Compared effect of immunosuppressive drugs cyclosporine A and rapamycin on cholesterol homeostasis key enzymes CYP27A1 and HMG-CoA reductase.

Hyperlipidaemia, i.e. increase in total cholesterol and triglycerides, is a common side-effect of the immunosuppressive drugs rapamycin (RAPA) and cyclosporine A (CsA), and is probably related to inhibition of the 27-hydroxylation of cholesterol (acid pathway of bile acid biosynthesis). This might be one of the causes for the increase in plasma cholesterol, as 27-hydroxycholesterol is a potent suppressor of 3-hydroxy-3-methyl-glutaryl-CoA reductase (HMGR), a key enzyme of cholesterol synthesis. As the sterol 27-hydroxylase (CYP27A1) inhibition by CsA is well known, we evaluated the effect of another immunosuppressive drug, RAPA, on this enzyme in HepG2 mitochondria, which confirmed the dose-dependent inhibition of mitochondrial CYP27A1 by cyclosporine (10-20 microM), while the inhibition by RAPA required a higher dose (50-100 microM). Corresponding K(i) was 10 microM for CsA (non-competitive inhibition) and 110 microM for RAPA (competitive inhibition). Cotreatment with both immunosuppressive drugs showed an additive inhibitory effect on CYP27A1 activity. Later, we analysed the effect of these immunosuppressants on HMGR expression in HepG2 cells, and a dose-dependent up-regulation of HMGR gene expression was observed. The results suggest that RAPA and CsA are both inhibitors of CYP27A1 activity with slightly different mechanisms and that they may accordingly increase HMGR expression.

Biological variations, genetic polymorphisms and familial resemblance of TNF-alpha and IL-6 concentrations: STANISLAS cohort.

Cytokines are involved in the development of several inflammatory diseases and atherosclerosis. Their variations in healthy individuals are not well defined. The aims of this study were: firstly, to identify factors affecting biological variation of interleukin-6 (IL-6) and tumour necrosis factor-alpha (TNF-alpha); secondly, to study their family resemblance; and thirdly, to evaluate the effect of two TNF-alpha (-308G/A and -238G/A) and two IL-6 polymorphisms (174G/C and -572G/C) on their corresponding circulating levels. A total of 171 healthy families selected from the STANISLAS cohort were studied. Age was negatively related to TNF-alpha concentrations in offspring only (both sons and daughters). Additionally, IL-6 and TNF-alpha levels were differently influenced by gender, white blood cells, tobacco consumption, and HDL-cholesterol level. A weak significant familial resemblance for TNF-alpha concentration was observed in siblings only. There was no significant familial resemblance for IL-6 levels. The TNF-alpha -308A allele was associated with decreased TNF-alpha concentrations in both offspring aged less than 18 and males without overweight (BMI<25 kg/m(2)). Fathers carrying the IL-6 -174CC genotype had higher IL-6 levels than those with the IL-6 -174G allele. Parents with the IL-6 -572GG genotype had higher IL-6 concentrations than the C allele carriers. In this sample of healthy families, plasma levels of IL-6 and TNF-alpha were differently affected by biological parameters including age, gender and smoking, and the impact of their respective polymorphisms was influenced by gender, age and BMI.

Genetic and environmental contributions to serum retinol and alpha-tocopherol concentrations: the Stanislas Family Study.

BACKGROUND: Although numerous environmental factors are documented to influence serum retinol and alpha-tocopherol concentrations, little is known about the genetic versus the environmental contributions to variations in these traits. OBJECTIVE: The aim of this study was to estimate additive genetic heritability and household effects for serum retinol and alpha-tocopherol concentrations in a variance component analysis. DESIGN: In a sample of 387 French families, information on serum retinol and alpha-tocopherol concentrations, usual dietary intake, lifestyle, and serum lipid profiles and related polymorphisms (apolipoprotein E, apolipoprotein C-III, apolipoprotein B, cholesteryl ester transfer protein, and lipoprotein lipase) was obtained. RESULTS: For serum retinol--after adjustment for sex, age, body mass index, alcohol consumption, oral contraceptive use, and serum albumin, triacylglycerol, and apolipoprotein A-I concentrations--additive genetic effects and shared common environment contributed 30.5% and 14.2% of the total variance, respectively. For serum alpha-tocopherol, approximately 22.1% of the total variance was due to the additive effects of genes and 18.7% to those of household environment, after adjustment for the covariates sex, age, vitamin E intake, oral contraceptive use, and cholesterol, triacylglycerol, and apolipoprotein A-I concentrations. For both vitamins, the influence of measured polymorphisms was not significant. Moreover, heritability and household effect estimates were not significantly different between the 4 classes of relatives and did not vary significantly when families shared more meals at home. CONCLUSIONS: The results show that serum retinol and alpha-tocopherol concentrations are under genetic control in healthy families.

Effect of acute and chronic psychostimulant drugs on redox status, AP-1 activation and pro-enkephalin mRNA in the human astrocyte-like U373 MG cells.

In order to approach the astroglial implication of addictive and neurotoxic processes associated with psychostimulant drug abuse, the effects of amphetamine or cocaine (1-100 microM) on redox status, AP-1 transcription factor and pro-enkephalin, an AP-1 target gene, were investigated in the human astrocyte-like U373 MG cells. We demonstrated an early increase in the generation of radical oxygen species and in the formation of 4-hydroxynonenal-adducts reflecting the pro-oxidant action of both substances. After 1 h or 96 h of treatment, Fos and Jun protein levels were altered and the DNA-binding activity of AP-1 was increased in response to both substances. Using supershift experiments, we observed that the composition of AP-1 dimer differed according to the substance and the duration of treatment. FRA-2 protein represented the main component of the chronic amphetamine- or cocaine-activated complexes, which suggests its relevance in the long-term effects of psychostimulant drugs. Concomitantly, the pro-enkephalin gene was differently regulated by either 6 h or 96 h of treatment. Because astrocytes interact extensively with the neurons in the brain, our data led us to conclude that oxidation-regulated AP-1 target genes may represent one of the molecular mechanisms underlying neuronal adaptation associated with psychostimulant dependence.

Charge-based heterogeneity of human plasma lipoproteins at hypertriglyceridemia: capillary isotachophoresis study.

To reveal the metabolic links between and within pools of pro-atherogenic triglyceride(TG)-rich lipoproteins and anti-atherogenic high density lipoproteins (HDL), the changes in lipoprotein profile at hypertriglyceridemia were analyzed by capillary isotachophoresis. Plasma samples from patients with apoE3/3 phenotype were stained with a fluorescent probe NBD-C6-ceramide and lipoproteins resolved into six H-, one (V+I) and four L-components which belong to HDL, very low and intermediate density (VLDL+IDL) and low density lipoproteins (LDL), respectively. The expected correlation between the relative size of the combined fractions and lipid and apolipoprotein values was obtained confirming the validity of the approach. The new findings were obtained as follows. (1) The fast L-component correlated inversely with HDL-cholesterol (Chol), while intermediate and slow H-components correlated inversely with plasma and LDL-Chol and apoB. (2) The content of intermediate and slow H-components increased within H-pool and decreased relative TG-rich lipoproteins as hypertriglyceridemia rose due to the impairment of triglyceride hydrolysis by lipoprotein lipase within TG-rich particles. (3) A predictive value of the ratios of fast to slow H-components as an indicator of lecithin:cholesterol acyltransferase activity was demonstrated which tended to decrease at hypertriglyceridemia. (4) The L1/L2 ratio may be considered as an indicator of the accumulation of small dense LDL, which is a feature of clinically manifested atherogenic B-pattern. The competition between H(DL) and L(DL) particles for hepatic lipase and significant contribution of apoE to functional deficiency of H(DL) particles at hypertriglyceridemia are suggested.

Association between Gly241Arg ICAM-1 gene polymorphism and serum sICAM-1 concentration in the Stanislas cohort.

Intracellular adhesion molecule-1 (ICAM-1), a cellular adhesion molecule that mediates the interaction of activated endothelial cells with leukocytes, is involved in various inflammatory and cardiovascular disorders. The relation between these markers and genetic polymorphism, however, remains to be elucidated. The aim of this study is to estimate the effect of a single-base polymorphism at codon 241 in exon 4 of ICAM-1 gene on serum sICAM-1 concentration in a healthy population, taking into account other biological determinants of sICAM-1 level. Serum sICAM-1 levels and the G/R241 polymorphism of the ICAM-1 gene were measured in a large healthy population consisting of 413 children aged 6-21 years and 363 adults aged 38-55 years extracted from the Stanislas cohort. The R241 allele was significantly associated with lower sICAM-1 levels and explained 3.4 and 1.9% of the sICAM-1 variability in children and adults, respectively. A codominant pattern contributed better to the model after adjustment for covariates as the RR homozygote effect was higher than that of the GR heterozygote. Moreover, significant independent associations were found between sICAM-1 and smoking, insulin resistance index (HOMA IR), interleukin-6 level, and alkaline phosphatase and aspartate aminotransferase activities. In conclusion, this study revealed a significant association between the G/R241 ICAM-1 polymorphism and serum sICAM-1 levels, probably due to the impairment in binding of ICAM-1 to leukocyte integrin Mac-1 protein.

Detection of putative functional angiotensinogen (AGT) gene variants controlling plasma AGT levels by combined segregation-linkage analysis.

Previous studies have suggested that angiotensinogen (AGT) gene variants are associated with increased plasma AGT levels, and may also contribute towards the inherited component of predisposition to essential hypertension in humans. To explore the potential functionality of several AGT polymorphisms and estimate their effects, together with other sources of familial correlations, on plasma AGT, we undertook a large study involving 545 healthy French volunteers in 130 nuclear families that include 285 offspring. Plasma AGT levels were measured in all participants, and bi-allelic AGT variants were analysed as candidate functional variants at three sites in the 5'-flanking region (C-532T, A-20C, G-6A), two sites in exon 2 (M235T, T174M) and two newly identified variant sites in the untranslated sequence of exon 5 and the 3'-flanking region (C+2054A, C+2127T) of the gene. Analysis with the class D regressive model showed significant effects influencing plasma AGT levels of all AGT polymorphisms tested, with the exception of T174M. The most significant result was found at C-532T (P=0.000001), which accounts for 4.3% of total plasma AGT variability in parents and 5.5% in offspring, with substantial residual familial correlations. Maximum likelihood estimates of haplotype frequencies and tests of linkage disequilibrium between each AGT polymorphism and a putative QTL are in agreement with a complete confounding of C-532T with the QTL, when taking into account sex and generation specific effects of the QTL. However, further combined segregation-linkage analyses showed significant evidence for additional effects of G-6A, M235T and C+2054A polymorphisms after accounting for C-532T, which supports a complex model with at least two functional variants within the AGT gene controlling AGT levels.

High-resolution genetic mapping of the ACE-linked QTL influencing circulating ACE activity.

Fine-mapping of trait loci through combined linkage and association analysis is an important component of strategies designed to identify causative gene variants, particularly in situations where the trait may be influenced by one or more of many polymorphisms within the same gene. Angiotensin-1 converting enzyme (ACE) provides one of the best models for developing and testing such methodologies, as a major fraction of the heritable variation in the activity of the angiotensin-1 converting enzyme (ACE) is tightly linked to the ACE gene. Moreover, ACE contains many frequent polymorphisms that are in strong linkage disequilibrium with each other. Although none of these variants induces a significant amino-acid change, one or more, either singly or in combination, are likely to have a strong effect on the quantitative phenotype. Here, we show that measured-haplotype analysis of SNP data from a large European family cohort can be used to localise the major ACE-linked genetic factors influencing the trait to a 16 kb interval within the gene, thus limiting the number of ACE variants that need to be considered in future studies designed to elucidate their biological effects. The approaches developed will be applicable to the fine-mapping of other quantitative trait loci in humans.

Biological determinants of serum ICAM-1, E-selectin, P-selectin and L-selectin levels in healthy subjects: the Stanislas study.

BACKGROUND: Circulating levels of adhesion molecules increase in various inflammation-related diseases, such as atherosclerosis. However, data about factors influencing their concentrations in physiological conditions are scarce. METHODS: We have studied the determinants of serum levels of intercellular adhesion molecule-1 (ICAM-1), E-selectin, P-selectin and L-selectin in a sample of healthy individuals: 303 children (4-17 years) and 493 adults (18-55 years). The concentrations of these molecules have been measured by enzyme-linked immunosorbant assay. RESULTS: As far as the children are concerned, a decrease in the levels of ICAM-1, E-selectin, P-selectin and L-selectin has been noticed for both boys and girls aged 4-17 years, without any difference between genders. For the adults, no age-related variation has been found for the ICAM-1, E-selectin and P-selectin levels, while the L-selectin level decreased until 55 years old. In the adult group, no sex-related difference in the concentrations of ICAM-1, E-selectin and L-selectin has been seen. As to the P-selectin level, men had significantly higher levels than women. Multiple regression analysis showed that smoking, homeostasis model assessment (HOMA) index, aspartate aminotransferase (AST), alkaline phosphatase (ALP) and C-reactive protein (CRP) were significant positive determinants of the ICAM-1 concentration, whereas age and apo AI were negative ones. The E-selectin level was positively associated with body mass index (BMI), leukocyte, platelet and erythrocyte counts, glucose, ALP and tumor necrosis factor-alpha (TNF-alpha), and negatively related to the use of oral contraceptive (OC). Positive determinants of the P-selectin concentration were leukocyte, platelet and erythrocyte counts, whereas sex, the use of oral contraceptive, glucose and TNF-alpha were negative determinants of P-selectin. Only two determinants have been noticed for the concentration of serum L-selectin: age, which was negatively correlated, and leukocyte count, which was positively associated. CONCLUSION: Our study contributes to the understanding of the regulation of adhesion molecules in physiological conditions.

IL-6, TNF-alpha and atherosclerosis risk indicators in a healthy family population: the STANISLAS cohort.

There are no satisfactory data on circulating concentrations of inflammatory cytokines and their potential relationship with traditional and nontraditional atherosclerosis risk factors in a large healthy young population. The present study was conducted to examine, in 179 healthy families selected from the STANISLAS cohort, the association between interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), C-reactive protein (CRP), orosomucoid, haptoglobin, cell-adhesion molecules (ICAM-1, E-, L- and P-selectin) and lipid parameter concentrations. Age, BMI, white blood cells and tobacco consumption contributed to the variation of IL-6 concentrations. Age and tobacco contributed also to TNF-alpha variation. Taking into account potential covariates, we showed strong positive correlation between IL-6 and both inflammatory markers TNF-alpha and CRP in parents and in offspring (P<0.001). In parents, IL-6 was associated with ICAM-1 and L-selectin (P<0.01), while IL-6 and TNF-alpha predicted E-selectin in offspring only (0.001

Myeloperoxidase polymorphisms in brain infarction. Association with infarct size and functional outcome.

Myeloperoxidase (MPO) has been shown to contribute to several diseases and more particularly to atherosclerosis through excessive ROS production via the MPO/H(2)O(2)/Cl(-) oxidation system. The aim of this study was to determine whether there is an association between MPO polymorphisms and brain infarction (BI), one of the main consequences of atherosclerosis. We investigated MPO G-463A and G-129A polymorphisms in 450 patients with BI confirmed by magnetic resonance imaging (MRI) and 450 controls of the GENIC (Génétique de l'Infarctus Cérébral) Study. Genotype determination of MPO was performed by polymerase chain reaction and allele-specific oligonucleotide hybridization (ASO). Genotype distributions for each of both MPO polymorphisms were found to be similar between cases and controls overall, and according to etiologic subtypes or gender. The frequency of the A allele of the G-463A polymorphism was 22% (95% confidence interval, 19.4 to 24.9) and the frequency of the A allele of the G-129A polymorphism was 6.8% (95% confidence interval, 5.3 to 8.6). The odds ratio (OR) for BI in carriers of the A allele of the G-129A polymorphism was 0.92 (95% confidence interval, 0.61 to 1.39), and the OR for BI in carriers of the A allele of the G-463A polymorphism was 1.15 (95% confidence interval, 0.88 to 1.52). No association between the main risk factors for BI such as hypertension, cholesterol, diabetes and MPO polymorphisms was found. In analyses restricted to cases, we identified an association between the A allele of the G-129A polymorphism and the size of the brain infarct (P=0.01). Furthermore, the A allele of the G-463A polymorphism was associated with a poorer functional short-term outcome as evaluated by the Rankin score (P=0.02). In conclusion, MPO polymorphisms were associated with the extent of brain damage and the functional outcome rather than with the risk of developing a BI.

Genetic influences on blood pressure within the Stanislas Cohort.

OBJECTIVE: To investigate the association of 21 polymorphisms within 13 genes, APOE, APOB, APOC3, CETP, LPL, PON1, MTHFR, FGB, F5, GPIIIa, SELE, ACE and AGT, with inter-individual blood pressure (BP) variation. PARTICIPANTS: Seven hundred and seventy-six men and 836 women, free of antihypertensive and lipid-lowering medications, were selected from the Stanislas Cohort. RESULTS: ANOVA on blood pressure values after adjustment for covariates [age, body mass index (BMI), contraceptive pill, tobacco and alcohol] showed that lipoprotein lipase (LPL) Ser447Ter and glycoprotein IIIA (GpIIIa) Pl polymorphisms were significantly associated with BP in women (0.01 < or = P < or = 0.05), whereas BP levels in men were significantly different according to apolipoprotein CIII (APOC3) 3206T/G and -482C/T polymorphisms (P < or = 0.05). In women, compared to the most common allele, the GpIIIa Pl allele was associated with increased mean arterial pressure (MAP) (P < 0.05) and pulse pressure (PP) (P < 0.001), and the LPL 447Ter allele was associated with decreased systolic blood pressure (SBP) and PP levels (0.001 < or = P < or = 0.05). These two polymorphisms appeared to act independently. In men, the APOC3 3206GG genotype was related to decreased diastolic blood pressure (DBP) and MAP levels (P < or = 0.01), and the APOC3 -482T allele with decreased PP levels (P < or = 0.05). The presence of both the -482C allele and the 3206GG genotype was related to decreased DBP, suggesting that specific haplotypes might be involved. CONCLUSION: The APOC3, LPL and GpIIIa genes were found to be associated with BP levels. The contributions of these genes, although modest, are consistent with the polygenic nature of BP levels.

The Leu554Phe polymorphism in the E-selectin gene is associated with blood pressure in overweight people.

BACKGROUND: Associations between circulating concentrations of E-selectin, blood pressure and obesity, and between the Leu554Phe (L/F554) polymorphism and blood pressure have been documented. OBJECTIVES: To investigate how the E-selectin L/F554 polymorphism is involved in longitudinal blood pressure changes, and how this polymorphism interacts with body mass index (BMI) on blood pressure. DESIGN AND PARTICIPANTS: For this study, 478 men and 546 women were selected from the Stanislas cohort, a French longitudinal study of volunteers for a free health check-up. These individuals underwent two examinations (t(0) and t(+5)) and were not taking medication that can affect blood pressure. RESULTS: At t(0), no relationship was observed between L/F554 polymorphism and blood pressure. However at t(+5), systolic blood pressure (SBP) was greater in individuals carrying the F allele, and the L/F554 polymorphism was associated with SBP in interaction with BMI (P < 0.001 in men and P < 0.05 for women). There was a steeper increase in SBP with BMI greater than 25 kg/m2 in carriers of the F allele than in LL homozygotes. Similar results were observed for diastolic blood pressure in men (P = 0.0103). CONCLUSION: These results suggest a BMI-specific effect of L/F554 polymorphism of the E-selectin gene on blood pressure, and strengthen the hypothesis that E-selectin is implicated in hypertension.

Family study of the relationship between height and cardiovascular risk factors in the STANISLAS cohort.

BACKGROUND: Short stature is a risk factor for coronary heart disease and is associated with an adverse cardiovascular profile. Mechanisms responsible for this association remain unknown. A genetic contribution to this association would imply a familial clustering between height and cardiovascular risk factors. METHODS: This study investigated whether lipids and blood pressure (BP) levels shared a common familial component with height. The sample included 865 nuclear families from the French STANISLAS cohort volunteering for a free health examination between 1993 and 1994. Within-individual correlations and familial intra-trait and cross-trait correlations were estimated using the Estimating Equation technique extended to a bivariate phenotype. RESULTS: Height negatively correlated to total and low density lipoprotein cholesterol (LDL-C) and triglycerides in both parents and offspring, and positively correlated to high density lipoprotein cholesterol (HDL-C) in parents only. In offspring, the correlation between height and HDL-C markedly increased with sexual maturation to reach after puberty the same value as in parents. The correlation of height with systolic BP was negative in fathers and positive in sons, whereas it was non-significant in mothers and daughters. The pattern of cross-trait familial correlations between height and LDL-C was compatible with the existence of a weak transmissible component explaining the relationship between these two traits. By contrast, the pattern observed for HDL-C and triglycerides was rather compatible with the influence of shared environmental factors. No familial clustering between height and BP levels was detected. CONCLUSIONS: The association between short stature and increased LDL-C might be partly of familial origin.

Myeloperoxidase G-463A polymorphism and Alzheimer's disease in the ApoEurope study.

Alzheimer's disease (AD) is the most common cause of dementia in the elderly. Epidemiological and molecular genetic studies have shown the existence of several genes associated with increased risk of AD, the major genetic susceptibility locus coding for apolipoprotein E (apoE). A polymorphism in the myeloperoxidase gene (MPO) has previously been associated with AD susceptibility. However, results in the literature are controversial and seem to be dependent on several factors such as gender, apoE polymorphism or the genetic structure of the population. We investigated MPO G-463A and apoE polymorphism in 265 cases and 246 controls from the ApoEurope Study. In females, we found a significant association between MPO genotype and AD (P=0.034), GG genotype frequency being lower in cases (52.4%) as compared to controls (64.2%). In men, there was no significant effect of MPO polymorphism. No interaction was found between MPO polymorphism and apoE epsilon 4 allele. In conclusion, the G-463A polymorphism of MPO was statistically associated with AD in a gender-specific manner. However, given the low significance of P value we suggest no causal effect of the MPO gene in AD, as also evidenced in a recent meta-analysis. Our results support the hypothesis of a possible linkage disequilibrium between the MPO G-463A gene polymorphism and another functional variant involved in AD.

Rapid spectrophotometric method for serum glutathione S-transferases activity.

BACKGROUND: Glutathione S-transferases (GST) are a family of polymorphic enzymes responsible for the metabolism of a broad range of xenobiotics including carcinogens and endogenous compounds. METHODS: The aim of this study was to develop a fully automated spectrophotometric method to determine serum GST activity. RESULTS: The method had a detection limit of 7 U/l. The within- and between-run repeatability reached 2% and 3%, respectively. The reproducibility tested over a 6-month period was 7%. Serum GST activity is stable for at least 1 week at +37, +4 and -20 degrees C, 1 month at -20 degrees C and 6 months at -80 and -196 degrees C. The GST activities were determined on 374 supposedly healthy subjects coming to the Center of Preventive Medicine for a health screening. The sample population comprised 178 males and 196 females 4-80 years of age. The mean value of the serum GST activity calculated for this healthy population was 28.2+/-3.0 U/l without gender difference (range from 20.0 to 38.0 U/l). The global interindividual variability reached 11%. CONCLUSIONS: This report describes a spectrophotometric assay for monitoring the serum GST activities in human. This method will be useful in examining the relationships between serum GST activities and the GST polymorphisms, especially pi-GST.

Time-dependent lipid response on fluvastatin therapy of patients with hypercholesterolemia sensitive to apoE phenotype.

Sixty-seven male patients with hypercholesterolemia, divided into three groups according to apolipoprotein E phenotype (33 with apoE3/ 3 phenotype, E3 group; 23 with 2/2 or 2/3, E2+ group; 11 with 4/4 or 4/3, E4+ group), received daily 20-40 mg of fluvastatin for 12 weeks. The levels of triglyceride (TG), cholesterol (Chol), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were measured after 0, 4, 8 and 12 weeks on fluvastatin and after 4 weeks washout period. Lipid percentage changes (delta) were not associated with apoE phenotype for any treatment time. Cholesterol decreased by 14% after 12 weeks and HDL-C increased by 14-16% after 12 weeks for three phenotype groups. deltaTG, deltaChol, deltaLDL-C were associated positively, while negatively for deltaHDL-C, with the corresponding basal lipid values for the three groups. The positive deltaTG values occurred at a low basal TG0 level and became negative at TG0 > 1.6-1.9 mM. For E3 and E4+ groups, only a single parameter contributed significantly into a variation of lipid percentage changes. For the E2+ group, TG0 and Chol0 contributed in a reciprocal manner into deltaTG12/0, both positively into deltaChol8/0; Chol0 and HDL-C0 both negatively contributed into deltaHDL-C12/0. HDL-C0 contributed reciprocally into LDL-C variability for E2+ and E4+ groups. Three effects seem to contribute differently into lipid response among patients with different apoE phenotype: the inhibition of hepatic and lipoprotein lipase activities, the competition between TG-rich and low-density lipoproteins for LDL-receptor and the accumulation of intermediate-density lipoproteins in patients bearing E2 isoform.

Homo- and hetero-complexes of exchangeable apolipoproteins in solution and in lipid-bound form.

The self-association state of human plasma apolipoprotein E (apoE) in solution and in complexes with dimyristoylphosphatidylcholine (DMPC) varying in stoichiometry was studied in sub-micromolar concentration range by gel filtration, fluorescence anisotropy, fluorescence quenching and energy transfer measurements with apolipoprotein labeled with lysine-specific fluorescent dyes. Together, these results confirm the equilibrium scheme for various apoE structures in solution: oligomer (in aged preparations) <==> 'closed' tetramer <==> 'open' tetramer ('molten globule' state) <==> native or partially denatured monomer <==> fully denatured monomer. Within DMPC:apoE discoidal complex (125:1) the apolipoprotein association state seems to be intermediate between that in solution and in larger vesicular complex (1000:1); for both complexes, the degree of exposure of fluorescein chromophores into water phase decreased. Hetero-associates of apoA-I and apoC-III-1 in solution and in the complexes with DMPC appear to behave similarly to apoE. When extrapolated to native HDL particles, 'molten globule' state seems to be a structure responsible for the interaction of exchangeable apolipoproteins with phospholipid. For a first time, the location of various apolipoprotein molecules on disc periphery was confirmed. The lysine residue(s) seems to locate closely to reacting residue(s) within apolipoprotein molecules in associates, however, with different package constraints for discoidal versus vesicular complexes with phospholipid.

Cardiovascular risk-associated allele frequencies for 15 genes in healthy elderly French and Chinese.

In order to investigate possible ethnic differences in genetic and environmental determinants, we investigated several cardiovascular disease-associated genetic variations in successful ageing populations of France (Nancy) and China (Hong Kong). Allelic frequencies of these genetic variations were compared between healthy elderly Chinese (n=103) and French populations (n=100). A multi-locus assay was used to genotype 15 genes for 29 biallelic sites, genes implicated in lipid and homocysteine metabolism, thrombosis, leukocyte adhesion, and blood pressure regulation. For most of the candidate markers within lipid metabolism genes, the less frequent alleles were more common in the Chinese population compared with the French population, while the less frequent alleles of the majority of the other markers were detected only or more commonly in the French population. In conclusion, polymorphisms in 13 genes exhibited statistically significant differences in allelic frequencies between the two populations. Since the two populations were selected as examples of successful ageing, we could hypothesise that genetic factors that could play a role in a successful ageing process may be different between the two populations.

The composition, structural properties and binding of very-low-density and low-density lipoproteins to the LDL receptor in normo- and hypertriglyceridemia: relation to the apolipoprotein E phenotype.

The composition, apolipoprotein structure and lipoprotein binding to the LDL receptor were studied for very-low-density (VLDL) and low-density lipoprotein (LDL) particles isolated from subjects with apoE phenotype E3/3 (E3), E2/2 or E2/3 (E2+) and E3/4 or E4/4 (E4+) and a wide range of plasma triglyceride (TG) contents. The data combined for all three phenotype groups can be summarized as follows. (i) A decrease in accessibility of VLDL tryptophan residues to I- anions with a decrease in tryptophan surface density, concomitant with an increase in VLDL dimensions, reflects the increased efficiency of protein-protein interactions. (ii) A gradual increase in the quenching constant for LDL apoB fluorescence with an increase in TG/cholesterol (Chol) ratio reflects the 'freezing' effect of Chol molecules on apoB dynamics. (iii) Different mechanisms specific for a particular lipoprotein from E3/3 or E2/3 subjects are responsible for apoE-mediated VLDL binding and apoB-mediated LDL binding to the LDL receptor in a solid-phase binding assay. (iv) The 'spacing' effect of apoC-III molecules on apoE-mediated VLDL binding results in a decrease in the number of binding sites. (v) The maximum of the dependence of the LDL binding affinity constant on relative tryptophan density corresponds to LDL intermediate size. VLDL particles from hypertriglyceridemic E2/3 heterozygotic individuals had remnant-like properties (increased cholesterol, apoE and decreased apoC-III content) while their binding efficiency was unchanged. Based on the affinity constant value and LDL-Chol content, increased competition between VLDL and LDL for the binding to the LDL receptor upon increase in plasma TG is suggested, and LDL from hypertriglyceridemic E3/3 homozygotic individuals is the most efficient competitor.