Inhibition of platelet aggregation by alpha1-acid glycoprotein

In view of known abnormalities of plasma proteins in diseases showing changes in platelet aggregation, the effect of one of the major serum proteins, alpha1-acid glycoprotein, on platelet aggregation was evaluated. This protein, when added to platelet-rich plasma, markedly inhibited platelet aggregation induced by both adenosine diphosphate (ADP) and epinephrine. Transferrin, similarly studied, had no effect. These results are consistent with the hypothesis that the relative concentration of alpha1-acid glycoprotein may influence platelet aggregation in diseases associated with abnormal concentrations of this protein.     

Inhibition of platelet aggregation by acetylsalicylic acid and other inhibitors

Effects on platelet aggregation were examined of acetylsalicylic acid (ASA), indomethacin and a number of other agents including dipyridamole, phenylbutazone and sulfinpyrazone under standardized conditions. The Born turbidometric method of measuring platelet aggregation was used with collagen as the stimulus for aggregation. ASA and indomethacin were shown to be among the most potent inhibitors of aggregation, being active at minimal effective concentrations of 1-3 mug/ml using a 10 min time of pre-incubation with the platelet-rich plasma (degree of aggregation inhibition was time dependent). Most of the other agents tested were also active in vitro and both prostaglandin E1 and adenosine were more potent than ASA or indomethacin. However, these agents were shown not to exert significant inhibitory effects when administered orally to rats (dose 10 and 30 mg/kg). ASA proved to be effective in doses as low as 3 mg/kg, and indomethacin in doses as low as 1 mg/kg orally. The inhibitory effects of ASA on aggregation remained for several days after a single oral dose, whereas the effects of indomethacin disappeared within 24 h.     

Inhibition of long-chain fatty acid metabolism does not affect platelet aggregation responses

A number of anti-anginal agents (perhexiline, amiodarone, trimetazidine) have been shown to inhibit myocardial carnitine palmitoyltransferase-1, which controls access of long-chain fatty acids to mitochondrial sites of beta-oxidation. In view of clinical data suggesting that perhexiline improves symptomatic status in unstable angina pectoris, and the known role of mitochondrial beta-oxidation in platelet metabolism, we compared the platelet carnitine palmitoyltransferase-1 inhibitory and putative anti-aggregatory effects of perhexiline, amiodarone and trimetazidine with those of specific carnitine palmitoyltransferase-1 inhibitors: etomoxir and hydroxyphenylglyoxylate in both normal subjects and patients with stable angina. All of the compounds examined inhibited platelet carnitine palmitoyltransferase-1 activity; rank order of potency etomoxir > malonyl-CoA > hydroxyphenylglyoxylate > amiodarone > or = perhexiline > trimetazidine. However, only perhexiline, amiodarone and trimetazidine inhibited platelet aggregation. We conclude that (a) the carnitine palmitoyltransferase-1 inhibitors perhexiline, amiodarone and trimetazidine exert significant anti-aggregatory effects which may be therapeutically relevant and, (b) these effects are independent of carnitine palmitoyltransferase-1 inhibition.     

Limits of platelet aggregation tests for investigating thromboses (author's transl)

Three methods--spontaneous aggregation, ADP-induced aggregation and levels of circulating platelet aggregates--were used to test for platelet hyperaggregation in 87 adult patients divided into three groups. Group A comprised 26 patients with severe arterial diseases, including cerebral vascular accidents (16) and peripheral thrombosis (10) ; group B consisted of 31 patients with venous conditions, including deep phlebitis (22) and recurrent thrombo-embolic disease (9) ; group C, which served as control, comprised 30 patients with various non-vascular disorders. Spontaneous aggregation and enhanced reactivity to ADP correlated well with each other and were more frequent in patients with vascular diseases. However, one-third of patients in group C had pathological results, while one-third of patients in groups A and B had normal results, independently of any clinical particularity or therapeutic regimen. There were no significant differences between the three groups with regard to circulating platelet aggregates. These tests, therefore, appear to be of statistical and epidemiological interest, but of questionable practical value.     

Shear-induced platelet aggregation (SIPA) monitoring of hemostatic effect during platelet transfusion in a patient with Glanzmann's thrombasthenia: a comparison between SIPA and conventional platelet aggregation

Shear-induced platelet aggregation (SIPA) in a patient with Glanzmann's thrombasthenia was examined during platelet infusion therapy. Prior to platelet infusion, SIPA measured with the modified cone-and-plate type viscometer, as well as ADP-and collagen-induced platelet aggregation measured with the conventional platelet aggregometer, were absent. The patient's SIPA after multiple infusions of platelet, in parallel with the bleeding time and the clinical hemostatic effect, improved to the normal level, while ADP-and collagen-induced platelet aggregation remained abnormal. These observations imply that SIPA is physiologically more relevant than the conventional agonist-induced platelet aggregometry in this type of the disease.     

Evaluation of factors influencing platelet aggregation in patients with chronic glomerulonephritis (CGN)

Platelets (PLT) play an important role in hemostasis, modulation of immunological and inflammatory processes. There is also evidence that PLT takes part in the development of atherosclerosis and glomerulosclerosis. The aim of presented study was to determine morphological and functional changes of platelets and their relation to the lipid, protein and coagulation factors disturbances in patients with chronic glomerulonephritis (CGN). The studies were carried out in 60 patients with CGN diagnosed by renal biopsy: 30 patients without nephrotic syndrome (NS)-CGN and 30 patients with NS-CGN+NS. Protein and lipid disturbances, coagulation factors were estimated using routine laboratory methods. Platelet count (PLT), mean platelet volume (MPV) and modal platelet volume (PLT-Mode) were measured using Technicon H1 hematological autoanalyser. Platelet function was assessed by aggregometry using turbidimetric method (inductors: ADP 1-3 microM, collagen 50g/ml, epinephrine 0.25-5 microM). Spontaneous platelet aggregation (SPA) was measured in platelet rich plasma (PRP) without inductors for 15 min, in 1-2 hours after venesection. SPA was observed in 9 of 30 patients with CGN and in 19 of 30 patients with CGN+NS. MPV and PLT Mode were significantly higher in patient showing SPA compared with those without. Significant correlations between SPA and the concentration of plasma albumin (r = -0,70; p < 0.02) TG and CH-LDL (r = 0,61; p < 0.05) were found in CGN+NS patients. APTT was significantly shorter in patients showing SPA compared with those without and negative significant correlation between SPA and APTT was found. Platelet aggregation to inductors in CGN and CGN+NS patients was diminished compared with control group. Lack of second phase aggregation in response to aggregation inducers was observed in patients with SPA. Conclusions. 1. Platelet hyperaggregation play an important role in hypercoagulation state in CGN patients. 2. SPA in vitro was observed in majority of CGN+NS patients and in some without NS. 3. Pathomechanism of SPA is probably multifactorial (hypoalbuminemia, dyslipidemia, changes in concentration of coagulation parameters).     

High shear stress induced platelet aggregation (h-SIPA) and effects of antiplatelet therapy

A physiologic time averaged mean shear stress in stenosed coronary artery reach more than 350 dyne/cm2. Pathologic stenosis can directly lead to shear-induced aggregation of platelets. Platelet aggregation in response to pathologically elevated shear stress is depend on the presence of plasma von Willebrand factor (vWF) and platelet receptor glycoprotein (GP) Ib/IX and GPIIb/IIIa. Fibrinogen bridging thrombus play as key factor at low shear rate, however, vWF is most important factor at high shear rate. When high shear stress are applied to vWF, vWF change the shape round to linear, and bind to extracellular matrix such as collagen type I or III exposed to blood by rupture of atheromatous plaque. Consequently vWF interact with GP Ib/IX for initial adhesion without agonist stimulation, which is followed by activation of GPIIb/IIIa receptor and co-binding with GPIIb/IIIa and vWF. The binding of platelets via vWF is strengthen to sustain the opposing effect of high shear forces in coronary artery. In our study, significant increases of h-SIPA and plasma vWF levels were observed in patients with acute coronary syndrome compared with patients with chronic coronary artery disease. The additional application of ticlopidine or cilostazol to aspirin therapy significantly inhibition of h-SIPA in patient with acute coronary syndrome, however, less effective than patients with chronic coronary artery disease.     

Effects of some antineoplastic drugs (vincristine, doxorubicin and epirubicin) on human platelet aggregation

The effects of some antineoplastic drugs (vincristine, doxorubicin and epirubicin) on collagen- and ADP-induced human platelet aggregation are investigated. Platelet rich plasma (PRP) and platelet poor plasma (PPP) from healthy male and female donors were used. The PRP was adjusted with analogous PPP to 300,000 platelets/microliters. Platelet aggregation was studied according to Born's turbidimetric technique using an Aggrecorder II PA 3220 with collagen at a concentration of 10 micrograms/ml and ADP at a concentration of 30 microM. Vincristine, doxorubicin and epirubicin significantly (p < 0.01) inhibited collagen- and ADP-induced platelet aggregation. The vincristine induced inhibition was higher than that induced by doxorubicin or epirubicin. The effects of doxorubicin and epirubicin were more intense on ADP-induced platelet aggregation than on the collagen induced one. Moreover, the doxorubicin inhibition of ADP-induced platelet aggregation was greater than the epirubicin one. In conclusion, our study shows that vincristine, doxorubicin and epirubicin inhibit human platelet aggregation. The present results may improve the therapeutic use of these drugs since it has been clearly shown that drugs with antiplatelet activity could block metastases.     

Imidazole derivatives, platelet aggregation inhibitors

Study of the platelet aggregation inhibition has shown the efficiency of compounds with imidazole ring alone or fused. All the compounds resulting of the molecular design starting from these structures has an in vitro activity. We have been able to discuss the correlation existing between activities, toxicities and structures with a particular emphasis to the lipophilicity.     

N-Acetylcysteine potentiates nitroglycerin-induced reversal of platelet aggregation

N-Acetylcysteine (N-AC) potentiates the systemic and coronary hemodynamic and antianginal effects of nitroglycerin (NGT) in humans; NTG/N-AC reduces the incidence of acute myocardial infarction in patients with unstable angina pectoris. Although previous studies have demonstrated that NTG exerts antiaggregatory effects on platelets, little information is available concerning the possible potentiation by N-AC of NTG antiplatelet effects. In the present study, we examined the in vitro effects of NTG and the combination of NTG with N-AC on reversal of ADP-induced aggregation in platelet-rich plasma (PRP) obtained from normal subjects and patients with stable angina pectoris. We also examined the potential effect of background aspirin therapy on this interaction. NTG, added to platelets 0.5 min after the beginning of aggregation, suppressed the incipient aggregation and provoked the appearance of a disaggregation phase, resulting in a concentration-dependent reversal of platelet aggregation. Platelet responsiveness to NTG was significantly less (p < 0.01) in both groups of patients (receiving and not receiving aspirin) as compared with normal subjects. N-AC (10(-5) M), which did not in itself affect aggregation, induced a threefold potentiation (p < 0.05) of the antiaggregating effect of NTG that was similar in degree for all tested groups of individuals. This potentiation of the antiplatelet effects of NTG by N-AC may contribute to the efficacy of combined NTG/N-AC therapy in patients with acute ischemic syndromes.     

Investigations of platelet aggregation and platelet counts from stored canine whole blood

The effects of the long term storage of canine blood at 4 to 6 degrees C in PVC-bags containing CPDA-1 as a stabiliser on platelet aggregation and platelet counts were investigated. Aggregation induced by collagen, adenosine diphosphate or a calcium ionophore was preserved well during the first six hours, but there was then a decrease of 24 to 46 per cent in the ability of the platelets to aggregate, after which during the next three to four weeks of storage there was no further decrease in their aggregation properties. The formation of aggregates of platelets reduced the numbers of platelets counted as single thrombocytes by more than 30 per cent during the first four days. The numbers of platelets recorded varied widely with the counting method used (counting chamber or automatically) and with the ethylenediamine tetra-acetic acid content of the dilution fluid.     

On the measurement of spontaneous platelet aggregation. The platelet aggregation test III. Methods and first clinical results

A new measuring device was developed for the study of "spontaneous" aggregating activity of thrombocytes. In the photometric platelet aggregation test (PAT III) 0.6 ml of platelet-rich plasma (PRP) are rotated in a disc-shaped cuvette at 20 rpm and 37 degrees C. Changes in optical density of PRP which are induced by the formation of platelet aggregates are continuously registered using a chart recorder. PAT III was developed for the detection of enhanced platelet aggregation, indicating a risk of thrombosis and thromboembolic complications. In 146 healthy individuals a certain percentage showed slight primary aggregation (alpha1) which in some cases was followed by marked aggregation (alpha2) at a certain time (Tr) after the beginning of rotation. The percentage of individuals showing alpha2 increased with age. An increase of plasma pH in the rotating sample, which was caused by diffusion of CO2, was an important conditioning factor for aggregation. The test results depended on the platelet count in PRP. Aggregation curves were suppressed by admixture of erythrocytes and lipid turbidity. The tendency of platelets to aggregate increased within 60-90 min following blood sampling. During this period the interval to the onset of aggregation (Tr) became shorter and the maximum aggregation speed (alpha 2) increased with time. PAT III yielded reproducible results when it was carried out more than 60 min after blood drawing. In a group of 327 diabetic patients "spontaneous" aggregation occurred more frequently in all age groups as compared with the controls. Additional equipment was available for the registration of ADP-, collagen-, or epinephrine-induced aggregation similar to Born's and O'Brien's method. The device can easily be mounted on an Eppendorf photometer without further alterations.     

Effect of xanthinol nicotinate treatment on platelet aggregation

Treatment of 24 male patients with 3 g/day of xanthinol nicotinate did not change the in vitro measurements of ADP-induced platelet aggregation but produced a marked inhibition of collagen-induced platelet aggregation. This effect may be connected with the drug-induced depression of the ATP level in platelet-rich plasma. Changes in the platelets in the patients' blood or in the lipid composition and the concentration of uric acid in their serum were ruled out as reasons for the decrease of the collagen-induced aggregation. The activity of the three serum enzymes y-GT, GOT, and GPT and the concentration of the blood sugar did not change.     

Measurement of platelet aggregation peptide inhibitors by ultrasonic interferometry

In healthy subjects, basal hepatic glucose production is (partly) regulated by paracrine intrahepatic factors. It is unknown if these paracrine factors also influence basal glucose production in infectious diseases with increased glucose production. We compared the effects of 150 mg indomethacin (n = 9), a nonendocrine stimulator of glucose production in healthy adults, and placebo (n = 7) on hepatic glucose production in Vietnamese adults with uncomplicated falciparum malaria. Glucose production was measured by primed, continuous infusion of [6,6-2H2]glucose. After indomethacin, the plasma glucose concentration and glucose production increased in all subjects from 5.3 +/- 0.1 mmol/L to a maximum of 7.1 +/- 0.3 mmol/L (P < .05) and from 17.6 +/- 0.8 micromol x kg(-1) x min(-1) to a maximum of 26.2 +/- 2.5 micromol x kg(-1) x min(-1) (P < .05), respectively. In the control group, the plasma glucose concentration and glucose production declined gradually during 4 hours from 5.4 +/- 0.2 mmol/L to 5.1 +/- 0.1 mmol/L (P < .05) and from 17.1 +/- 0.8 micromol x kg(-1) x min(-1) to 15.1 +/- 1.0 micromol x kg(-1) x min(-1) (P < .05), respectively. There were no differences in plasma concentrations of insulin, counterregulatory hormones, or cytokines between the groups. We conclude that indomethacin administration results in a transient increase in glucose production in patients with uncomplicated falciparum malaria in the absence of changes in plasma concentrations of glucoregulatory hormones or cytokines. Thus, this study indicates that in uncomplicated falciparum malaria, the rate of basal hepatic glucose production is also regulated by paracrine intrahepatic factors.     

Effects of insulin on calcium metabolism and platelet aggregation

Poppy seeds from seven different origins (Dutch, Australian, Hungarian, Spanish, Czech, and two Turkish) were analyzed for the amount of opiates present. Four grams of each kind of seeds, equivalent to the amount of seeds on two bagels, were ingested by volunteers. One volunteer also ingested four times the same amount of poppy seeds from the same origin (Spanish). During 24 hours urine samples were obtained and screened for the presence of morphine and codeine using the FPIA technique (cut-off = 200 ng/mL) and a GC/MS confirmation with a limit of detection (LOD) of 25 ng/mL for codeine and morphine. Poppy seeds from different origins contain a wide variation of morphine (2-251 micro g/g) and codeine (0.4-57.1 micro g/g) content. No other opiate could be detected. After ingestion a large interindividual variation of excretion of opiates exists. The testing results from the same kind of seeds ingested four times with a one week interval by the same volunteer also show a poor reproduceability. Several kinds of poppy seeds can give positive testing results (Australian, Hungarian, Spanish and one kind of Turkish seeds). Within 24 hours all testing results became negative.     

Relationship of platelet aggregation to bleeding after cardiopulmonary bypass

Excessive bleeding after cardiopulmonary bypass operations is a persistent problem. This study assessed the influence of platelet function on blood loss for 134 patients undergoing cardiopulmonary bypass. Platelet function was measured by platelet aggregation in platelet-rich plasma and whole blood using collagen as the agonist. Adenosine triphosphate release was assessed concurrently. Measurements were made 1 day before operation and 1 hour after the cessation of cardiopulmonary bypass. Three important findings were made. First, statistically significant correlations were shown between preoperative and postoperative platelet aggregation and blood drainage for the first 3 hours postoperatively. Second, correlations were greatest when preoperative measurement was performed on whole blood and postoperative measurement was performed on platelet-rich plasma. Third, patients with reduced postoperative platelet aggregation in platelet-rich plasma had significantly greater transfusion requirements in the first 24 hours postoperatively. In defining the 16 patients who bled excessively among the 134 patients studied, the preoperative aggregation in whole blood had a sensitivity of 62%, specificity of 75%, positive predictive value of 26%, and negative predictive value of 94%. The postoperative aggregation in platelet-rich plasma had a sensitivity of 86%, specificity of 69%, positive predictive value of 28%, and negative predictive value of 97%. These results indicate that preoperative and postoperative measurement of platelet aggregation may provide a rationale for the prophylaxis or treatment of patients to reduce blood loss after cardiopulmonary bypass.