Effect of an alpha-blocker (Nicergoline) and of a beta-blocker (Acebutolol) on the in vitro biosynthesis of vascular extracellular matrix.

The effect of an alpha-blocking agent and of a beta-blocking agent on the biosynthesis of extracellular matrix macromolecules of the arterial wall was investigated. Rabbit aorta explants were cultured up to 48 hours with radioactive proline, lysine or glucosamine. In presence of these drugs, at concentration shown to be effective for the inhibition of platelet-endothelial cell interactions (10(-7) M), the incorporation of 14C proline in total macromolecular proline was higher than in macromolecular hydroxyproline suggesting a relatively higher rate of biosynthesis of non-collagenous proteins as compared to collagens. The alpha-blocking increased the incorporation of 14C proline in collagenous and non-collagenous proteins after 18 hours of incubation. beta-blocking also increased the incorporation of proline in macromolecular proline and hydroxyproline as compared to control cultures. Both increased the incorporation of 3H glucosamine in newly synthesised glycosaminoglycans. beta-blocking increased mainly the neosynthesis of heparan sulphate, alpha-blocking that of hyaluronan. The incorporation of 14C-lysine in crosslinked, insoluble elastin was not modified. These experiments confirm that alpha and beta-blocking agents can influence not only the tonus of aortic smooth muscle cells but also the relative rates of biosynthesis of extracellular matrix macromolecules. This effect should be taken in consideration for the evaluation of the long range effect of alpha and beta-blocking drugs on the vascular wall.

[Role of diet in cardiovascular prevention]. / Place de la diététique dans la prévention cardiovasculaire.

Several dietary components increase the risk of ischemic cardiovascular disease: excess dietary cholesterol and fats (especially saturated fats with low mono- and polyunsaturated fatty acid content), deficiency of antioxidants, fibers. These dietary factors modify the levels and the physicochemical structure of plasma lipoproteins and consequently their metabolic activities. Some of them also modify platelet aggregation and endothelial function. To reduce the cardiovascular risk, different nutritional schemes have been proposed and applied in prevention studies. Mediterranean-type diet, or traditional Asian food provide a very favorable cost/effectiveness ratio. These diets must be associated with the usual measures for improving lifestyle (increased physical activity, reduction of smoking) and with drug treatment of major risk factors (dyslipidemia, hypertension, diabetes).

Autosomal dominant type IIa hypercholesterolemia: evaluation of the respective contributions of LDLR and APOB gene defects as well as a third major group of defects.

Autosomal dominant type IIa hypercholesterolaemia (ADH) is characterised by an elevation of total plasma cholesterol associated with increased LDL particles. Numerous different molecular defects have been identified in the LDL receptor (LDLR) and few specific mutations in the apolipoprotein B (APOB) gene resulting in familial hypercholesterolaemia and familial defective apoB-100 respectively. To estimate the respective contribution of LDLR, APOB and other gene defects in this disease, we studied 33 well characterised French families diagnosed over at least three generations with ADH through the candidate gene approach. An estimation of the proportions performed with the HOMOG3R program showed that an LDLR gene defect was involved in approximately 50% of the families (P = 0.001). On the other hand, the estimated contribution of an APOB gene defect was only 15%. This low estimation of ADH due to an APOB gene defect is further strengthened by the existence of only two probands carrying the APOB (R3500Q) mutation in the sample. More importantly and surprisingly, 35% of the families in the sample were estimated to be linked to neither LDLR nor APOB genes. These data were confirmed by the exclusion of both genes through direct haplotyping in three families. Our results demonstrate that the relative contributions of LDLR and APOB gene defects to the disease are very different. Furthermore, our results also show that genetic heterogeneity is, generally, underestimated in ADH, and that at least three major groups of defects are involved. At this point, the contribution of the recently mapped FH3 gene to ADH cannot be assessed nor its importance in the group of 'non LDLR/non APOB' families.

Plasma lipoprotein distribution and lipid transfer activities in patients with type IIb hyperlipidemia treated with simvastatin.

The aim of the present study was to search in type IIb hyperlipidemic patients for putative concomitant effects of simvastatin on the physicochemical characteristics of low density lipoproteins (LDL) and high density lipoproteins (HDL), as well as on the activities of the cholesteryl ester transfer protein (CETP) and the phospholipid transfer protein (PLTP) that were determined in both endogenous lipoprotein-dependent and endogenous lipoprotein-independent assays. In a double-blind, randomized trial, patients received either placebo (one tablet/day; n = 12) or simvastatin (20 mg/day; n = 12) for a period of 8 weeks after a 5-week run-in period. Simvastatin, unlike placebo, reduced the lipid and apolipoprotein B contents of the most abundant LDL-1, LDL-2, and LDL-3 subfractions without inducing significant changes in the overall size distribution of LDL and HDL. Whereas simvastatin significantly increased PLTP activity in an endogenous lipoprotein-dependent assay (P < 0.01), no variation was observed in a lipoprotein-independent assay. Simvastatin significantly decreased plasma CETP activity in an endogenous lipoprotein-dependent assay (P < 0.01), and the reduction in plasma cholesteryl ester transfer rates was explained by a 16% drop in CETP mass concentration (P < 0.01). In contrast, the specific activity of CETP was unaffected by the simvastatin treatment reflecting at least in part the lack of significant alteration in plasma triglyceride-rich lipoprotein acceptors. The simvastatin-induced changes in plasma CETP mass levels correlated positively with changes in plasma CETP activity (r = 0.483, P = 0.0561), in total cholesterol levels (r = 0.769; P < 0.01), and in LDL-cholesterol levels (r = 0.736; P < 0.01). Whereas the observations suggest that simvastatin might exert concomitant beneficial effects on plasma CETP and LDL levels, neither plasma cholesteryl ester transfer activity nor plasma phospholipid transfer activity appeared as the main determinants of the LDL and HDL distribution profiles in type IIb hyperlipidemic patients.

Abnormal capacity to induce cholesterol efflux and a new LpA-I pre-beta particle in type 2 diabetic patients.

In this study, we first characterized the lipoprotein components of serum samples obtained from a group of well-controlled diabetic patients and from healthy subjects in fasting and postprandial states. We then explored some aspects of reverse cholesterol transport in the same population. Patients showed high levels of fasting triglycerides, postprandial triglyceride responses and LpC-III levels (3.18+/-0.86 vs 2.17+/-0.54 mg/dl, P < 0.001). There were also positive correlations between LpC-III and fasting triglycerides (r = 0.82, P < 0.001), total triglyceride area (r = 0.75, P < 0.001) and incremental triglyceride area (r = 0.54, P < 0.001). HDL-C and apo A-I were significantly decreased in diabetic patients due to a selective reduction in LpA-I subfraction, whose antiatherogenic role is generally accepted (37.4+/-8.0 vs 49.2+/-12.5 mg/dl, P < 0.001). In addition, HDL from patients proved to be triglyceride enriched and cholesteryl ester depleted, alterations which were further amplified in the postprandial state. The molar ratio HDL-C/apo A-I + apo A-II, already defined as a predictor of apo A-I fractional catabolic rate, was significantly diminished in the patient group (15.1+/-2.2 vs 20.8+/-3.3, P < 0.001), thus suggesting an accelerated catabolism of apo A-I. For the first time, we describe here the presence of a small apo A-I-containing particle, isolated by two-dimensional electrophoresis and characterized by immunoblotting, only in samples from diabetic patients. This particle that we named pre-beta0, has an apparent molecular weight of 40 kDa. As regards the capacity of serum samples to promote cholesterol efflux from [3H]cholesterol-labeled Fu5AH rat hepatoma cells, patient samples were found to induce significantly lower cholesterol efflux than controls only in the postprandial state (21.2+/-3.3 vs 23.8+/-1.8%, P = 0.012). The presence of pre-beta0 in samples from diabetic patients might therefore be associated to an altered capacity of these serum samples to promote cellular cholesterol efflux. Overall, these abnormalities may contribute to a delay in the reverse cholesterol transport pathway in type 2 diabetic patients.

Elastin-elastase-atherosclerosis revisited.

This review proposes reinvestigation of a topic studied in the author's laboratory over the last decades concerning the age-dependent modifications of the vascular extracellular matrix (ECM) as related to atherogenesis and its recognized risk-factors: blood lipids, lipoproteins. Most salient previous results are confronted with recent publications in this field. Age-dependent modifications of the vascular wall discussed in this review include upregulation of elastolytic enzymes, demonstrated for the first time in the vascular wall in this laboratory, matrix biosynthesis and receptor function. The progressive deposition of lipids in elastic tissues as well as the addition of lipoproteins or lipids to cell and organ cultures were shown to modify matrix biosynthesis and upregulate elastase expression. Lipid-elastin interactions exhibit a great deal of specificity as shown by the nature and amount of lipids accumulating in elastin in vivo and in vitro. Recent epidemiological studies (the EVA study) enables the confrontation of blood lipid parameters with matrix related components (serum elastase and inhibitors, elastin peptides, fibronectin) in the same blood samples. The elastin laminin receptor present on vascular cells was shown to trigger NO dependent vasodilation, and downregulation of cholesterol synthesis. Both of these functions decrease or disappear with age except the upregulation of elastase release which is preserved and increased. Recent experiments extended these findings to T-lymphocytes present also in the atherosclerotic plaque. Finally several recent publications are analyzed which give more precision on the cellular mechanisms underlying the above-described modifications.

Comparison between extra virgin olive oil and oleic acid rich sunflower oil: effects on postprandial lipemia and LDL susceptibility to oxidation.

The aim of our study was to determine whether the minor polar components of virgin olive oil could have favorable effects (1) on fasting and postprandial lipid profile and (2) on low-density lipoprotein (LDL) composition and susceptibility to oxidation in vitro. Ten normolipidic subjects were included in a crossover study (two diet periods of 3 weeks) and received either virgin olive oil (OO diet) or oleic acid rich sunflower oil. An oral fat load was performed at the end of each period. The plasma lipid levels were not significantly different after both diets in the fasting and postprandial states. A few minor variations of the LDL composition were observed only in the postprandial lipemia, and they were different after both diets. The LDL oxidation susceptibility was evaluated by the formation of conjugated dienes. With LDL isolated in the fasting state, the diene production decreased (p = 0.0573) only after the OO diet. The dienes determined at time 0 and the maximal dienes obtained during the oxidation reaction decreased (p = 0.0145 and p = 0.0184, respectively) only after the OO fat load. Nevertheless, the diene production decrease was not significant (p = 0.0848). Our results suggest a mild effect of minor components of virgin olive oil related to a decrease of LDL susceptibility to oxidation; further analyses are necessary to give clear conclusions about their role.

Mutations at the lipoprotein lipase gene locus in subjects with diabetes mellitus, obesity and lipaemia.

1. The common association of obesity, diabetes mellitus and hyperlipidaemia may have a primary aetiological basis. Insulin resistance has been postulated as a possible cause, but defects in the plasma transport of triacylglycerol or fatty acids could also be primary determinants. 2. We have therefore studied 18 patients with diabetes mellitus, obesity and severe hypertriglyceridaemia for defects of a key protein involved in the clearance of plasma triacylglycerols, lipoprotein lipase. 3. DNA was prepared from leucocytes of 18 patients with the above syndrome, and exons encoding lipoprotein lipase were amplified by PCR. The products were sequenced using the dideoxy chain-termination method. 4. Eight of the subjects were found to possess genetic variants at the lipoprotein lipase gene locus. These were: (a) G579-->A, V108V; (b) G818-->A, G188E; (c) C829-->T, R192; (d) A1127-->G, N291S; (e) C1308-->G, F351L; (f) C1338-->A, T361T; and (g) C1595-->G, S447. Three of these, (c), (e) and (f), have not hitherto been described. Variant (f), appears to be a population polymorphism whose allele frequency in normolipidaemic diabetics was found to be 0.12 (162 chromosomes studied). The others are all rare at frequencies of < 0.01 and may contribute to the phenotype by impairing clearance of plasma triacylglycerols. 5. We conclude that genetic variants at the lipoprotein lipase locus occur commonly in subjects with this syndrome (four out of 18 subjects with probably functional mutants) and may affect the individual's response to obesity and diabetes mellitus for the development of lipaemia.

Role of lipoprotein-bound NEFAs in enhancing the specific activity of plasma CETP in the nephrotic syndrome.

Plasma cholesteryl ester transfer protein (CETP) activity, evaluated by the transfer of radiolabeled cholesteryl esters from a tracer dose of tritiated HDL to the plasma apolipoprotein B-containing lipoproteins, was significantly higher in patients with untreated idiopathic nephrotic syndrome (n = 15) than in normolipidemic control subjects (n = 22) (81.5 +/- 8.4 versus 43.1 +/- 3.1 micrograms CE.mL-1.h-1, respectively; P < .001). The increased CETP activity in nephrotic plasma was explained by a significant rise in both the CETP mass concentration (3.2 +/- 0.2 versus 2.1 +/- 0.1 mg/L; P < .001), and the specific CETP activity, calculated as the ratio of CETP activity to CETP mass (25.3 +/- 1.7 versus 20.4 +/- 1.6 micrograms CE.mg-1.h-1; P < .05). Elevated CETP activity in nephrotic patients was shown to be associated with a significant decrease in the mean size of LDL (24.4 +/- 0.5 versus 26.3 +/- 0.5 nm; P < .0001) as well as in the relative abundance of HDL2a (29.6 +/- 1.6% versus 34.8 +/- 1.1%; P < .05). The nephrotic syndrome was characterized by a significant increase in the relative proportion of lipoprotein-bound nonesterified fatty acids (NEFAs) (35.4 +/- 7.7% versus 7.6 +/- 3.0% of total; P < .01), leading to a significant increase in the electronegative charge of LDL (-4.3 +/- 0.1 versus -3.9 +/- 0.1 mV; P < .05) and HDL (-11.5 +/- 0.1 versus -11.1 +/- 0.2 mV; P < .05). Compared with native, non-supplemented plasma, removal of lipoprotein-bound NEFAs by addition of fatty acid-poor albumin to total plasma from nephrotic patients or control subjects significantly decreased CETP activity and specific CETP activity. Specific CETP activity no longer differed between nephrotic and control groups after albumin supplementation (19.7 +/- 1.5 versus 17.7 +/- 1.5 micrograms CE.mg-1.h-1; NS). It is concluded that, in addition to elevated CETP mass concentration, lipoprotein-bound NEFAs, by increasing the negative electrostatic charge of nephrotic lipoproteins, can facilitate the CETP-mediated neutral-lipid transfer reaction in total plasma from nephrotic patients.

Lipoprotein kinetics in patients with analbuminemia. Evidence for the role of serum albumin in controlling lipoprotein metabolism.

In vitro data suggested that albumin is a key factor controlling apolipoprotein (apo) synthesis by hepatocytes. Studies in analbuminemic rats have shown an increase in secretion of apoB-containing lipoprotein from the liver. We studied the kinetic aspects of apoB- and apoAI-containing lipoprotein metabolism in two sisters with analbuminemia using a constant 14-hour infusion of leucine labeled with stable isotopes. Compared with control subjects, total cholesterol was higher in the two patients (432 and 461 versus 155 +/- 14 mg/dL), as was apoB (257 and 230 versus 72 +/- 7 mg/dL). Triglycerides were slightly increased (134 and 105 versus 89 +/- 9 mg/dL), whereas apoAI was lower (109 and 105 versus 124 +/- 6 mg/dL). VLDL-apoB production was higher, as was the production of IDL-apoB and LDL-apoB (32.8 and 36.0 versus 24.8 +/- 5.9, 32.1 and 27.2 versus 16.4 +/- 2.3, and 14.1 and 17.6 versus 10.3 +/- 1.2 mg.kg-1.d-1, respectively). The fractional catabolic rate of all the apoB-containing lipoproteins was decreased (0.23 and 0.37 versus 0.48 +/- 0.05, 0.27 and 0.28 versus 0.62 +/- 0.08, and 0.012 and 0.009 versus 0.022 +/- 0.002.h-1, respectively). A similar mechanism could explain the dyslipidemia observed in other conditions associated with low albumin levels, such as nephrotic syndrome.

Oleic acid rich diet protects against the oxidative modification of high density lipoprotein.

Oxidative modifications of lipoproteins could contribute to the development of atherosclerosis, but the influence of dietary fats on high density lipoprotein (HDL) oxidative modification is unknown. This study was designed to determine whether a diet rich in oleic acid could modulate the oxidative modification of HDL3. Twenty two healthy men were randomly placed on a 32-wk crossover study of an oleic acid rich diet supplied by a variant of sunflower oil vs a linoleic acid rich diet provided by conventional sunflower oil. Plasma HDL3 obtained after the diet rich in oleic acid showed a significantly higher oleic acid content in the phospholipid than lipoprotein isolated after the linoleic acid rich diet. HDL3 isolated after the oleic acid rich diet had lower values of thiobarbituric acid reactive substances (TBARS) than HDL3 obtained after the linoleic acid rich diet both for native (mean +/- SE; 0.24 +/- 0.02 vs 0.42 +/- 0.08 nmol MDA/mg protein; p < 0.01) and copper oxidized HDL3 (0.75 +/- 0.06 vs 0.95 +/- 0.07 nmol MDA/mg protein; p < 0.01). Indeed, TBARS for native HDL3 were negatively correlated with the oleic acid to linoleic acid ratio and positively with the percentage of linoleic acid in their phospholipids. Interestingly, HDL3 after both diets had similar antioxidant vitamins A and E content. HDL3 overall composition and fluidity were similar after the two diets. Moreover, HDL3 obtained after both diets produced identical [3H] free cholesterol efflux from human monocyte-derived macrophages (29%) and fibroblasts (26%). In conclusion, HDL3 rich in oleic acid was less easily oxidized regardless of the content of antioxidants such as vitamins A and E. Therefore, dietary monounsaturated fatty acid prevent the oxidative modification of lipoproteins.

Lipid-lowering drugs and mitochondrial function: effects of HMG-CoA reductase inhibitors on serum ubiquinone and blood lactate/pyruvate ratio.

1. Statins inhibit synthesis of mevalonate, a precursor of ubiquinone that is a central compound of the mitochondrial respiratory chain. The main adverse effect of statins is a toxic myopathy possibly related to mitochondrial dysfunction. 2. This study was designed to evaluate the effect of lipid-lowering drugs on ubiquinone (coenzyme Q10) serum level and on mitochondrial function assessed by blood lactate/pyruvate ratio. 3. Eighty hypercholesterolaemic patients (40 treated by statins, 20 treated by fibrates, and 20 untreated patients, all 80 having total cholesterol levels > 6.0 mmol l-1) and 20 healthy controls were included. Ubiquinone serum level and blood lactate/pyruvate ratio used as a test for mitochondrial dysfunction were evaluated in all subjects. 4. Lactate/pyruvate ratios were significantly higher in patients treated by statins than in untreated hypercholesterolaemic patients or in healthy controls (P < 0.05 and P < 0.001). The difference was not significant between fibratetreated patients and untreated patients. 5. Ubiquinone serum levels were lower in statin-treated patients (0.75 mg l-1 +/- 0.04) than in untreated hypercholesterolaemic patients (0.95 mg l-1 +/- 0.09; P < 0.05). 6. We conclude that statin therapy can be associated with high blood lactate/ pyruvate ratio suggestive of mitochondrial dysfunction. It is uncertain to what extent low serum levels of ubiquinone could explain the mitochondrial dysfunction.

Prevention of recurrent acute pancreatitis in patients with severe hypertriglyceridemia: value of regular plasmapheresis.

The association between acute pancreatitis and severe hypertriglyceridemia has long been recognized. We report two cases of severe primary hypertriglyceridemia (types 1 and V) with recurrent acute pancreatitis. In both patients, observance of appropriate diet and drug therapy was insufficient. Recurrent episodes of pancreatitis were precipitated by dietary fat or alcohol abuse. A plasmapheresis was performed every 4 weeks to decrease the incidence of pancreatitis. It appears that plasmapheresis is a safe and highly effective method for quickly removing serum triglycerides. Moreover, plasma-pheresis may be useful for preventing acute pancreatitis.

Increased cholesteryl ester transfer activity in plasma from analbuminemic patients.

Hypercholesterolemia associated with analbuminemia, an inherited disease manifesting low plasma albumin concentration, is characterized by enhanced LDL cholesterol levels and reduced HDL cholesterol levels. In addition, compared with normal counterparts, the esterified cholesterol:triglyceride ratio tends to be higher in analbuminemic apoB-containing lipoproteins and lower in analbuminemic HDL. The aim of the present study was to investigate the mechanism that may account for the association of a hypoalbuminemic state with alterations in the concentration and composition of plasma lipoprotein fractions. To this end, endogenous cholesterol esterification activity, phospholipid transfer activity, and cholesteryl ester transfer activity were measured in total plasma from three analbuminemic patients and five control subjects. Whereas endogenous cholesterol esterification and phospholipid transfer rates were not significantly affected in analbuminemia, the transfer of radiolabeled cholesteryl esters from HDL toward apoB-containing lipoproteins was constantly higher in analbuminemic plasmas than in normal control plasma (473.6+/-107.3% x h(-1) x mL(-1) versus 227.5+/-84.0% x h(-1) x mL(-1), respectively; P=.036). The rise in cholesteryl ester transfer protein (CETP) activity in analbuminemic plasma was due to a significant increase in the transfer of radiolabeled cholesteryl esters toward LDL but not toward the triglyceride-rich lipoproteins. The CETP mass was higher in analbuminemic patients than in control subjects, but the difference did not reach the significance level (5.18+/-0.82 mg/L versus 3.13+/-1.19 mg/L respectively; P=.07). Since abnormally elevated nonesterified fatty acid (NEFA) levels were shown to be associated with analbuminemic lipoproteins, mostly LDL, the direct role of lipoprotein-bound NEFA in enhancing CETP activity was suspected. In support of this view, supplementation of total plasmas with fatty acid-poor albumin was shown to reduce CETP activity to a significantly greater extent in analbuminemic plasmas than in normal control plasma. It is concluded that hyperlipidemia associated with the hypoalbuminemic state can relate, at least in part, to the combined effect of CETP and NEFA in promoting the transfer of cholesteryl esters from the antiatherogenic HDL toward the proatherogenic apoB-containing lipoproteins.

[Homocysteine: relations to ischemic cardiovascular diseases]. / L'homocystéine: ses liens avec les maladies cardiovasculaires ischémiques.

Homocysteine, a sulfur-containing amino acid, is an intermediate metabolite of methionine. Patients with homocystinuria and severe hyperhomocysteinemia develop premature arteriosclerosis and arterial thrombotic events, and venous thromboembolism. Studies suggest that moderate hyperhomocysteinemia can be considered as an independent risk factor in the development of premature cardiovascular disease. In vitro, homocysteine has toxic effects on endothelial cells. Homocysteine can promote lipid peroxidation and damage vascular endothelial cells. Moreover, homocysteine interferes with the natural anticoagulant system and the fibrinolytic system. Homocysteinemia should be known in patients with premature vascular diseases, especially in subjects with no risk factors. Folic acid, vitamin B6 can lower homocysteine levels.