G.G. Neri Serneri

DIFFERENTIAL INHIBITION OF PROSTACYCLIN PRODUCTION AND PLATELET AGGREGATION BY ASPIRIN

Abstract The effect of aspirin ingestion on platelet aggregation, malondialdehyde (MDA) formation, and prostacyclin (PGI 2 ) production by the blood-vessel wall was investigated in twenty-five healthy young volunteers. PGI 2 production induced by ischaemia in the arm was assayed in venous blood. MDA formation and platelet aggregation induced by adenosine disphosphate (ADP), collagen, and adrenaline were inhibited by doses of aspirin smaller than those inhibiting prostacyclin production. The doses which inhibited 50% of platelet aggregation (ID 50) were 3·2 mg/kg for adrenaline, 3·4 for ADP, and 3·2 for collagen, whereas the ID 50 for prostacyclin production was 4·9 mg/kg. Inhibition of platelet aggregation increased only slightly with increasing doses, whereas inhibition of PGI 2 production increased linearly up to 8 mg/kg. Inhibition of platelet aggregation and MDA formation was still present after 72 h, whereas prostacyclin inhibition reversed within 24 h in all subjects after 2, 3·5, and 5 mg/kg and in 6 out of 9 subjects after 8 and 10 mg/kg. These findings indicate that inhibition of platelet cyclo-oxygenase occurs with smaller doses of aspirin and lasts longer than inhibition of vessel-wall cyclo-oxygenase. 3·5 mg/kg is the dose of aspirin most likely to produce a consistent inhibition of platelet aggregation and only a slight inhibition of prostacyclin production.

Effect of heparin, aspirin, or alteplase in reduction of myocardial ischaemia in refractory unstable angina

399 out of 474 inpatients with unstable angina were monitored for 48 h and 97 of these were found to be refractory to conventional antianginal treatments and entered a randomised double-blind study. With the initial protocol heparin infusion or bolus were compared with aspirin; with a modified protocol, heparin infusion, the best of these three treatments, was compared with alteplase. Patients were monitored for 3 days after starting treatment and then observed clinically for 4 more days. On the first days of treatment heparin infusion significantly decreased the frequency of angina (by 84-94%), episodes of silent ischaemia (by 71-77%), and the overall duration of ischaemia (by 81-86%). Heparin bolus and aspirin were not effective. Alteplase caused small (non-significant) reductions on the first day only. Only minor bleeding complications occurred.

TxA2 production by human arteries and veins

Human arterial and venous segments from patients under-going operations when incubated in Tris buffer both alone and with arachidonic acid were able to produce thromboxane B2 (assessed by radioimmunoassay). Thromboxane B2 (TxB2) production was progressive in time (till 40 min.) and was enhanced by the addition of 1mM norepinephrine. Contamination of tissues by platelet was checked and platelets did not contribute to thromboxane formation. The investigation of the conversion of 1-14C arachidonic acid by vascular tissue indicated that human vascular tissues produce the metabolites of the cyclooxygenase dependent pathway and that prostacyclin is the main metabolite with a PGI2/TxA2 ratio of 4:1. The arterial wall was found to possess an active lipoxygenase dependent pathway. Thromboxane production by intimal cells was negligible and the main source of thromboxane was the media. The production of thromboxane did not change in relation to age, but arterial segments from men produced significantly larger amounts of thromboxane than those from women.

Randomised comparison of subcutaneous heparin, intravenous heparin, and aspirin in unstable angina

Intravenous heparin has been used in the control of myocardial ischaemia in patients with unstable angina. We set out to assess the efficacy of subcutaneous heparin in reducing myocardial ischaemia in patients with unstable angina. 343 of 399 patients with unstable angina were monitored for 24 h and 108 were refractory to conventional antianginal treatment and were entered into a randomised multicentre trial. 37 patients were assigned to heparin infusion (partial thromboplastin time 1.5-2 times baseline), 35 to subcutaneous heparin (adjusted dose with partial thromboplastin time 1.5-2 times baseline), and 36 to aspirin (325 mg daily). All had additional conventional antianginal therapy. After the run-in patients were monitored for 3 days. The primary endpoint was reduced myocardial ischaemia assessed by the number of anginal attacks, silent ischaemic episodes, and duration of ischaemia per day. At 1 week and 1 month we accounted for anginal attacks and other clinical events (myocardial infarction, revascularisation procedures, and death). Aspirin did not significantly affect the incidence of myocardial ischaemia. On the first 3 days, infused and subcutaneous heparin significantly decreased the frequency of angina (on average by 91% and 86%, respectively), episodes of silent ischaemia (by 56% and 46%), and the overall duration of ischaemia (66% and 61%) versus run-in day and aspirin (p < 0.001 for all variables). The favourable effects of heparin therapy remained evident during follow-up. Only minor bleeding complications occurred. Subcutaneous heparin is effective in the control of myocardial ischaemia in patients with unstable angina.

Silent ischemia in unstable angina is related to an altered cardiac norepinephrine handling.

BackgroundInferential evidence suggests that silent ischemia might be related to sympathetic activity. Study of [3H]norepinephrine kinetics is a suitable tool to assess the regional sympathetic activity. This method was applied to investigate whether silent myocardial ischemia in unstable angina is related to and depends on cardiac sympathetic overactivity Methods and ResultsPatients with active unstable angina were compared with patients with inactive unstable angina, stable effort angina, and controls. Silent myocardial ischemia was evaluated by three 24-hour Holter monitoring periods on alternate days, and [3H]norepinephrine kinetics was assessed under rest conditions and following the cold pressor test. Simultaneously, catecholamine concentrations were measured in the aortic, coronary sinus, and peripheral venous blood. Different than the other groups (p=0.0013), in patients with active unstable angina, the majority of silent ischemic episodes occurred without increase in heart rate. These patients had a positive coronary sinus-aorta norepinephrine gradient, both at rest and following the cold pressor test. [3H]Norepinephrine kinetics demonstrated an increased selective cardiac spillover, both at rest and, even more, after the cold pressor test. Reduced cardiac [HI norepinephrine extraction also was found. A significant relation was found between the number of ischemic episodes or the overall duration of silent ischemia and norepinephrine spillover, both at rest and following cold application. ConclusionDuring the acute phase of unstable angina (but not in the quiescent phase or in stable effort angina), a disorder in cardiac norepinephrine handling occurs. This results in a reflex cardiac sympathetic overactivity that plays a major role in the occurrence of silent myocardial ischemia.

Enhanced prostacyclin production by dipyridamole in man

SummaryThe effects of some phosphodiesterase (PDE) inhibitors (dipyridamole, theophylline, papaverine and SH-869) on prostacyclin (PGI2) production have been studied in vitro and in vivo. PGI2 was bioassayed by Vanes superfusion technique. In rabbit aortic rings, only dipyridamole in concentrations from 1 to 12 µM was able to stimulate PGI2 biosynthesis in a dose-dependent manner. This effect was also detected with so-called “exhausted” rabbit aortic rings. The other PDE inhibitors used, both in µM and mM concentration, did not affect PGI2 biosynthesis. Dipyridamole was found to increase PGI2 production in healthy volunteers, when given both by infusion (8 µg/kg/min × 2 h) and by oral administration (375 mg/day for seven days). Circulating PGI2 and PGI2 production induced by a 3-min period of ischaemia were increased by an average of 137% (p<0.001) and 30.8% (p<0.001) respectively. Saline and theophylline (as aminophylline) infusions used as controls did not affect PGI2 production.

Cyclooxygenase and lipoxygenase metabolite synthesis by polymorphonuclear neutrophils: in vitro effect of dipyrone

Functional activity of polymorphonuclear neutrophils (PMN) is associated with the metabolism of Arachidonic Acid (AA) released from membrane phospholipids. In this study the in vitro effect of dipyrone, a non steroidal anti-inflammatory drug, on the production of AA metabolites through cyclooxygenase (CO) and lipoxygenase (LO) pathways by stimulated PMN has been investigated. PMN isolated by counterflow centrifuge elutriator were greater than 98% pure and viable. Metabolite production was evaluated by RIA of Thromboxane A2 (TxA2), Prostaglandin E2 (PGE2), Leukotriene B2 (LTB4) and Leukotriene C4 (LTC4) after PMN stimulation with calcium ionophore A 23187 (20 microM). The levels of beta-thromboglobulin (RIA) lower than 5 ng/ml allowed us to rule out activation of residual contaminant platelets. In these experimental conditions, in the absence of dipyrone the products (ng/10(6) cells) of AA metabolism were LTB4 (3.51 +/- 0.22), LTC4 (0.81 +/- 0.08), TxB2 (0.144 +/- 0.025) and PGE2 (0.150 +/- 0.017). Incubation with dipyrone induced changes of PGE2 and TXB2 production in a dose dependent fashion (r = 0.83 and r = 0.87, p less than 0.001), obtaining already at the lowest drug concentration (5 micrograms/ml) a significant inhibition (33 and 40% for TxB2 and PGE2 p less than 0.005). No significant changes of LTB4 and LTC4 production have been observed. The results of this study indicate that dipyrone relevantly affects CO metabolite synthesis by stimulated PMN at concentrations comparable to those reached in therapeutic use. The inhibition of PGE2 synthesis which is present in inflamed tissues and actively participates in inflammatory reactions, could contribute to the therapeutic anti-inflammatory action of dipyrone.

Altered membrane fatty acid composition and increased thromboxane A2 generation in platelets from patients with diabetes.

Lipid composition of platelet membranes and thromboxane A2 (TxA2) generation by platelets were investigated in 42 diabetic patients (14 with macroangiopathic complications, 10 with microangiopathy and 18 without vascular complications) and in 42 clinically healthy subjects of similar age. All subjects were on a similar dietary regimen and the adherence to diet was checked by analysis of red blood cell lipids. Platelets from all groups of diabetic patients produced increased amounts of TxA2 than platelets from controls (at least p less than 0.01) and patients with macroangiopathy (p less than 0.01). Platelet cholesterol and total platelet phospholipids were higher in patients with macroangiopathy, while the relative percentage of the different phospholipid fractions in platelet membrane and their saturated and unsaturated fatty acids were similar in the different groups. Arachidonic acid (AA) content in phosphatidylcholine (PC) was found to be significantly higher in diabetic patients than in controls (at least p less than 0.005). Moreover patients with macroangiopathy had higher AA (p less than 0.001) and lower eicosapentaenoic (EPA) and docosahexaenoic acid (DHA) levels in PC (p less than 0.001) than the other groups of patients and controls.

Enhanced peripheral vasoconstrictor response and increased thromboxane A2 synthesis after the cold pressor test in patients with angina at rest.

Peripheral vascular resistance (PVR) and thromboxane A2(TxA2) synthesis after the cold pressor test were investigated in different subsets of patients with angina (10 with stable effort angina, 36 with resting angina [24 in an active phase and 12 in an inactive phase], and five with Prinzmetals variant angina) and in 41 control subjects of equivalent age and risk factors. Left ventricular end-diastolic pressure, ejection fraction, extent of coronary angiographic lesions, and baseline PVR were not significantly different among the various patient groups. In all patient groups, except those with variant angina, the cold pressor test resulted in a higher increase in PVR than in the control subjects (p less than .001 for all groups). In patients with variant angina the vasoconstrictor response was increased only in proximity (about 1 hr) to ischemic attacks. In patients with active resting angina the vasoconstrictor response was on the average four times longer than that in patients with effort angina and with inactive resting angina (p less than .001). This exaggerated vasoconstrictor response was associated with elevated TxA2 resting levels in plasma and with increased TxA2 synthesis after the cold pressor test. A linear relationship was found between the area of the vascular response and the area of TxA2 production after the cold pressor test in patients with active resting angina (r = .87, p less than .001). The increased TxA2 synthesis and the inappropriate increase of peripheral vascular response to sympathetic stimulation revert back to normal in the inactive phase.(ABSTRACT TRUNCATED AT 250 WORDS)

Repeated sympathetic stimuli elicit the decline and disappearance of prostaglandin modulation and an increase of vascular resistance in humans.

To investigate the role of prostaglandins I2 and E2 in modulating the vasoconstrictor response to sympathetic stimulation, repeated and proximate cold pressor tests were performed in 23 healthy young volunteers. Limb vascular resistance (blood flow measured by venous occlusion plethysmography), prostaglandin I2 (as 6-ketoprostaglandin F1 alpha) and prostaglandin E2 plasma levels (detected by radioimmunoassay), and plasma catecholamines (detected by high-performance liquid chromatography and electrochemical detection) were measured. A progressive increase in the duration of the vasoconstrictor response was observed with repetition of cold applications (p less than 0.001, by analysis of variance for trends). This increase was associated with a progressive decrease in cold-induced elevation of 6-ketoprostaglandin F1 alpha and prostaglandin E2 plasma levels until, after five stimulations, neither prostaglandin was detectable. The maximum detected concentration of norepinephrine did not significantly change, but its area under the curve in time showed a trend toward an increase. Epinephrine levels did not significantly change. The increase of vascular resistance was significantly correlated with the decrease of both prostaglandins (r = 0.93, p less than 0.05 for prostaglandin E2 and r = 0.89, p less than 0.05 for 6-ketoprostaglandin F1 alpha), whereas no significant correlations were found between variations of vascular resistance and catecholamines. Prostaglandin blockade induced by diclofenac sodium administration caused, from the first cold application, a pattern of the vasoconstrictor response and plasma prostaglandin and norepinephrine changes similar to that observed at the fifth cold application in untreated subjects, when prostaglandins are no longer detectable in plasma. We conclude that an increased vasoconstrictor response to sympathetic stimulation in humans may result from a diminished inhibitory influence of prostaglandins on adrenergic transmission.