P. Maderna

Hypertriglyceridemia and regulation of fibrinolytic activity.

A relation between elevated triglyceride (TG) levels and alterations of the fibrinolytic system has been recognized in studies of patients with coronary heart disease. In this investigation, the total fibrinolytic activity and the levels of specific components of the fibrinolytic system were evaluated in plasma samples from a highly selected group of patients with type IV hyperlipoproteinemia before and after a dietary treatment aimed at reducing TG levels. The fibrinolytic response of type IV patients was comparable to that of normolipidemic subjects, whereas tissue-type plasminogen activator antigen levels before and after venous occlusion (p less than 0.01) and resting plasminogen activator inhibitor-1 (PAI-1) antigen (p less than 0.01) and activity (p less than 0.01) were significantly higher in hypertriglyceridemic subjects compared with controls. After dietary treatment, a 22% reduction in TG levels was attained in type IV patients, with no appreciable modification of fibrinolytic parameters. The analysis of the single-patient data revealed a tendency toward normalization of PAI-1 levels only in those patients who showed a TG reduction greater than or equal to 20%. Very low density lipoproteins (VLDLs) from both normal and type IV patients concentration-dependently stimulated PAI-1 release by endothelial cells and HepG2 cells, with the effect of VLDL from type IV patients being more pronounced on HepG2 cells. The release of PAI-1 induced by VLDL in competent cells may thus account for the elevated levels of this antifibrinolytic protein that occur in hypertriglyceridemic patients.

Increased platelet sensitivity and thromboxane B2 formation in type-II hyperlipoproteinaemic patients

Abstract. Platelet aggregation induced by collagen, ADP and epinephrine, was monitored in 150 type‐II patients (115 type IIA and 35 type IIB) and compared with a reference group of normolipidaemic controls; in addition, malondialdehyde formation and thromboxane B2 were examined in a subsample of the type‐IIA patients. Threshold aggregatory concentrations were significantly lower in the whole group of type‐II patients for all three aggregating agents; no difference in terms of aggregatory response was detected between platelets from type‐IIA and ‐IIB patients. Only 56% of type‐II patients, however, exceeded the 95th percentile of the threshold aggregatory concentrations in controls. The formation of malondialdehyde in platelet‐rich plasma stimulated with thrombin and collagen, was significantly higher in platelets from type‐IIA patients. The production of thromboxane B2 by platelets, from endogenous arachidonic acid in type‐IIA patients, was significantly higher and exceeded the highest level found in controls.

Reverse cholesterol transport: Physiology and pharmacology

Reverse cholesterol transport identifies a series of metabolic events resulting in the transport of cholesterol from peripheral tissues to the liver and plays a major role in maintaining cholesterol homeostasis in the body. High density lipoproteins (HDL) are the vehicle of cholesterol in this reverse transport, a function believed to explain the inverse correlation between plasma HDL levels and atherosclerosis. An attempt to stimulate, by the use of drugs, this transport process seems to be of great promise in the prevention and treatment of arterial disease. Only few drugs are now known that can modify the activity of the various factors involved in the process. Clofibrate reduces cholesterol esterification, but the newer fibric acids are generally ineffective as anion-exchange resins. Probucol directly increases the activity and mass of cholesteryl ester transfer protein, thus possibly improving the physiological process of cholesterol removal from tissues. The few available data on the effects of drugs on reverse cholesterol transport should stimulate the search for new agents specifically stimulating this antiatherogenic process.

Increased synthesis of plasminogen activator inhibitor-1 by cultured human endothelial cells exposed to native and modified LDLs : an LDL receptor-independent phenomenon

The effects of native and acetylated low density lipoproteins (LDLs and acetyl-LDLs, respectively) on the release of plasminogen activator inhibitor type 1 (PAI-1) by cultured human umbilical vein endothelial cells (ECs) were evaluated. LDL and acetyl-LDL incubated with ECs for 16-18 hours increased the PAI-1 antigen levels in conditioned medium. At a concentration of 100 micrograms/mL, LDL and acetyl-LDL increased PAI-1 by 10.8 and 12.0 ng/mL, respectively (p < 0.05 and p < 0.01 versus control). The increases in PAI-1 antigen levels exerted by the lipoproteins paralleled the changes in PAI-1 activity. The effect of LDL and acetyl-LDL was concentration dependent and specific for PAI-1 because tissue-type plasminogen activator and expression of procoagulant activity were not affected by either lipoprotein. In addition, total protein synthesis evaluated in [35S] methionine-labeled ECs was not affected, and studies with cycloheximide showed that the effect of LDL and acetyl-LDL on PAI-1 release was due to de novo protein synthesis. Experiments using the C7 monoclonal antibody against the LDL receptor and binding-defective LDL indicated that the effect of LDL on the synthesis of PAI-1 was not dependent on the interaction of the LDLs with their specific receptors. Finally, extensive oxidation of LDL prevented and even reversed the effect of LDL on PAI-1 release by ECs. It is concluded that LDL specifically increases the synthesis of PAI-1 by ECs with mechanisms that are not receptor mediated.

n-3 fatty acid ethyl ester administration to healthy subjects and to hypertriglyceridemic patients reduces tissue factor activity in adherent monocytes.

n-3 Fatty acids are known to influence several functions of monocytes, including adhesion, cytokine synthesis, and superoxide generation. Monocytes express tissue factor, a membrane-bound glycoprotein, that acts as a catalyst in the coagulation cascade. In this study we evaluated the effects of administration of n-3 fatty acid ethyl esters to healthy volunteers and to hypertriglyceridemic patients on tissue factor activity (TF activity) in adherent monocytes. n-3 Fatty acids containing 75% eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (ratio of EPA to DHA, 1.34) were administered (3 g/d) to normal volunteers for 18 weeks. In addition, the effects of this treatment were evaluated in 30 hypertriglyceridemic patients for 24 weeks by using a double-blind, placebo-controlled study. TF activity in adherent monocytes was evaluated with a one-stage clotting assay. Plasma and monocyte fatty acid compositions were determined by gas-liquid chromatography. In healthy volunteers, n-3 fatty acids significantly reduced TF activity in adherent monocytes either in the unstimulated condition or after exposure to endotoxin. The inhibitory effect was observed after 12 weeks of treatment and was more pronounced after 18 weeks (> 70%, P < .001 versus baseline). Concomitantly, levels of EPA and DHA increased in plasma and monocyte lipids. Interestingly, after stopping treatment, monocyte TF activity remained inhibited for at least 14 weeks. Treatment with n-3 fatty acids for 24 weeks also resulted in a significant reduction of TF activity in adherent monocytes from hypertriglyceridemic patients (-31% and -40% in unstimulated and endotoxin-stimulated cells; P < .05 versus baseline).(ABSTRACT TRUNCATED AT 250 WORDS)

Platelet Aggregation and Malondialdehyde Formation in Type IIA Hypercholesterolemic Patients

Platelet aggregation induced by threshold concentrations of ADP, adrenaline, collagen and Thrombofax was evaluated in 25 type IIA hypercholesterolemic patients in comparison with 15 control subjects. Malondialdehyde (MDA) formation induced by collagen and thrombin was also studied in a subgroup of 8 patients. In the patient group, irreversible platelet aggregation was induced by significantly lower concentrations of both adrenaline (p is less than 0.05) and Thrombofax (p is less than 0.01). MDA formation induced by both aggregating agents was markedly increased (p is less than 0.01) in type IIA hypercholesterolemic patients.

Bezafibrate lowers plasma lipids, fibrinogen and platelet aggregability in hypertriglyceridaemia

SummaryThe effects of bezafibrate 400 mg/day (slow release formulation) on plasma lipids/lipoproteins and on selected haemostatic parameters were evaluated in a double blind cross-over study in patients with Type IIb and IV hyperlipoproteinaemia.Placebo treatment did not influence any of those parameters, but the drug significantly reduced plasma triglycerides (−45%) and VLDL cholesterol, as well as causing a 12 % fall in total cholesterol and a 20 % increase in HDL cholesterol. Apo AI levels were significantly increased following bezafibrate and Apo B was reduced by about 20 %. In addition to changes in the plasma lipid profile, bezafibrate reduced the sensitivity of platelets to the aggregatory effect of collagen, with no effect on TXB2 production. Fibrinogen levels after bezafibrate treatment were significantly lowered, the effect being more marked in patients with hyperfibrinogenaemia. Bezafibrate did not influence the plasma fibrinolytic profile.It is concluded that bezafibrate, besides its beneficial effects on the plasma lipoprotein profile, can exert beneficial changes on specific haemostatic parameters.

Effects of PGI2 on platelet aggregation and adenylate cyclase activity in human type IIa hypercholesterolemia

The sensitivity to PGI2 of platelets of 20 selected type IIa hypercholesterolemic patients was studied and compared to that of platelets of 14 normocholesterolemic subjects. Type IIa subjects required higher concentrations of PGI2 to inhibit platelet aggregation elicited by 1 microM ADP, 1 microgram/ml collagen and 1.4 microM epinephrine. Adenylate cyclase activity was also measured in washed platelet membranes from the two groups of subjects. Adenylate cyclase activity, both basal and PGI2-stimulated, was not statistically different in the two groups examined. Therefore changes at the level of PGI2 receptors coupled to adenylate cyclase are not likely to be responsible for the different platelet sensitivity to prostacyclin.

Prostacyclin-lipoprotein interactions: studies on human platelet aggregation and adenylate cyclase

The in vitro effects of different lipoprotein fractions (VLDL, LDL and HDL) on human washed platelet aggregation, induced by collagen and thrombin, were evaluated in the presence and absence of PGI2. Although VLDL and LDL increased the platelet aggregation while HDL showed an opposite effect, none of the tested lipoprotein fractions affected the potency of PGI2 as inhibitor of platelet aggregation (IC50). In addition, studies were performed to evaluate the effects of lipoproteins on adenylate cyclase activity in human platelet membranes. The three lipoprotein classes inhibited both basal and PGI2-stimulated adenylate cyclase without affecting the EC50 for PGI2. This inhibitory activity was not specifically elicited by any protein or lipid since neither bovine serum albumin nor a lipid emulsion (Intralipid) displayed any inhibition. The effect on adenylate cyclase elicited by VLDL, LDL and HDL does not seem to be correlated with the activity on platelet aggregation. It is concluded that mediators other than cAMP might be involved in the control of platelet function by lipoproteins.

Metformin reduces platelet hypersensitivity in hypercholesterolemic rabbits

The effects of Metformin treatment on platelet responsiveness to aggregating agents was studied in cholesterol-fed rabbits. Three groups of animals were fed, for one month, either a normal (N), or a hypercholesterolemic (HC), or a hypercholesterolemic + 0.5% Metformin diet (HC + Met), Platelets from the HC rabbits required significantly lower collagen and arachidonic acid concentrations to aggregate, as compared to platelets from N rabbits. The platelet response from the HC + Met rabbits was not significantly different from that of normals. The cholesterol/phospholipid ratio in platelets was increased in both dietary groups (HC, HC + Met). The serum thromboxane B2 concentrations did not show any significant difference between the groups. Plasma exchange experiments failed to indicate a specific effect of the plasma environment on platelet behaviour. In view of the inactivity of metformin on the platelet cyclo-oxygenase pathway, the reported results suggest that metformin may act by an as yet unexplored mechanism.