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The findings of a large prospective study designed to identify primary and/or secondary haemostatic disorders before surgical interventions are presented. A total of 5649 unselected adult patients were enrolled to identify impaired haemostasis before surgical interventions. Each patient was asked to answer a standardized questionnaire concerning bleeding history. Activated partial thromboplastin time (aPTT), prothrombin time (PT), and platelet counts (PC) including PFA-100 (platelet function analyzer): collagen-epinephrine (C/E), and collagen-ADP (C/ADP) were routinely done in all patients. Additional tests, bleeding time (BT), von Willebrand factor (VWF:Ag, VWF:Rcof) and a further haemostaseological diagnostic was performed only in patients with a positive bleeding history and/or evidence of impaired haemostasis; e.g., drug ingestion. The bleeding history was negative in 5021 patients (88.8%) but positive in the remaining 628 (11.2%). Impaired haemostasis could be verified only in 256 (40.8%) of these patients. The vast majority was identified with PFA-100: C/E (n = 250; 97.7%). The sensitivity of the PFA-100: collagen-epinephrine was the highest (90.8%) in comparison to the other screening tests (BT, aPTT, PT, VWF : Ag). The positive predictive value (to detection of impaired haemostasis) of the PFA-100: collagen-epinephrine with the standardized questionnaire was high (82%), but the negative predictive value was higher (93%). The use of a standardized questionnaire and, if indicated, the PFA-100: C/E and/or other specific tests not only ensure the detection of impaired haemostasis in almost every case but also a significant reduction of the costs. Based on these data, national regards are formulated or under construction.

Therapy with acetylsalicylic acid (ASA) and/or clopidogrel is used to achieve prophylactic inhibition of platelet aggregation in patients with arterial thrombosis. We examined if aggregometry can be used to see the effect of antiplatelet drugs (ASA 30, 50, 100, 300 mg/d, clopidogrel 75 mg/d or ASA 100 + clopidogrel 75 mg/d). A modified platelet aggregation test was used to investigate maximum aggregation in response to ADP, collagen, adrenalin and arachidonic acid. Reference values were established based on healthy individuals. We devised a simple scoring system for detection of inadequate platelet inhibition. Compared with the control group, we detected a significant delay of maximum aggregation in response to all agonists in patients on ASA and combination therapy ASA + clopidogrel. Patients on clopidogrel alone were found to have prolonged aggregation when induced with ADP, collagen and arachidonic acid. The failure rate to achieve adequate platelet inhibition on 100 mg/d ASA, 75 mg/d clopidogrel or combination therapy was 27%, 26% and 7%, respectively. Our results demonstrate that platelet inhibition in aggregometry is inadequate in many patients with arterial thrombosis.

The influence of heparin on different tests of platelet function was investigated in 5 healthy dogs receiving subcutaneously 1000 I.E./kg BW of a commercial unfractionated Sodium heparin preparation. Blood samples were collected before and 4 hours after heparin injection. Besides plasma activity of heparin platelet count, capillary bleeding time with two different methods, platelet aggregation according to Breddin and to Born with the inductors ADP, collagen, and thrombin as well as in vitro bleeding time were measured. The activity of heparin four hours after the subcutaneous heparin application was 1.20 +/- 0.11 I.E./ml. Compared to the starting values no significant influence of this heparin level could be demonstrated on platelet count, platelet aggregation according to Born with the inductors ADP and collagen, platelet aggregation according to the Breddin method as well as in vitro bleeding time (p > 0.05). Only one of the two methods used for measuring the capillary bleeding time showed a slight prolongation compared to the starting value (mean = 77 sec) to 88 sec (p < 0.05). The most significant influence was seen on the platelet aggregation induced by 1 I.U./ml thrombin, whereby the aggregation maximum decreased from 92.0% (mean) to 12.2% (p < 0.001). Whereas the latter result has to be interpreted primarily as a consequence of the anti-thrombin effect of heparin, the results of the other tests in summary illustrate the clinical unimportant influence of heparin on platelet function in dogs.

The possibility of storage of canine platelet concentrates (PC) was investigated using PC from dogs which were obtained with an automatic cell separator in C4-cell separation sets with low gasdiffusionable Polyvinylchlorid (PVC) storage containers or in C4L-sets developed for storage with high gasdiffusionable Polyolefin (PO) containers, respectively. The storage was carried out for a period of 10 days under permanent agitation at 22 degrees C (C4/22 degrees C, n = 10; C4L/22 degrees C, n = 11) or at 4 degrees C (C4L/4 degrees C, n = 6), respectively. Measurements were done directly after production of the PC, after 6 hours and then daily during the 10-day storage period. In the first part of this paper the results of platelet count (determined automatically with a blood cell differentiation automat and visually), the number of platelet aggregates, the mean platelet volume (MPV) as well as the platelet function with regard to the platelet aggregation induced by collagen or ADP and the resonance-thrombogram (RTG) are presented. The platelet count, measured automatically as well as visually, remained preponderantly constant over the complete storage time in all storage conditions. Dependent on the storage conditions--especially under storage at 22 degrees C--an increase of the number of platelet aggregates and a decrease of MPV was determined. In addition, the loss of platelet function measured by aggregation induced by collagen as well as by ADP showed a significant dependency of storage conditions. The stored platelets lost their ability to aggregate under C4/22 degrees C-conditions after a storage period of 2 days, under C4L/22 degrees C-conditions after 4 days and under C4L/4 degrees C-conditions not before 8 days of storage. Previous resuspending of platelets in fresh plasma delayed the loss of platelet function. Because the loss of platelet function described in the RTG became significant at nearly the same point in time, a storage of canine PC under corresponding conditions can be recommended for upto 2 days (C4/22 degrees C), for 4 days (C4L/22 degrees C) or 8-10 days (C4L/4 degrees C), respectively.

The influence of acetylsalicylic acid (ASA) in vitro and in vivo on several parameters of the hemostatic system was evaluated in healthy cats. The aggregation response of cat platelets in vitro was not influenced by 10 micrograms ASA/ml plasma, whereas ASA concentrations of 35, 70, 100 micrograms/ml and of 1 mg/ml consistently inhibited the platelet aggregation induced by 0.1 (n = 4 cats) and 0.25 micrograms collagen/ml (n = 6 cats). The aggregation response to ADP and thrombin was partially (n = 2) or completely (n = 8) inhibited by an ASA concentration of 5 mg/ml. Administration of ASA at a dosage of 10 and 25 mg/kg BW orally and intravenously yielded a plasma concentration of salicylic acid of 30-42 micrograms/ml (10 mg/kg BW) and 50-70 micrograms/ml (25 mg/kg BW). All the dosages of ASA produced an inconsistent inhibition of the collagen-induced platelet aggregation (1/5 and 2/5 for the respective groups), whereas the response to ADP and thrombin was not altered. Capillary bleeding time was not significantly altered after the ASA administration. Platelet count and screening tests were not influenced by ASA application. According to the results of this study, ASA as a platelet aggregation inhibitor is not useful for the prophylaxis of thromboembolism in cats.

In order to establish an univariate nonparametric pediatric tolerance region platelet function has been investigated in 105 healthy children and adolescents. In comparison to adult normal values, the bleeding time is shortened, spontaneous platelet aggregation is enhanced as well as collagen-induced platelet aggregation. 30% of the children showed an increased disaggregation in ADP-induced aggregation. A slight delay was found in the spreading of thrombocytes. Platelet volume shifted to the left. Values of beta-thromboglobulin were raised. Compared to adult values no alterations could be found in platelet shape-change. Changes of platelet functions were more apparent in the younger children.

Platelet count, spontaneous platelet aggregation, ADP- and collagen-induced platelet aggregation platelet adhesion, platelet volume, shape change, beta-thromboglobulin and von-Willebrand-factor have been investigated in 51 insulin dependent diabetic children without clinical signs of diabetic angiopathy. Compared to an age matched healthy control group diabetic children showed a significant enhancement of spontaneous platelet aggregation, elevated plasma levels of von-Willebrand-factor, increased platelet shape change and adhesion. No alterations could be found in ADP--and collagen--induced platelet aggregation and in beta-thromboglobulin levels. Significant correlations could be found between the total glycosylated haemoglobin concentrations (Hb A1) and spontaneous platelet aggregation, as well as between duration of diabetes Hb A1, and platelet volume. In this study we could demonstrate changes in platelet function in diabetic children without clinical signs of diabetic angiopathy. However these changes could be due to metabolic adjustment and may precede diabetic vasculopathy.

In blood samples of 12 healthy volunteers (5 female, 7 male) the influence of Gallopamil (Procorum, CAS 16662-47-8) on the adenosine-diphosphate- (ADP-) and collagen-induced platelet aggregation was studied. Measurements in platelet rich plasma after induction with 1 mumol ADP and 0.5 microgramml collagen with gallopamil were performed. After incubation mit 250 mumol gallopamil a second test was carried out. The parameters were compared with the U-test. After application of 250 mumol gallopamil a significant reduction of ADP- and collagen-induced platelet aggregation could be observed. This is a sign for a protective effect of gallopamil in atherosclerotic disorders due to its calcium-channel blocking property.

Platelet aggregation in healthy and sick cats after adding various aggregating agents is described. Feline platelets aggregate irreversibly in response to 0.15-1.0 micrograms/ml collagen, 1 microM ADP, 0.3 IU/ml test-thrombin and 0.71 NIH/ml Topostasin. Epinephrine, ristocetin and kaolin failed to cause aggregation. The aggregation function was decreased in a cat with liver damage and icterus; in 2 cats with uremia platelet aggregation was normal. Acetylsalicylic acid (ASA) (10 mg/kg iv) inhibits platelet aggregation in the presence of collagen in low concentrations; high concentrations of collagen succeeded in inducing platelet aggregation.

Sera of patients with RA and SLE contain platelet-reactive IgG antibodies, which bring about the release of serotonin from platelets. They can also induce the aggregation of normal human platelets. In addition, such antibodies cause a decreased in vitro disaggregation of platelets after ADP exposure, as well as a significantly reduced collagen-induced aggregation. Thus, IgG platelet antibodies in RA and SLE can cause a functional impairment of patients' platelets, and may also be instrumental in systemic inflammatory rheumatoid disease by releasing proinflammatory mediators.

Patients with thrombocythaemia due to myeloproliferative disorders (n = 21), with secondary thrombocytosis of various origin (n = 16), and a control group of healthy donors (n = 20) were investigated with respect to the aggregation behaviour and the total calcium content of blood platelets. The calcium content was significantly lower in both groups of patients as compared to controls (2 p less than 0.001). In 16 of 21 patients with myeloproliferative disorders platelet rich plasma did not respond to epinephrine (15 mumol/l), a concentration which induced at least weak aggregation in 14 of 16 patients with secondary thrombocytosis and also in healthy subjects. In patients with thrombocythaemia the mean extent of aggregation induced by epinephrine, collagen or adenosin diphosphate was significantly lower as compared to controls (2 p less than 0.001).

The ADP-induced primary aggregation of human thrombocytes in vitro was inhibited by 1.5X10(-4) mol/l (+/-)bupranolol. At this concentration a strong desaggregation was obtained. The stereoisomers of bupranolol did not differ in effectiveness. Fractional addition was just as effective as a single dose. Not beta-receptor blocking but a membrane stabilizing effect is, therefore, the probable mechanism -- as in the case of propranolol. The less lipophilic metabolites hydroxybupranolol and carboxybupranolol were one-fourth and one-fifth as effective, respectively, as bupranolol. 5X10(-4) mol/l ASA (final concentration) showed the same inhibition of primary aggregation as did 1.5X10(-4) mol/l bupranolol but elicited less desaggregation. Doubling this dose of ASA resulted in a weaker inhibition than the subsequent addition of the aforementioned doses of bupranolol and ASA or vice versa, while this combination, in turn, inhibited less than a double dose of bupranolol. 1.5X10(-4) mol/l bupranolol inhibited the formation of pseudopodia by thrombocytes not stimulated with ADP. At 3X10(-4) mol/l this inhibition was drastic but no morphological damage was seen.

Lipoxygenase from rabbit reticulocytes cause disruption of mitochondrial membranes and peroxidation of their lipids as judged by electronmicroscopy, release of matrix enzymes and formation of malonyldialdehyde. Without substrate mitochondria become orthodox and strong lysis by lipoxygenase appears. The lysis is prevented by ATP or ADP plus succinate; in this case mitochondria remain condensed or partly condensed. The protection by substrate was even observed in the presence of 2,4-DNP, although the mitochondria were transformed to the condensed state. Lysis was more pronounced in hypotonic than in hypertonic sucrose, condensed mitochondria are also attacked. No relation seems to exist between lipoxygenase attack and the conformational state of mitochondria. Lysis of mitochondria is dependent on the susceptibility of the fatty acid moiety of phospholipids, which may be influenced by both metabolic and structural events via alteration of protein-lipid interactions.

The secretolytic agent bromhexine and its active metabolite ambroxol prevent the collagen- and ADP-induced decrease in circulating blood platelets in rabbits, while they do not inhibit the aggregation induced by ADP, collagen or arachidonic acid of human and rabbit blood platelets in vitro. Bromhexine was found to exert beneficial effects on the disturbances of pulmonary function that might be caused by a reduced surfactant production observed during thrombin-induced disseminated intravascular coagulation in rats. The results suggest that bromhexine has a protective effect on the acute impairment of pulmonary function due to microthrombosis.

The oxidoreductase inhibitor is not formed from NADH as previously thought, but only from NAD under alkaline conditions. Analogues of NAD (e.g. NADP) and components of the NAD molecule (e.g. ADP) have no effect on the formation of the inhibitor. The most favourable pH, temperature, duration of incubation, type of buffer and NAD concentration for the formation of the inhibitor were investigated. The method for the formation and chromatographic isolation of the oxidoreductase inhibitor is briefly described.

The incubation of rabbit venous rings (vena cava) is followed by the synthesis and release of prostacyclin (prostaglandin I2, PGI2), as measured by the inhibition of ADP-induced platelet aggregation. The incubation of the venous rings in dihydroergotamine leads to a stimulation of PGI2 synthesis and/or release. This effect of dihydroergotamine, apart from the vasoactive properties of this compound, might prove advantageous as a prophylactic measure in venous thrombosis.

Platelet function (ADP-induced platelet aggregation) in 20 patients with juvenile-onset diabetes mellitus was not different from that of age- and sex-matched controls. Platelet sensitivity to exogenous PGI2 was not diminished in diabetic patients. No sex differences were detected. There was no correlation between concomitant blood glucose levels and platelet sensitivity to PGI2.

3-(2-Diethylaminoethyl)4-methyl-7-(carbethoxy-methoxy)-2-oxo-1,2-chromene-hydrochloride (carbocromen; Intensaïn) shows dose dependent platelet aggregation inhibitory activity in vitro according to the methods of Born (following ADP, epinephrine and collagen) and of Breddin (platelet agglutination test--PAT--, adhesively). The effect was found to be more pronounced than that of acetylsalicylic acid. Furthermore, in vivo carbocromen inhibited the increased spontaneous platelet aggregation in man.

The influence of inductors and inhibitors of platelet aggregation on certain functional properties and structural peculiarities of platelets are investigated. Inductors of aggregation, such as ADP and thrombin, will cause the calcium binding to the platelet membrane to be diminished, whereas inhibitory substances of platelet aggregation will cause an increase of calcium binding. Inductors increase the number of partially degranulated platelets. Inhibitors, such as triphtazin and papaverin, inhibit these morphological changes.

ADP and collagen induced platelet aggregation (PA) was studied in 18 patients suffering from arterial occlusive disease (stage II Fontaine) before and after a standardized treadmill test before and after 24 days of exercise training. At the beginning of this therapy collagen induced PA after treadmill testing was increased (amplitude +11.29%, p less than 0.05, maximal slope +18.57%, p less than 0.05, integrated area +15.23%, p less than 0.05), whereas ADP induced PA showed no consistent alteration. After the 24-day treatment period increase in collagen induced PA was considerably smaller (amplitude +7.48%, n.s., maximal slope +7.35%, n.s., integrated area +11.99%, p less than 0.05), whereas ADP induced PA again showed no consistent changes. Since the treadmill exercise induced PA showed a negative linear correlation with the claudication provoking walking distance the smaller increase in PA after treatment was considered to be due to the improved metabolic situation following exercise therapy.

The effects on platelet function of a four-week administration of aspirin at a low dosage (100 mg daily) were compared in two groups, 14 healthy young volunteers and 14 patients with coronary heart disease. In both groups there occurred a clear inhibition of platelet aggregation with collagen (1 and 5 micrograms/l) and arachidonic acid (1 mmol/l) during the aspirin period. The inhibitory effect reached its maximum after three days, remaining at maximum for the remainder of the four weeks. Platelet functions returned to normal within eight days of discontinuing aspirin. The inhibitory effects went together with a definite in-vitro decrease in thromboxane synthesis. In both groups there was no change in aggregation velocity with adenosine diphosphate (ADP) as aggregation-inducing substance, while the frequency of irreversible aggregation decreased with submaximal concentrations of ADP (0.5 and 1.0 mumol/l). The results indicate that low-dose aspirin causes a definite inhibition of platelet function, in a similar manner, in both healthy subjects and patients with coronary heart disease.

It is the purpose of this study to standardize platelet aggregation according to the method of Born. It was found that aggregation is influenced by the time of storage, the pH and temperature of plasma. However, there is no significant correlation between platelet number versus aggregation in healthy subjects. To get reproducible results, the plasma samples should be investigated within 2 hours after venipuncture. During storage temperature of all samples should be constant.

Heparin added to citrated platelet rich plasma influences shape change and aggregation of platelets in different ways. In the presence of heparin neither ADP nor collagen induces shape change, while shape change after thrombin or arachidonic acid remains unaltered. Heparin potentiates the first aggregation step induced by ADP and epinephrine but inhibits aggregation induced by thrombin and ristocetin. The second phase of aggregation and the release reaction are not directly influenced by heparin no matter which aggregation agent is used.

Polarographic titration in a closed cell of blood samples and concentrated Hb solutions was used to study the regulation of the release of oxygen by the red cell. The effect of temperature, polyethylenglycol and metabolites normally found in the erythrocyte was determined by spectrophotometric and polarographic methods. Some pecularities of O2-transport and release in beta-thalassemia, functional hypoxia and Cooley-disease are described.

A long term incubation in NBT-solution leads not only to NBT-reduction by polymorphnuclear leukocytes and monocytes, but also in washed human platelets. The reaction is enhanced by ADP and to a smaller and very variable extent also by Thrombin and Ristocetin. It is inhibited by incubation in plasma environment as well by DBcAMP and PGE1. After a detailed demonstration of the applied method, some possible biochemical mechanisms of the NBT-reduction in human platelets are discussed.

ADP and collagen induced platelet aggregation were determined in 34 newborns immediately after birth. Blood was drawn from the pulsating not clamped umbilical vein. Parameters of aggregation were correlated to pH values of umbilical artery and vein, Apgar score and to the duration of the first and second stage of labor. No significant correlations were found between parameters of labor and platelet aggregation. It is demonstrated, however, that a decrease of the pH values in the umbilical artery and vein is well correlated with an increase of the main parameters of platelet aggregation. Changes of disaggregation of platelets were only moderate, they showed a tendency to decrease with falling pH values. Altogether there is an increase of platelet aggregability if pH values go down. The results emphasize that the changes of platelet function described should be regarded in the concept of therapy of newborns with severe acidosis or shock as they may accentuate preexisting hypercoagulability.

A new measuring device for the estimation of the "spontaneous" aggregating activity of thrombocytes has been developed. In this photometric platelet aggregation test (PAT III) a small amount (0.6 ml) of platelet-rich plasma (PRP) is being rotated in a disc-shaped cuvette at 20 rpm, at 37% C. Changes in optical density of PRP which are induced by the formation of platelet aggregates are continuously registered using a chart recorder. The decisive trigger mechanism for aggregation is an increase of plasma pH in the rotating sample which is caused by evaporation of CO2. The results of the test depend on the platelet count in PRP. Aggregation curves are misrepresented by admixture of erythrocytes and lipid turbidity. The tendency of platelets to aggregate increases within 60-90 min following blood sampling. During this period the time interval to the onset of aggregation(Tr) is shortening, and the maximum aggregation speed (alpha2) is increasing. The spontaneously enhanced aggregation tendency of thrombocytes may be reliably measured from 60 min after drawing the blood. The reason for these time-dependent changes which are also demonstrable in ADP-collagen-or epinephrine-induced aggregation is probably the primary shape change of platelets, which occurs after blood drawing and makes them "stickly" and aggregable. PAT III was developed for the detection of enhanced platelet aggregation, indicating a risk of thrombosis and thromboembolic omplications. The new measuring device has been designed as "universal" aggregometer. Additional equipment is available for the registration of ADP-collagen-or epinephrine-induced aggregation similar to Born's and O'Brien's methods. The device may be mounted easily on an Eppendorf photometer without further modifications.

The action of 3-[4,5-bis-(4-chlorophenyl)-oxazolyl-(2)-thio]-propionic acid (tioxaprofen, EMD 26 644), a non-steroidal antiinflammatory substance, on platelet function has been studied in vitro and ex vitro. Collagen-induced aggregation and the second phase of adrenaline-induced aggregation are inhibited by small concentrations of tioxaprofen, The ADP-induced aggregation being inhibited only by higher concentrations. The generation of malonedialdehyde (MDA) is blocked. The pathological spontaneous aggregation found by the method of Born was reversed after treatment, and the pathologically increased aggregation according to platelet aggregation test (PAT I) of Breddin was normalized. Tioxaprofen blocks the formation of thromboxane most probably by inhibition of cyclo-oxygenase, but an inhibition of thromboxane synthetase cannot be excluded by our results. The in vivo action starts immediately after ingestion of tioxaprofen and is maintained by small doses. By checking the routine parameters of the blood a slight increase of creatinine and blood urea nitrogen (BUN) and a slight reduction of gamma-GT and alkaline phosphatase were found, all values remaining within the reference interval.

Platelet aggregation was measured in rat plasma at pH values between 6.0 and 8.0 at temperatures, of 21 degrees, 37 degrees and 43 degrees C, respectively, and under aerobic and anaerobic conditions. Platelet aggregation inhibition was predominant at neutral and weakly acidic pH values. At 37 degrees C, aggregation and desaggregation were markedly accelerated as compared to 21 degrees C. In contrast, platelet aggregation at 43 degrees C was not or at most insignificantly increased in comparison to 37 degrees C-Oxygen depletion had no measurable effect. It is concluded from the results obtained that the microcirculation inhibition detected in rats and clinically utilized in the Cancer Multistep Therapy (tumor acidification plus hyperthermia) is highly likely not mediated by platelet aggregation.

The platelet aggregation inhibiting effect of N-[3-(1-benzyl-cycloheptyloxy)-propyl]-N,N-dimethyl-amine (bencyclan, Fludilat) was studied by a new method for quantification of platelet aggregation in viscometric flow. In vitro-addition of bencyclan shows significant inhibition in a concentration of 1 mg/100 ml, total inhibition of platelet aggregation is found at a concentration of 5 mg/100 ml. Peroral application of bencyclan shows no significant effect on platelet aggregation. Intravenous application of bencyclan studied in three patients results in inhibition of platelet aggregation, however, no information about the duration of this platelet-inhibiting effect is yielded.