M.A. Barradas

Effect of ethanol on vascular prostacyclin (prostaglandin I2) synthesis, platelet aggregation, and platelet thromboxane release

A series of experiments with platelets from healthy volunteers showed a concentration related inhibitory effect of ethanol on platelet aggregation and release of thromboxane A2. This effect was observed at blood alcohol concentrations ranging between 66 and 132 mg/dl (14.3 and 28.6 mmol/l), which are commonly found in alcoholics. Investigations carried out by incubating ethanol with platelet rich plasma in vitro also showed an inverse linear correlation between ethanol concentration and platelet thromboxane synthesis. In contrast, the incubation of a wide range of concentrations of ethanol with human endothelial cells and rat aortic rings did not alter the ability of these systems to synthesise prostacyclin (prostaglandin I2). This finding of a selective inhibition of thromboxane A2 synthesis and platelet aggregation without an alteration of prostaglandin I2 synthesis may provide an explanation for the reported ethanol mediated protection against vascular disease. This effect of ethanol may also be relevant to the induction of acute gastrointestinal haemorrhage that occurs after bouts of excessive alcohol consumption.

Intraplatelet serotonin in patients with diabetes mellitus and peripheral vascular disease

Abstract Intraplatelet serotonin (5‐HT) content was determined in 23 patients with type I (insulin‐dependent) diabetes mellitus (IDDM), 23 patients with type II (non‐insulin‐dependent) diabetes mellitus (NIDDM), 29 patients with peripheral vascular disease (PVD) and 34 age‐matched normal subjects. Intraplatelet 5‐HT content in normal subjects showed an age‐related decline (r=–0·45; P < 0·008), as has been previously demonstrated. The median 5‐HT content in platelets of the young normal subjects was 4·36 (range: 3·62–6·79) nmol 10‐9platelets, while that in the elderly normal subjects was 3·87 (range: 2·8–6·0) nmol 10‐9platelets and that in young+elderly subjects was 4·05 (range: 2·8–6·8) nmol 10‐9platelets. The median intraplatelet 5‐HT content was significantly lower (P < 0·002) in IDDM patients: 3·0 (range 1·3–6·3), NIDDM patients: 2·5 (range 1·7–5·8), PVD patients: 2·42 (range 0·94–4·98) nmol 10‐9platelets than that in all young+elderly healthy subjects. The presence of hypertension in DM patients caused a small but significant (P < 0·05) decrease in intraplatelet 5‐HT content, whilst its presence had no effect in PVD patients. In a smaller study, it was established that NIDDM and PVD patients have significantly (P < 0·002) greater plasma 5‐HT concentrations than controls. Insulin‐dependent diabetes mellitus patients had greater plasma 5‐HT concentrations but this did not achieve statistical significance despite a 66% increment in its value. We conclude that the diminished 5‐HT content in platelets and the increased plasma levels may reflect enhanced release of 5‐HT by hyperactive platelets. This increase in plasma 5‐HT may contribute to the pathogenesis of atherosclerosis and vasospasm.

Heparin-induced platelet aggregation in anorexia nervosa and in severe peripheral vascular disease

Abstract. We have previously demonstrated that platelets obtained from patients with anorexia nervosa or severe peripheral vascular disease are hyperaggregable. Since conventional heparins are known to activate platelets in vitro and occasionally induce thrombosis and consumptive thrombocytopenia in vivo, we have investigated the direct effect of a conventional heparin on platelets obtained from patients with anorexia nervosa or severe peripheral vascular disease. Heparin at therapeutic concentrations was found to induce platelet aggregation of such platelets in vitro. In contrast, a recently developed low molecular weight heparinoid (Org 10172), at therapeutic concentrations, had no effect on these hyperaggregable platelets. We conclude that: (i) heparin may be potentially harmful to patients with hyperaggregable platelets; (ii) thrombocytopenia and thrombosis associated with heparin therapy may be mediated through a direct effect of heparin on platelets; (iii) it is unlikely that heparin induced thrombocytopenia is always mediated by classical immunological mechanisms, especially in patients with hyperaggregable platelets; and (iv) low molecular weight heparinoids may be safer anticoagulants in patients with platelet hyperaggregability.

Fibrinogen mediated activation of platelet aggregation and thromboxane A2 release: pathological implications in vascular disease.

The effect of a human fibrinogen preparation on in vitro platelet aggregation was assessed. Platelets were obtained from healthy volunteers. Human fibrinogen induced platelet aggregation in 65% of platelet rich plasma samples and enhanced submaximal platelet aggregation induced by heparin or by several conventional agonists in all samples. Aggregation induced by fibrinogen alone was reversed by the in vitro addition of human albumin. Fibrinogen induced aggregation was associated with the release of the vasoconstrictor, thromboxane A2. Preincubation with indomethacin inhibited both the aggregation and the release of thromboxane A2. Fibrinogen had no effect on in vitro vascular prostaglandin I2 synthesis (rat aortic rings) during a 60 minute incubation. The observed effects of fibrinogen on platelet function may be relevant to clinical conditions in which hyperaggregability of platelets is associated with hyperfibrinogenemia and thrombosis.

Platelet Function in Patients Admitted with a Diagnosis of Myocardial Infarction

Platelet function and thromboxane A2 release were measured in 71 patients admitted to a coronary care unit with a provisional diagnosis of acute myocardial infarction (AMI). All measurements were carried out within twenty-four hours of admission. Of these, 35 patients had the diagnosis of AMI confirmed. The remainder (n=36), who did not have AMI (NMI), were divided into two groups: (a) those (n = 18) with an unequivocal history of previous vascular disease and (b) those without vascular disease (n=18). Platelet aggregation and thromboxane A2 (TXA2) release were significantly increased in the AMI group when compared with those in the NMI without vascular disease group or a healthy control group with similar age and sex distribution. Aggregation and TXA 2 release in the NMI patients with vascular disease were greater than those in controls and did not differ significantly from those in the AMI group. Patients in the AMI or NMI with vascular disease groups who were taking β-blockers or calcium channel antagonists at the time of admission showed significantly less platelet aggregation than those who were not taking these drugs. Heparin, added in vitro at therapeutic concentrations, induced significantly more aggregation in patients in the AMI and NMI with vascular disease groups than in the NMI without vascular disease group. We conclude that: (1) platelets obtained from patients with AMI are hyperaggregable and release more TXA2; (2) platelets from patients with significant vascular disease are hyperaggregable, even in the absence of AMI, although they are not as hyperaggregable as those from AMI; (3) treatment with nifedipine and β-blockers protects these patients from platelet hyperaggregability; (4) heparin induces significant aggregation of platelets from patients with AMI and NMI with vascular disease. These observations are of importance in considering the pathogenesis and treatment of AMI and ischemic heart disease.

Platelet function defects in chronic alcoholism.

Platelet function in alcoholic patients was assessed on admission and during abstinence in hospital. On admission platelets from these patients were significantly less responsive (percentage aggregation and thromboxane A2 release) to conventional in vitro aggregating agents (adrenaline, adenosine diphosphate, and collagen) than platelets from healthy, moderate drinkers. Initially, platelet counts in platelet rich plasma tended to be low and the Simplate II bleeding times frequently prolonged. Platelet aggregation and thromboxane A2 release, however, were inhibited even in patients with normal platelet counts on admission. Platelet aggregation and thromboxane A2 release returned to normal or became hyper-responsive during two to three weeks of abstinence. Platelet counts rose during this period, the largest responses occurring in those patients with the lowest counts on admission. Bleeding times reverted to normal during abstinence and correlated significantly with changes in platelet aggregation, thromboxane A2 release, and platelet count and with the estimated ethanol consumption during the week before admission. Chronic, heavy alcohol ingestion evidently exerts an inhibitory effect on platelet function even in the absence of alcohol in the blood, and this phenomenon is reversible on abstaining. The impaired platelet function, together with the reduced platelet count, may contribute to the bleeding diathesis associated with chronic alcoholism and to the increased incidence and recurrence of gastrointestinal haemorrhage associated with excessive alcohol intake.

Comparison of the platelet pro-aggregatory effect of conventional unfractionated heparins and a low molecular weight heparin fraction (CY 222).

Summary. Two unfractionated heparins (UH), a porcine intestinal mucosal heparin (PIM), a bovine lung heparin (BLH) and a low molecular weight heparin (LMWH), CY 222, were assessed for their capacity to enhance platelet aggregation in vitro. Their potential proaggregatory effect was investigated in four systems: (a) enhancement of submaximal platelet aggregation induced by conventional agonists in platelet rich plasma and (b) in whole blood; (c) reversal of inhibition of platelet aggregation induced by iloprost, a stable analogue of prostacyclin; (d) the direct aggregatory effect of these anticoagulants on hyperactive platelets prepared from patients with severe peripheral vascular disease or anorexia nervosa. Whereas BLH and PIM, at therapeutic concentrations, had a proaggregatory effect in all four systems, CY 222 had no significant effect when compared with the controls. BLH was more potent than PIM in three of the four systems studied. These observations confirm that conventional UH are more proaggregatory than LMWH, and thus the latter may be potentially safer. These observations are also consistent with the fact that BLH administration causes thrombocy‐topenia more frequently than PIM. The direct activation by UH of platelets from patients not previously exposed to heparin also challenges the hypothesis that heparin‐induced platelet activation and thrombocytopenia is solely mediated by classical heparin‐dependent immune mechanisms.

Iron chelators inhibit human platelet aggregation, thromboxane A2 synthesis and lipoxygenase activity

The iron chelators desferrioxamine and 1,2‐dimethyl‐3‐hydroxypyrid‐4‐one (L1) inhibited human platelet aggregation in vitro as well as thromboxane A2 synthesis and conversion of arachidonate to lipoxygenase‐derived products. Non‐chelating compounds related to L1 were without effect on cyclooxygenase or lipoxygenase activity. Since both cyclooxygenase and lipoxygenase are iron‐containing enzymes, it is suggested that the inhibition of platelet function by these iron chelators may be related to the removal or binding of iron associated with these enzymes. These iron chelators may therefore be of potential therapeutic value as platelet antiaggregatory agents and of possible use in the treatment of atherosclerotic and inflammatory joint diseases.

Plasma histamine concentrations are elevated in patients with diabetes mellitus and peripheral vascular diasease

Previous work has shown that plasma and tissue concentrations of histamine are elevated in rats with experimental diabetes mellitus and that leucocytes and platelets from patients with peripheral vascular disease have a higher histamine content than those from controls. In the present study, we have measured: (a) plasma histamine concentrations; (b) leucocyte and platelet histidine decarboxylase (the enzyme responsible for the biosynthesis of histamine) in patients with diabetes mellitus (Types I and II) and peripheral vascular disease; and (c) platelet and leucocyte histamine content. Plasma histamine concentration was significantly higher in patients with diabetes and peripheral vascular disease respectively than that in age-matched controls. Leucocyte histidine decarboxylase activity in diabetic and peripheral vascular disease patients was similar to that in controls, while platelets had no histidine decarboxylase activity. The leucocyte and platelet content of histamine were greater in patients with peripheral vascular disease than those in controls, but they were not altered in diabetic patients. There was no correlation between plasma histamine concentration, leucocyte and platelet histamine content, and histidine decarboxylase activity. We conclude that plasma histamine is elevated in diabetics and in patients with peripheral vascular disease and that platelet and leucocyte histamine content is increased in the latter. This increase in platelet and leucocyte histamine content is not due to an increase in histidine decarboxylase activity of these cells. The increase in plasma and cellular histamine content may contribute to the pathogenesis of increased endothelial permeability in diabetes and to the pathogenesis of intimal damage in atherosclerosis.

The effect of olive oil supplementation on human platelet function, serum cholesterol-related variables and plasma fibrinogen concentrations: a pilot study.

Abstract Seven healthy volunteers ingested 21 g olive oil daily for 8 weeks. Platelet aggregation (in platelet rich plasma and in whole blood) and thromboxane A 2 release (in platelet rich plasma) were reduced after supplementation; some of these values achieved statistical significance. There were no significant changes in platelet count and median platelet volume or in the concentrations of plasma fibrinogen or serum cholesterol, low density lipoprotein and high density lipoprotein after supplementation. Platelet membrane oleic acid content increased significantly ( P P 2 release, since arachidonic acid is the substrate for thromboxane A 2 . It is of interest that populations consuming large amounts of olive oil have a low incidence of ischaemic heart disease. Olive oil should no longer be considered an inactive placebo in studies assessing platelet function.