production. Neither EPA nor DHA significantly affected the thrombotic tendency of oxidized LDL compared with the placebo, although DHA tended to decrease it. In conclusion, there are subtle differences in the effects of EPA and DHA on the oxidizability and thrombogenicity of LDL. DHA does not appear to increase the susceptibility of LDL to oxidation to the same degree as EPA and has a tendency to decrease LDL-supported thrombin generation. (C) 2004 Elsevier Ireland Ltd. All rights reserved.

Moderate fish-oil supplementation reverses low-platelet, long-chain n-3 polyunsaturated fatty acid status and reduces plasma triacylglycerol concentrations in British Indo-Asians

Background: The mechanisms involved in the increased mortality from coronary artery disease in British Indo-Asians are not well understood. Objectives: This study aimed to investigate whether British Indo-Asian Sikhs have higher plasma triacylglycerol concentrations, lower platelet phospholipid levels, and lower dietary intakes of long-chain n-3 polyunsaturated fatty acids (PUFAs) than do age- and weight-matched Europeans and whether moderate dietary fish-oil intake can reverse these differences. Design: A randomized, double-blind, placebo-controlled, parallel, fish-oil intervention study was performed. After a 2-wk run-in period, 44 Europeans and 40 Indo-Asian Sikhs were randomly assigned to receive either 4.0 g fish oil [1.5 g eicosapentaenoic acid (EPA) and 1.0 g docosahexaenoic acid (DHA)] or 4.0 g olive oil (control) daily for 12 wk. Results: At baseline, the Indo-Asians had significantly higher plasma triacylglycerol, small dense LDL, apolipoprotein B, and dietary and platelet phospholipid n-6 PUFA values and significantly lower long-chain n-3 PUFAs (EPA and DHA) than did the Europeans. A significant decrease in plasma triacylglycerol, plasma apolipoprotein B-48, and platelet phospholipid arachidonic acid concentrations and a significant increase in plasma HDL concentrations and platelet phospholipid EPA and DHA levels were observed after fish-oil supplementation. No significant effect of ethnicity on the responses to fish-oil supplementation was observed. Conclusions: Moderate fish-oil supplementation contributes to a reversal of lipid abnormalities and low n-3 PUFA levels in Indo-Asians and should be considered as an important, yet simple, dietary manipulation to reduce CAD risk in Indo-Asians with an atherogenic lipoprotein phenotype.