OBJECTIVE

Evaluate the consistency of the contribution of interactions between single nucleotide polymorphism (SNP) genotype effects to variation in measures of lipid metabolism across ethnic strata within gender.

RESULTS

We considered 80 SNPs within the apolipoprotein (APO) A1/C3/A4/A5 gene cluster using an over-parameterized general linear model to identify SNPs whose genotype effects combine non-additively to influence plasma levels of high density lipoprotein cholesterol (HDL-C), total cholesterol (TC) and triglycerides (TG) in a consistent manner across ethnic strata. We analyzed population-based samples of unrelated 18 to 30 year old African-Americans (n = 1,858) and European-Americans (n = 1,973) ascertained without regard to health at four field centers (Birmingham, Ala.; Chicago, Ill.; Minneapolis, Minn. and Oakland, Calif., USA) by the Coronary Artery Risk Development in Young Adults (CARDIA) study. To identify which SNP genotype effects combine non-additively we used a two-tier analysis strategy. We first required that pairs of SNPs show statistically significant non-additivity in both ethnic strata within a gender, where experiment-wise significance was evaluated using a permutation test to determine the probability of observing the number of tests significant in both ethnic strata by chance alone. Second, we required no significant evidence of heterogeneity of the relationship between the phenotype and the two SNP genotypes across ethnic strata and across field centers within each ethnic group. From this strategy we identified ten pairs of SNPs, involving thirteen SNPs, that displayed statistically significant non-additivity of SNP genotype effects on TC. Only one of these thirteen SNPs had statistically significant genotype effects that were consistent across samples.

CONCLUSIONS

Our analyses suggest that ignoring the contribution of interactions between SNP genotype effects when modeling multi-SNP genotype-phenotype relationships may result in an underestimate of the contribution of genetic variation to variation in quantitative cardiovascular disease (CVD) risk factor traits.

OBJECTIVE

In this study, we investigated the impact of enhancing cholesterol delivery to mitochondrial sterol 27-hydroxylase, via steroidogenic acute regulatory protein (StAR), on the expression of genes involved in macrophage cholesterol homeostasis and efflux of cholesterol to apolipoprotein (apo) AI.

RESULTS

Stably transfected, murine (RAW 264.7) macrophages were used to investigate the role of StAR in cholesterol homeostasis. Cellular responses were analysed using quantitative PCR, immunoblotting, and an LXRE reporter plasmid; [3H]cholesterol efflux was measured in the presence or absence of apoAI. Macrophage overexpression of mitochondrial cholesterol trafficking protein, StAR, activates and induces expression of liver X receptors (LXRs), and significantly alters expression of genes involved in cholesterol homeostasis, decreasing Fdps, Hmgcr, Mvk, Ldlr, and Scap, and markedly increasing Abca1 mRNA and protein. Overexpression of StAR, but not mutated 'loss-of-function' (R181L) StAR, enhanced efflux of [3H]cholesterol to apoAI, and this effect was maintained in macrophages pretreated with LDL or acetylated LDL. The effect of StAR overexpression on apoAI-dependent [3H]cholesterol efflux was mimicked by non-sterol agonist, T901317, and 27-hydroxycholesterol, and blocked by LXR inhibitor, geranylgeranyl pyrophosphate, sterol 27-hydroxylase inhibitor, GW273297x, and probucol, inhibitor of ATP binding cassette transporter A1 (ABCA1). Importantly, all observed effects of StAR overexpression were dependent upon cyclic AMP (cAMP analogue, dibutyryl cAMP), which is required for the full activity of the StAR protein to be manifested.

CONCLUSIONS

Macrophage overexpression of StAR significantly enhances LXR-dependent apoAI- and ABCA1-dependent cholesterol efflux, by which disposal of excess arterial cholesterol deposits and atheroma regression can be achieved.

Long chain n-3 polyunsaturated fatty acids (n-3 LCPUFA) lower risk of coronary heart disease (CHD), but mechanisms are not well understood. We used proteomics to identify human serum proteins that are altered by n-3 LCPUFA. Such proteins could identify pathways whereby they affect CHD. Eighty-one healthy volunteers entered a double blind randomised trial to receive 3.5 g of fish oil or 3.5 g of high oleic sunflower oil daily. Serum was collected before and after 6 wk of intervention. Serum was analysed by proteomics using 2-DE. Proteins that were differentially regulated were identified by MS. We also analysed serum apolipoprotein A1 (apo A1), high-density lipoprotein (HDL) particle size and haptoglobin. Serum levels of apo A1, apo L1, zinc-alpha-2-glycoprotein, haptoglobin precursor, alpha-1-antitrypsin precursor, antithrombin III-like protein, serum amyloid P component and haemopexin were significantly downregulated (all p<0.05) by fish oil compared with high oleic sunflower oil supplementation. Fish oil supplementation caused a significant shift towards the larger, more cholesterol-rich HDL(2) particle. The alterations in serum proteins and HDL size imply that fish oil activates anti-inflammatory and lipid modulating mechanisms believed to impede the early onset of CHD. These proteins are potential diagnostic biomarkers to assess the mechanisms whereby fish oil protects against CHD in humans.

Our aims were to analyze the protein composition of the organic matrix of urinary stones and to investigate the role of albumin in its constitution. Five different morphological types of stones were studied. Proteins extracted from the stone were submitted to sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and analyzed by immunoblotting with antibodies to 13 urinary proteins. Nine of the 13 proteins were found in all types of stone: human serum albumin (HSA), alpha 1-acid glycoprotein (alpha 1-GP), alpha 1-microglobulin (alpha 1-M), immunoglobulins (Igs), apolipoprotein A1 (apo-A1), transferrin (Tr), alpha 1-antitrypsin (alpha 1-T), retinol-binding protein (RBP) and renal lithostathine (RL). The beta 2-microglobulin (beta 2-M) was present only in calcium oxalate and uric acid stones. In contrast, ceruloplasmin, haptoglobin and Tamm-Horsfall protein (THP) were detected in none of them. Because HSA appeared as the major protein component in all stones, we wondered whether it might play a specific role in the constitution of the stone matrix. Association of HSA with urinary proteins that were present in stones was demonstrated by showing that proteins present in the matrix comigrated with HSA on gel filtration, whereas proteins that were absent did not. Moreover, HSA induced the binding of stone matrix proteins to an albumin-specific affinity column. Finally, we evidenced HSA binding to calcium oxalate monohydrate (COM) crystals in a solution similar to urine.(ABSTRACT TRUNCATED AT 250 WORDS)

The effects of the phenolic compounds catechin (Cat), quercetin (Qer), and resveratrol (Res) present in red wine on early atherosclerosis were studied in hamsters. Hamsters (n = 32) were divided into 4 groups of 8 and fed an atherogenic diet for 12 weeks. They received by force-feeding 7.14 mL/(kg of body wt.day) Cat, Qer, or Res in water [2.856 mg/(kg of body wt.day) for Cat and 0.1428 mg/(kg of body wt.dday) for Qer and Res], mimicking a moderate consumption of alcohol-free red wine (equivalent to that supplied by the consumption of about two glasses of red wine per meal for a 70 kg human), or water as control. Plasma cholesterol concentration was lower in groups that consumed phenolics than in controls. The increase in plasma apolipoprotein (Apo) A1 concentration was mainly due to Cat (26%) and Qer (22%) and to a lesser extent, but nonsignificantly, Res (19%). Apo-B was not affected. Plasma antioxidant capacity was not improved, and there was no sparing effect on plasma vitamins A and E. Plasma iron and copper concentrations were not modified nor were liver super oxide dismutase and catalase activities. A sparing effect of Qer on liver glutathione peroxidase activity appeared, whereas Cat and Res exhibited a smaller effect. Aortic fatty streak area was significantly reduced in the groups receiving Cat (84%) or Qer (80%) or Res (76%) in comparison with the controls. These findings demonstrate that catechin, quercetin, and resveratrol at nutritional doses prevent the development of atherosclerosis through several indirect mechanisms.

It has been reported that micronized fenofibrate and losartan can significantly decrease serum uric acid levels by augmenting uric acid excretion. We undertook this study to evaluate the effects of the combination treatment with micronized fenofibrate and losartan in nondiabetic hypertensive dyslipidemic patients with hyperuricemia (serum uric acid, >7 mg/ dl). A total of 25 patients (15 men, 10 women) aged 21-66 years was studied. In all patients, serum lipid parameters, including Lp(a), fibrinogen, and uric acid levels, as well as fractional excretion of uric acid (FEUA) were obtained before treatment, 8 weeks after micronized fenofibrate treatment (200 mg daily), and 8 weeks after combination therapy with micronized fenofibrate (200 mg daily) and losartan (50 mg daily). Fenofibrate alone significantly decreased total cholesterol, low-density lipoprotein (LDL) cholesterol, triglycerides, Apo B, Lp(a), fibrinogen, and uric acid levels (from 7.6+/-0.55 to 5.6 +/-0.5 mg/dl; p < 0.0001) and increased high-density lipoprotein (HDL) cholesterol, Apo A1, and FEUA (from 6.5+/-1.8% to 12.2+/-4%; p < 0.0001). The addition of losartan beyond the decrease in blood pressure values did not significantly alter serum metabolic parameters. However, a small additional decrease in serum uric acid levels (from 5.6+/-0.5 to 4.9+/-1 mg/dl; p < 0.05) was found because of a further increase in FEUA (from 12.2+/-4% to 14+/-5.5%; p < 0.05). It is concluded that the combination of micronized fenofibrate and losartan is useful for the management of patients with multiple metabolic abnormalities, including hyperuricemia.

OBJECTIVE

Aim of this study was to evaluate implication of pregnancy induced hypertension on maternal plasma lipid, lipoprotein, apolipoprotein concentrations and lipid peroxidation products by a comparison of normal pregnancy vs. preeclampsia.

METHODS

Thirty-four women with preeclampsia and 32 healthy pregnant women (controls) in the third trimester were recruited for this study.

RESULTS

In the preeclamptic group plasma total triglyceride, low density lipoprotein cholesterol (LDL-C), malondialdehyde (MDA) and apolipoprotein B (apo-B) were significantly increased, while plasma high density lipoprotein cholesterol (HDL-C) was significantly decreased compared to that of control group. There was no significant difference in total cholesterol and apolipoprotein A1 (apo-A1) concentrations.

CONCLUSIONS

Our findings suggest that preeclampsia share some metabolic characteristics with coronary artery disease such as dislipidemia and increased lipid peroxidation. However lipoprotein concentrations may be better biochemical markers of dislipidemia in the preeclamptic state than the corresponding apolipoproteins.

OBJECTIVE

To determine the effect of 900 mg/day of dried garlic powder (standardised to 1.3% allicin) in reducing total cholesterol.

METHODS

Double-blind, randomised six-month parallel trial.

METHODS

115 individuals with a repeat total cholesterol concentration of 6.0-8.5 mmol/l and low-density lipoprotein (LDL) cholesterol of 3.5 mmol/l or above after six weeks of dietary advice.

METHODS

The active treatment group received dried garlic tablets (standardised to 1.3% allicin) at a dosage of 300 mg three times daily. The control group received a matching placebo.

METHODS

Primary end-point: total cholesterol concentration; secondary end-points: concentrations of LDL and high-density lipoprotein cholesterol, apolipoproteins (apo) A1 and B, and triglycerides.

RESULTS

There were no significant differences between the groups receiving garlic and placebo in the mean concentrations of serum lipids, lipoproteins or apo A1 or B, by analysis either on intention-to-treat or treatment received. In a meta-analysis which included the results from this trial, garlic was associated with a mean reduction in total cholesterol of -0.65 mmol/l (95% confidence intervals: -0.53 to -0.76).

CONCLUSIONS

In this trial, garlic was less effective in reducing total cholesterol than suggested by previous meta-analyses. Possible explanations are publication bias, overestimation of treatment effects in trials with inadequate concealment of treatment allocation, or a type 2 error. We conclude that meta-analyses should be interpreted critically and with particular caution if the constituent trials are small.

BACKGROUND

TNF-alpha is a critical mediator of inflammation with an important role in metabolic profile and insulin resistance. The regulation of these parameters by TNF-alpha in inflammatory bowel disease (IBD) is, however, poorly understood. The aim of this study was to assess the in-vivo TNF-alpha-mediated regulation of insulin resistance and of lipid levels in patients with IBD.

METHODS

Twenty-two patients with IBD (eight females; 19 Crohn's disease) received infliximab according to treating physician's assessment at weeks 0, 2 and 6 from baseline and subsequently every 8 weeks and were prospectively followed for 14 weeks. Fasting insulin, total cholesterol, triglycerides, HDL cholesterol, LDL cholesterol, apolipoprotein A1 (apo-A1), apolipoprotein B100 and lipoprotein a were measured in serum at baseline and at week 14. Insulin resistance was calculated with the use of the Homeostasis Model Assessment index.

RESULTS

Infliximab therapy induced clinical response or remission in 19 of the 22 patients. C-reactive protein levels were significantly decreased by week 14. Body mass index was increased in all patients. No difference was observed in insulin levels, Homeostasis Model Assessment index, triglycerides, LDL cholesterol, apolipoprotein B100 and lipoprotein a. In contrast, total cholesterol, HDL cholesterol and apo-A1 levels were significantly increased from baseline.

CONCLUSIONS

TNF-alpha inhibition does not alter insulin resistance in IBD patients. In contrast, total cholesterol, HDL cholesterol and apo-A1 levels are significantly increased after infliximab treatment compared with baseline. The importance of these alterations needs to be clarified in future studies.

Fish oil (FO) has traditionally been used as the dominating lipid component in fish feed. However, FO is a limited resource and the price varies considerably, which has led to an interest in using alternative oils, such as vegetable oils (VOs), in fish diets. It is far from clear how these VOs affect liver lipid secretion and fish health. The polyunsaturated fatty acids (PUFAs), eicosapentanoic acid (EPA) and docosahexanioc acid (DHA), reduce the secretion of lipoproteins rich in triacylglycerols (TAGs) in Atlantic salmon, as they do in humans. The mechanism by which n-3 fatty acids (FAs) in the diet reduce TAG secretion is not known. We have therefore investigated the effects of rapeseed oil (RO) and n-3 rich diets on the accumulation and secretion of (3)H-glycerolipids by salmon hepatocytes. Salmon, of approximately 90 g were fed for 17 weeks on one of four diets supplemented with either 13.5% FO, RO, EPA-enriched oil or DHA-enriched oil until a final average weight of 310 g. Our results show that the dietary FA composition markedly influences the endogenous FA composition and lipid content of the hepatocytes. The intracellular lipid level in hepatocytes from fish fed RO diet and DHA diet were higher, and the expressions of the genes for microsomal transfer protein (MTP) and apolipoprotein A1 (Apo A1) were lower, than those in fish fed the two other diets. Secretion of hepatocyte glycerolipids was lower in fish fed the EPA diet and DHA diet than it was in fish fed the RO diet. Our results indicate that EPA and DHA possess different hypolipidemic properties. Both EPA and DHA inhibit TAG synthesis and secretion, but only EPA induces mitochondrial proliferation and reduce intracellular lipid. Expression of the gene for peroxisome proliferator-activated receptor alpha (PPARalpha) was higher in the DHA dietary group than it was in the other groups.

Dietary fatty acids (FA) are the major determinants of blood lipids, and measurements of plasma phospholipid FA (PL-FA) composition that reflect the dietary intake of FA may provide insights into the relationships between diet and CHD. We assessed CHD mortality associations with PL-FA (SFA, PUFA and MUFA) levels measured in a nested case-control study of 116 cases of CHD death and 239 controls that were frequency-matched for age and employment grade. The participants had plasma levels of total cholesterol, LDL-cholesterol (LDL-C), HDL-cholesterol, apo B and apo A1, C-reactive protein (CRP) and fibrinogen recorded. SFA levels were significantly positively correlated with total cholesterol, LDL-C, apo B, CRP protein and fibrinogen. By contrast, phospholipid-PUFA were inversely associated with CRP, but not with any of the lipids. A higher SFA content (top v. bottom quarter) was associated with a 2-fold higher risk of CHD (OR and 95 % CI: OR 2.12; 95 % CI: 1.13, 3.99), and an equivalent difference in PUFA was associated with a halving in CHD risk (OR 0.49; 95 % CI: 0.26, 0.94), but MUFA was unrelated to CHD risk. These associations were substantially attenuated, after additional adjustment for lipids and inflammatory markers. Higher levels of saturated fat and lower levels of polyunsaturated fats were each associated with a higher risk of CHD in elderly men, and these associations were partly explained by their effects on blood lipids and biomarkers of inflammation.

Apo A1 is the major apoprotein of the human plasma high density lipoprotein (HDL). We have isolated apo A1 cDNA and genomic clones and used them to study the gene organisation as defined by its restriction enzyme map. These studies showed that apo A1 is coded by a unique gene. Cross hybridisation was not observed with functionally related apoprotein genes. Increased levels of HDL have been correlated with certain protection against coronary heart disease. If there is a genetic component that contributes to the variable levels of HDL found in the population, it may be possible to correlate these differences with distinct gene organisation patterns.

The caudal ventrolateral medulla (CVL) contains neurons that are vasodepressor and are a critical component of the baroreceptor reflex pathway. While electrophysiological studies suggest that CVL neurons are intercalated in the baroreceptor pathway between the nucleus tractus solitarius (NTS) and the rostral ventrolateral medulla (RVL), there is no direct evidence for this projection. Therefore, we identified CVL neurons that project to RVL by retrogradely labelling them with wheat germ agglutinin-apo-horseradish peroxidase conjugated to colloidal gold (WAHG) injected into the RVL. Retrogradely labelled neurons were seen in previously identified vasodepressor areas of the rostral CVL that are critical for the baroreceptor reflex. Double labelling for WAHG and tyrosine hydroxylase (TH) immunocytochemistry indicated that CVL neurons that project to the RVL (CVL ->> RVL neurons) are distinct from the noradrenergic neurons of the A1 cell group. To establish the presence of a direct projection from the NTS to CVL ->> RVL neurons, the retrograde tracer WAHG was pressure injected into the RVL and the anterograde tracer biocytin was iontophoresed into the NTS of anesthetized rats. After 4-6 h, anesthetized rats were perfused transcardially with 3.75% acrolein in 2% paraformaldehyde and sections through the CVL were processed for both markers. By light microscopy, numerous biocytin-labelled varicose processes overlapped neurons containing WAHG in the CVL. By electron microscopy, biocytin was found in myelinated and unmyelinated axons and in axon terminals (0.9 + 0.02 microns) that contained primarily small clear vesicles. These terminals formed predominantly asymmetric synapses on large (1.5-6.0 microns in diameter) dendrites within the CVL. Some of the post-synaptic perikarya and large dendrites contained WAHG associated with lysosomes and multivesicular bodies, indicating that they belong to neurons which project to the RVL. We conclude that CVL ->> RVL neurons are (a) distinct from A1 noradrenergic cells; (b) receive direct synaptic contacts from NTS efferent terminals; (c) are potently and monosynaptically excited (asymmetric synapses) by NTS efferent terminals. These data support the hypothesis that CVL neurons are intercalated between the NTS and the RVL in the baroreceptor reflex pathway.

Plasma HDL has been reported to protect against the initial development of inflammatory atherosclerosis. Although cellular and molecular mechanisms of this HDL protection are not fully understood, they may involve prevention of the endothelial trafficking of circulating leukocytes via its down-regulation of inflammatory genes. To test this hypothesis, HDL inhibition of monocyte adherence to the endothelium and of the expression of vascular cell adhesion molecule-1 (VCAM-1) and its transcriptional regulatory mechanism were elucidated in human umbilical vein endothelial cells (HUVECs) stimulated by pro-inflammatory TNF-alpha. Plasma HDL (1mg/ml apo A1) markedly prevented the monocyte adhesion to TNF-alpha-activated HUVECs with blunting the increased expression levels of VCAM-1 protein and its mRNA at a maximal TNF-alpha stimulation. Electrophoretic mobility shift assay showed that in the activated HUVECs HDL substantially inhibited DNA binding activities of transcription factors of nuclear factor-kappaB (NF-kappaB) and activator protein-1 (AP-1) that have binding sites in the promoter region of the VCAM-1 gene. In addition, immunocytochemical staining analysis confirmed that HDL inhibited TNF-alpha-induced VCAM-1 expression via reduced nuclear translocation of NF-kappaB. These results suggest that HDL down-regulates the expression of VCAM-1 gene in TNF-alpha-activated HUVECs at transcriptional levels via blunted translocation and transactivation of NF-kappaB and AP-1 transcription factors. Plasma HDL may block the initial atherosclerotic process via inhibited monocyte adhesion to the endothelium, which is independent of modulating the cholesterol reverse transport of plasma HDL.

NPC (Niemann-Pick type C) disease is a progressive neurological disorder characterized by defects in intracellular cholesterol trafficking, accumulation of cholesterol in the endosomal system and impaired cholesterol homoeostasis. Although these alterations appear to occur in all NPC1-deficient cell types, the consequences are most profound in the nervous system. Since glial cells are important mediators of brain cholesterol homoeostasis, we proposed that defective generation and/or function of lipoproteins released by glia might contribute to the neurological abnormalities associated with NPC disease. We found that, as in other cell types, Npc1-/- glia accumulate cholesterol intracellularly. We hypothesized that this sequestration of cholesterol in glia might restrict the availability of cholesterol for lipoprotein production. Cerebellar astroglia were cultured from a murine model of NPC disease to compare the lipoproteins generated by these cells and wild-type glia. The experiments demonstrate that the amount of cholesterol in glia-conditioned medium is not reduced by NPC1 deficiency. Similarly, cholesterol efflux to apo (apolipoprotein) A1 or glial expression of the transporter ATP-binding-cassette transporter A1 was not decreased by NPC1 deficiency. In addition, the ratio of apo E:cholesterol and the density distribution of lipoproteins in Npc1-/- and Npc1+/+ glia-conditioned medium are indistinguishable. Importantly, in a functional assay, apo E-containing lipoproteins generated by Npc1-/- and Npc1+/+ glia each stimulate axonal elongation of neurons by approx. 35%. On the basis of these observations, we speculate that the neuropathology characteristic of NPC disease can quite probably be ascribed to impaired processes within neurons in the brain rather than defective lipoprotein production by astroglia.

OBJECTIVE

To determine if there is an association between low levels of high-density lipoprotein cholesterol (HDL), apolipoprotein A1 (Apo A1), total cholesterol, and anticardiolipin antibody (aCL) in patients with systemic lupus erythematosus (SLE) who are not taking corticosteroids.

METHODS

We studied 75 outpatients with documented SLE who were attending our hospital clinics: 57 were aCL positive and 18 were aCL negative. Both IgG and IgM aCL levels were determined by enzyme-linked immunosorbent assay. Lipid fractions (total cholesterol, HDL, low-density lipoprotein, very-low-density lipoprotein, and triglycerides) were determined by standard enzymatic techniques. Apo A1 and Apo B levels were determined by nephelometry.

RESULTS

Patients with SLE who were IgG aCL+ had low levels of serum cholesterol (mean +/- SD 173.6 +/- 34.6 mg/dl) and HDL (43.9 +/- 16.3 mg/dl) compared with aCL- SLE patients, normal donors, and patients with other diseases. Apo A1 levels were also low in the aCL+ group (95.5 +/- 50.9 mg/dl) compared with the aCL- group (152.7 +/- 32.6 mg/dl). There was no association of total cholesterol level or aCL titer with clinical activity.

CONCLUSIONS

These data indicate that in SLE patients, there is an association between antibody against the phospholipid cardiolipin and low levels of cholesterol, HDL, and Apo A1.

OBJECTIVE

To investigate the lipid profiles in patients with active rheumatoid arthritis (RA) and to assess the relationship of inflammatory disease activity markers, sex, and menopausal status with lipid profiles.

METHODS

Three groups of patients with active RA (n = 184) were studied: men (n = 61, mean age 50.8 +/- 4.81 yrs), premenopausal women (n = 58, mean age 39.2 +/- 2.44 yrs), and postmenopausal women (n = 65, mean age 60.4 +/- 2.14 yrs), and healthy controls (n = 161): men (n = 65, mean age 50.9 +/- 3.42 yrs), premenopausal women (n = 47, mean age 40.3 +/- 1.66 yrs), and postmenopausal women (n = 49, mean age 61.3 +/- 3.16 yrs). We measured fasting plasma levels of total cholesterol (TC), triglyceride (TG), HDL-cholesterol (HDL-C), LDL-cholesterol (LDL-C), lipoprotein (a) [LP(a)], apolipoprotein A1 (apo A1), apolipoprotein B (apo B), and erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP).

RESULTS

Male RA patients had significantly higher apo B/apo A1 and LP(a) and lower HDL-C than male controls. Female RA patients had significantly higher TC, LDL-C, and LP(a) than female controls. Premenopausal RA patients had significantly higher LDL-C, TC/HDL-C, LDL-C/HDL-C, and apo B/apo A1 and lower TG and HDL-C than premenopausal controls. Postmenopausal RA women had significantly higher TG and LP(a) and lower TC than postmenopausal controls. Female RA patients had higher HDL-C, apo A1, and TC/HDL-C and lower apo B/apo A1 than male RA patients. Postmenopausal RA patients had significantly higher TC, TG, TC/HDL-C, apo B, LP(a), and LDL-C/HDL-C than premenopausal RA patients. CRP correlated positively with TC/HDL-C, LDL-C/HDL-C, and apo B/apo A1 and negatively with HDL-C in male RA patients. In female RA patients CRP had positive correlation with TC/HDL-C and LDL-C/HDL-C and negative correlation with HDL-C.

CONCLUSIONS

These findings suggest that patients with active RA have altered lipid profiles and that disease activity, sex, and menopausal status affect lipid profiles, and these would be expected to change the pattern of atherosclerotic events in RA.

In a sample of Dutch families consisting of parents aged 35-65 years and their twin offspring aged 14-21 years, a significant difference between generations was observed in phenotypic variances and in genetic heritabilities for plasma levels of total cholesterol, triglycerides, high density lipoprotein (HDL) and low density lipoprotein (LDL) cholesterol, and apolipoproteins (apo) A1, A2, B, and E. For all traits parents were more variable than their offspring. This increase in phenotypic variance was best explained by a genetic model in which individual specific environmental variance increased with increasing age. Genetic variance was the same across generations for nearly all traits except triglycerides and apoE, for which a decrease in genetic variance was observed. This model led to large intergenerational differences in genetic heritabilities. Heritabilities for children were between 65 and 87%, while heritabilities for their parents were between 10 and 50%. No evidence was found for effects of a shared family environment.

We have investigated the effects of two fibric acid derivatives, bezafibrate mono (400 mg daily) and gemfibrozil (600 mg b.d.), in 29 patients with type IIb hyperlipoproteinaemia. All patients received placebo and each drug for 8 weeks in randomised order in a double-blind, cross-over study designed to evaluate any different effects of the drugs on serum lipoproteins, cholesteryl ester transfer protein (CETP), cholesteryl ester transfer activity (CETA), plasma fibrinogen, plasminogen activator inhibitor-I (PAI-1) or paraoxonase. Serum cholesterol decreased (P < 0.05) with gemfibrozil, but the effect of bezafibrate on serum cholesterol did not achieve statistical significance (placebo 8.34 +/- 1.05 (mean +/- S.D.), gemfibrozil 7.70 +/- 1.23 and bezafibrate 7.8 +/- 1.37 mmol/l). Both drugs decreased the serum triglyceride concentration (both P < 0.001) (placebo 4.39 (3.13-5.75) (median (interquartile range)), bezafibrate 2.26 (1.89-3.89) and gemfibrozil 2.00 (1.30-3.30) mmol/l) and very low density lipoprotein (VLDL) cholesterol (both P < 0.001) (placebo 1.18 (0.74-2.30), bezafibrate 0.59 (0.34-0.85) and gemfibrozil 0.48 (0.34-0.68) mmol/l). Discontinuous gradient ultracentrifugation (DGU) revealed that Sf 60-400 (large VLDL) decreased by more than 50% and Sf 20-60 (small VLDL) by more than 30% with each of the drugs (both P < 0.001), neither of which affected the composition of these lipoproteins. Gemfibrozil decreased the concentration of Sf 12-20 lipoprotein (intermediate density lipoprotein; IDL) by 23% (P < 0.01), whereas the effect of bezafibrate on this lipoprotein did not achieve statistical significance. Neither drug altered the concentration of apolipoprotein B or of total Sf 0-12 lipoproteins (low density lipoprotein, (LDL)). Both, however, significantly increased the quantity of free cholesterol in Sf 0-12 lipoproteins (P < 0.05). Overall the concentration of triglycerides decreased significantly in all lipoproteins isolated by DGU (Sf 0-12, Sf 12-20, Sf 20-60, Sf 60-400) on gemfibrozil treatment, but only in Sf 20-60 and Sf 60-400 on bezafibrate (all P < 0.05). Both drugs also increased serum high density lipoprotein (HDL) cholesterol (placebo 1.15 +/- 0.29, bezafibrate 1.27 +/- 0.38 (P < 0.01) and gemfibrozil 1.26 +/- 0.49 (P < 0.05) mmol/l) and HDL3 cholesterol concentration (placebo 0.59 +/- 0.12, bezafibrate 0.72 +/- 0.23 (P < 0.001) and gemfibrozil 0.70 +/- 0.24 (P < 0.01) mmol/l). Serum apolipoprotein A1 (apo A1) was increased (P < 0.05) by bezafibrate compared to gemfibrozil (placebo 103 +/- 26, bezafibrate 111 +/- 28 and gemfibrozil 102 +/- 25 mg/dl) and CETA from HDL to VLDL and LDL was decreased (P < 0.05) by bezafibrate compared to placebo, but the apparent decrease with gemfibrozil did not achieve statistical significance (placebo 39.6 +/- 17.7, bezafibrate 32.3 +/- 14.7 and gemfibrozil 33.8 +/- 15.0 nmol/ml/h). Neither drug affected the circulating concentration of CETP. Plasma fibrinogen was increased (P < 0.05) by gemfibrozil (placebo 4.16 (3.38-4.71) and gemfibrozil 4.65 (4.05-5.77) g/l) and was significantly lower (P < 0.001) on bezafibrate (3.60 (3.18-4.54) g/l) than on gemfibrozil treatment. There was a significant (P < 0.05) increase in PAI-1 activity with bezafibrate and a similar trend with gemfibrozil (placebo 41.2 (25.6-64.5), bezafibrate 50.5 (35.1-73.9) and gemfibrozil 48.5 (31.5-5.4 U/l). Neither fibrate influenced plasma concentrations of PAI-1 nor were the activities of lecithin:cholesterol acyl transferase or paraoxonase affected. The major difference in the action of the two drugs on lipoprotein metabolism was the greater effect of gemfibrozil in decreasing the overall serum concentration of Sf 12-20 lipoproteins and the triglycerides in Sf 12-20 and 0-12 lipoproteins. Bezafibrate, however, increased serum apo A1 concentration and significantly decreased CETA. The two drugs also had different effects on the plasma fibrinogen levels, which increased with gemfibrozil and tended to decrea

BACKGROUND

The association of single nucleotide polymorphisms (SNPs) in the apolipoprotein (Apo) A1/C3/A4/A5 gene cluster and serum lipid profiles is inconsistent. The present study was undertaken to detect the association between the ApoA1/C3/A5 gene polymorphisms and their haplotypes with serum lipid levels in the general Chinese population.

METHODS

A total of 1030 unrelated subjects (492 males and 538 females) aged 15-89 were randomly selected from our previous stratified randomized cluster samples. Genotyping of the ApoA1 -75 bp G>A, ApoC3 3238C>G, ApoA5 -1131T>C, ApoA5 c.553G>T and ApoA5 c.457G>A was performed by polymerse chain reaction and restriction fragment length polymorphism combined with gel electrophoresis, and then confirmed by direct sequencing. Pair-wise linkage disequilibria and haplotype analysis among the five SNPs were estimated.

RESULTS

The levels of high-density lipoprotein cholesterol (HDL-C) and ApoA1 were lower in males than in femailes (P < 0.05 for each). The allelic and genotypic frequencies of the SNPs were no significant difference between males and females except ApoC3 3238C>G. There were 11 haplotypes with a frequency >1% identified in the cluster in our population. At the global level, the haplotypes comprised of all five SNPs were significantly associated with all seven lipid traits. In particular, haplotype G-G-C-C-A (6%; in the order of ApoA5 c.553G>T, ApoA5 c.457G>A, ApoA5 -1131T>C, ApoC3 3238C>G, and ApoA1 -75bp G>A) and G-A-T-C-G (4%) showed consistent association with total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), ApoA1, ApoB, and the ApoA1/ApoB ratio. In addition, carriers of haplotype G-G-T-C-G (26%) had increased serum concentration of HDL-C and ApoA1, whereas carriers of G-G-C-G-G (15%) had high concentrations of TC, triglyceride (TG) and ApoB. We also found that haplotypes with five SNPs explain much more serum lipid variation than any single SNP alone, especially for TG (4.4% for haplotype vs. 2.4% for -1131T>C max based on R-square) and HDL-C (5.1% for haplotype vs. 0.9% for c.553G>T based on R-square). Serum lipid parameters were also correlated with genotypes and several environment factors.

CONCLUSIONS

Several common SNPs and their haplotypes in the ApoA1/C3/A5 gene cluster are closely associated with modifications of serum lipid parameters in the general Chinese population.

The detection and quantitation of apolipoproteins, important markers for coronary heart disease, in serum by liquid chromatography/tandem mass spectrometry (LC/MS/MS) using multiple reaction monitoring (MRM) is reported. A tryptic digest of depleted human serum was analysed by nanoflow LC/MS/MS at a flow rate of 300 nL/min and several apolipoproteins (Apo), including Apo A1, A2, A4, C1, C2, C3, D, F and M, were successfully identified. The analysis of the same depleted serum digest by ultra-performance (UP)LC/MS/MS operating at 700 microL/min resulted in comparable sensitivity and selectivity to the nanoflow method, but with a dramatic ( approximately 20-fold) reduction in run time. The potential of UPLC/MS/MS for the rapid quantitation of proteins in biological matrices by representative tryptic peptides was further investigated using Apo A1 and its corresponding stable isotopically labelled tryptic AQUA peptide (DYVSQFEGSALGK). A set of serum-based Apo A1 calibrators from a clinical analyser kit were digested without depletion following the addition of the AQUA peptide and analysed using UPLC/MS/MS. A linear calibration curve was generated from peak area ratios to the labelled peptide with a coefficient of correlation of 0.9989. Standard curves were also generated for other apolipoproteins together with Apo B100, Apo E, lecithin cholesterol acyltransferase and albumin, which were also detected in the standards. The concentration of Apo A1 in five fresh undepleted human serum samples and a quality control (QC) sample were determined using both the UPLC/MS/MS method and a clinical analyser. Results were comparable and the quantitative study, involving 80 injections which took hours rather than days to complete, demonstrates the high-throughput potential of UPLC/MS/MS to quantify multiple serum proteins without the need for antibodies, and thus provide an alternative to the use of clinical analysers for serum protein biomarkers.

UNASSIGNED

Exosomes are closed-membrane nanovesicles that are secreted by a variety of cells and exist in most body fluids. Recent studies have demonstrated the potential of exosomes as natural vehicles that target delivery of functional small RNA and chemotherapeutics to diseased cells.

UNASSIGNED

In this study, we introduce a new approach for the targeted delivery of exosomes loaded with functional miR-26a to scavenger receptor class B type 1-expressing liver cancer cells. The tumor cell-targeting function of these engineered exosomes was introduced by expressing in 293T cell hosts, the gene fusion between the transmembrane protein of CD63 and a sequence from Apo-A1. The exosomes harvested from these 293T cells were loaded with miR-26a via electroporation.

UNASSIGNED

The engineered exosomes were shown to bind selectively to HepG2 cells via the scavenger receptor class B type 1-Apo-A1 complex and then internalized by receptor-mediated endocytosis. The release of miR-26a in exosome-treated HepG2 cells upregulated miR-26a expression and decreased the rates of cell migration and proliferation. We also presented evidence that suggest cell growth was inhibited by miR-26a-mediated decreases in the amounts of key proteins that regulate the cell cycle.

UNASSIGNED

Our gene delivery strategy can be adapted to treat a broad spectrum of cancers by expressing proteins on the surface of miRNA-loaded exosomes that recognize specific biomarkers on the tumor cell.

Alzheimer's disease (AD) brains contain high levels of advanced glycation endproducts (AGEs). Double immunostaining using anti-AGE and anti-apolipoprotein E (apoE) antibodies demonstrated that AGEs co-localized to a very high degree with apoE. We examined the binding of apoE to in vitro-prepared AGE-bovine serum albumin (AGE-BSA), using Western ligand blot analysis. ApoE exhibited AGE-specific binding activity in the presence of excess native BSA, with the dimeric form of apoE binding better than the monomeric form. Other apolipoproteins including apo A1, B, CI and CII, and serum beta2-microglobulin, did not bind AGE-BSA. ApoE4 exhibited a 3-fold greater AGE-binding activity than the apoE3 isoform. These results suggest that apoE may participate in aggregate formation in the AD brain by binding to AGE-modified plaque components. It is possible that enhanced binding of apoE4 might have pathogenic consequences in vivo.

OBJECTIVE

To examine the recent advances in our knowledge of HDL metabolism, composition, function, and coronary heart disease (CHD), as well as marked HDL deficiency states because of mutations in the apolipoprotein (apo) A-I, ATP-binding cassette transfer protein A1 and lecithin cholesterol acyltransferase (LCAT) gene loci.

RESULTS

It has been documented that apoA-I, myeloperoxidase and paraoxonase 1 (PON1) form a complex in HDL that is critical for HDL binding and function. Myeloperoxidase has a negative impact on HDL function, whereas PON1 has a beneficial effect. Patients who lack apoA-I develop markedly premature CHD. Patients who lack ATP-binding cassette transfer protein A1 transporter function have only very small discoidal preβ-1 HDL, and develop hepatosplenomegaly, intermittent neuropathy and premature CHD, although significant heterogeneity for these disorders has been reported. Patients with LCAT deficiency have abnormal small discoidal LDLs and HDL particles, and develop kidney failure. Enzyme replacement therapy is being developed for the latter disorder.

CONCLUSIONS

Recent data indicates that proteins other than apoA-I and apoA-II such as MPO and PON1 have important effects on HDL function. There has been considerable recent progress made in our understanding of HDL protein content and function.