Correlation between plasma levels of growth factors and von Willebrand factor.

Intimal proliferation of smooth muscle cells is an important characteristic of developing atherosclerotic lesions and late occlusion of venous bypass grafts. Platelet-derived growth factor, released from aggregating platelets at sites of endothelial injury, has been suggested as a main factor responsible for intimal hyperplasia. The von Willebrand factor is a platelet binding protein secreted by endothelial cells and increased plasma levels of this factor has been identified as a marker of endothelial injury. In the present study we have analysed plasma levels of von Willebrand factor and growth factors in healthy controls, young post-infarction patients and patients with recent coronary bypass surgery. The results demonstrate a significant correlation between plasma growth factor activity and the level of von Willebrand factor (R = 0.52, p less than 0.01) and support the notion of a coupling between endothelial injury and release of platelets mitogens.

Platelet activation in patients with mitral valve prolapse.

An increased frequency of thromboembolic events in patients with mitral valve prolapse has been demonstrated. It has been suggested that this association may be related to increased systemic platelet activity. Beta-thromboglobulin (BTG) is a platelet specific protein secreted during the platelet release reaction, with BTG levels reflecting ongoing platelet activation. Plasma BTG levels were measured in 14 normal volunteers, 23 patients with mitral valve prolapse and nonthickened mitral leaflets (group 1) and 13 patients with mitral valve prolapse and thickened mitral leaflets (group 2). The BTG level was 8.1 +/- 4.6 ng/mL (mean +/- SD) in normal subjects, 9.6 +/- 5.5 ng/mL in the nonthickened mitral valve prolapse group and 10.0 +/- 5.7 ng/mL in thickened mitral valve prolapse group. There was no significant difference in the BTG levels between groups. Five patients with multiple valvular prolapse did not show elevation of BTG levels. The present study did not demonstrate increased BTG levels in neurologically asymptomatic mitral valve prolapse patients.

Platelet specific proteins (beta-thromboglobulin and platelet factor 4) in normal pregnancy and in pregnancy complicated by preeclampsia.

Pregnancy is accompanied by significant alterations of platelet function. Platelet activity can be determined by measurement of plasma levels of secreted platelet proteins. In this study we determined plasma levels of beta-Thromboglobulin (beta-TG) and Platelet Factor 4 (PF4) simultaneously in 35 women with uncomplicated pregnancy and in 15 patients with preeclampsia in third trimester of gestation. Additionally, PF4 plasma levels were measured using a commercially available Radio Immunoassay (RIA) and an Enzyme Immunoassay (EIA) simultaneously and values obtained were compared. Platelet count and creatinine were in the normal range in both groups; however, significantly higher levels of beta-TG (P less than 0.0005) and PF4 (P less than 0.0001) were found in case of preeclampsia. High levels of platelet proteins emphasize the active role of the platelets in the alterations of hemostasis in cases of preeclampsia.

Prostaglandin D2 and histamine release in cold urticaria unaccompanied by evidence of platelet activation.

Six patients with acquired primary cold urticaria and six normal control subjects were challenged with a 5-minute immersion of an arm in cold water, at 10 degrees C, to induce cold urticaria. Venous blood draining the arm was sampled before and at 5 and 20 minutes after challenge. Prostaglandin D2 levels in the serum increased significantly after cold challenge but did not correlate with the severity of the urticaria. Significant elevations in histamine after cold challenge tended to be higher in the patients with a low threshold to cold reaction. Two markers of platelet activation, platelet factor 4 and beta-thromboglobulin, remained at basal levels 5 minutes and 20 minutes after challenge.

Influence of urapidil on in vitro platelet response to adrenaline and other aggregating agents.

The effects of hypotensive drug urapidil on human platelet functions were investigated. Urapidil failed to evidence direct aggregating properties or potentiating effects. Furthermore, drug high concentrations inhibited the platelet response to ADP, PAF, collagen, adrenaline and bovine thrombin, and influenced the platelet release reaction induced by ADP and PAF. Data indicate that urapidil possesses negligible agonistic effects on human platelet alpha 2-adrenoceptors and interferes at high concentrations with the platelet activation, as evidenced for other anti-aggregating compounds.

Periarticular fibrosis associated with idiopathic retroperitoneal fibrosis.

We describe a previously unreported association of retroperitoneal fibrosis with biopsy proven periarticular inflammatory fibrosis. Histologic features were strikingly similar at the 2 sites. Plasma levels of a platelet derived growth factor, connective tissue activating peptide III, were found to be elevated and may play a role in pathogenesis. A dramatic response of the periarticular process to corticosteroids correlated with improvement of other variables of disease activity.

Platelet alpha granule secretion in cerebral ischemia: effect of short and long term low dose aspirin treatment.

The effect of short and long-term therapy with aspirin (50 mg/day) on platelet alpha granule secretion was studied in 11 healthy controls and 57 patients suffering from transient cerebral ischemic attacks (TIA) with and without accompanying diabetes and hypertension. Plasma levels of beta-thromboglobulin (beta-TG) and platelet factor 4 (PF 4) were measured as indicators of platelet alpha granule secretion. beta-TG and PF 4 levels were increased following cerebral ischemia. Aspirin treatment failed to suppress plasma levels of both proteins when measured a month and then a year after initiation of treatment. Therefore, these proteins may be poor indicators of platelet inhibition by aspirin.

Effects of prolonged picotamide therapy on platelet activity in patients with peripheral arterial disease.

An open study was carried out in 14 patients with peripheral arterial disease to investigate the effects of prolonged therapy with picotamide on platelet activity. Patients received daily oral doses of 900 mg picotamide for 1 month, 600 mg per day during the second month and 300 mg per day from the third to the sixth month of the study. Measurements were made before and during therapy of blood coagulation parameters and factors influencing platelet function, i.e. plasma beta-thromboglobulin and serum thromboxane B2. The results showed that there were no significant variations in platelet count, prothrombin time, partially activated thromboplastin time, presence and amount of fibrinogen in blood, and antithrombin III. Examination for fibrinogen degradation products was constantly negative and unaltered during therapy. Although plasma beta-thromboglobulin values did not vary significantly, there was a significant and progressive reduction throughout treatment in serum levels of thromboxane B2.

Plasma and urine beta-thromboglobulin in severe preeclampsia.

Beta-thromboglobulin (BTG) has been shown to be a specific platelet protein and can be used as a marker of platelet activation in preeclampsia. Concomitant studies of BTG levels in plasma and urine were performed with eight primiparous severe preeclamptic patients and eight normal primiparous women matched for age. The mean plasma BTG in the severe preeclamptic patients was 186.62 +/- 29.93 ng/ml, and in the control group 45.38 +/- 31.84 ng/ml. The P-value for the difference was highly significant (P = 0.000). In contrast, the mean urine BTG in the study group was 8.42 +/- 4.61 ng/ml, while the mean value for the control group was similar, 5.00 +/- 3.20 ng/ml. The P-value for the difference was not significant (0.05 less than P less than 0.10). These results show that urinary BTG cannot be considered an indicator of platelet activation in severe preeclampsia. A low urinary BTG concentration in the presence of high plasma BTG levels may rather express renal impairment. Failure of BTG renal clearance would contribute to further raising the level of plasma BTG.

Caffeine stimulates in vivo platelet reactivity.

The effect of coffee drinking on platelet reactivity was studied in 12 healthy subjects. Plasma beta-thromboglobulin concentration was determined before and one hour after administration of 100 mg of caffeine, corresponding to one cup of coffee. Mean values were 47.0 +/- 19.3 and 179.3 +/- 85.5 ng/ml before and after caffeine administration respectively. The increase, 298 +/- 150%, is highly significant (p less than 0.001).

Low platelet arachidonic acid in young patients with brain infarction.

Fatty acid patterns of plasma and platelet lipids, platelet aggregation and thromboxane A2 (TxA2) production were studied in young patients (n = 12) with brain infarction and in healthy controls (n = 13). Platelet arachidonic acid content was significantly reduced in the stroke patients, but in vitro platelet aggregation was similar in the two groups. A low dose of acetosalicylic acid (ASA) (100 mg) suppressed thromboxane production and normalized the platelet arachidonic acid values. The low arachidonic acid in platelets is probably due to its increased consumption, indicating platelet activation in vivo.

[Prevention of thromboembolism during pregnancy in 3 sisters with congenital antithrombin III deficiency and decreased inducible fibrinolysis]. / Thromboembolieprophylaxe während der Schwangerschaft bei 3 Schwestern mit kongenitalem Antithrombin III-Mangel und vermindert stimulierbarer Fibrinolyse.

Antithrombin III (AT III) deficiency is associated with a high risk of venous thromboembolism, particularly in pregnancy. As prophylactic treatment it has been recommended that plasma levels of AT III be normalized by use of AT III concentrates, in addition to heparin. - We report on the prophylactic treatment of three sisters (age 21, 25, and 32 years) with congenital AT III deficiency (38-53%, normal 80-120%) and reduced inducible fibrinolytic activity (1.2, 5.8, 1.2%, normal greater than 8.5%), who had already had severe thromboembolism. During pregnancy prophylactic measures were taken individually, depending on the plasma level of beta thromboglobulin (BTG) determined every 2-3 weeks. In two patients prophylaxis with s.c. heparin (2 X 7500 IU/d) was started at the time of the first increase of BTG (around 10th week of gestation), leading to normalization of BTG. When BTG was again elevated, the dose of heparin was increased stepwise to 2 x 15,000 IU/d; in this way functional AT III levels remained in the range of 28-50%. Thus, these two patients received only heparin throughout pregnancy. However, in the third patient BTG levels could not be normalized by heparin alone (60-130 ng/ml, normal less than 43 ng/ml). Injections of AT III concentrate (1000 IU) led to reduction of BTG within 2 hours (60----42,220----61 ng/ml). Therefore, AT III was given from the 25th week of gestation in increasing amounts up to 5000 IU/week (funct. AT III in plasma 51-72%) in addition to heparin (2 x 12,500 IU/d), resulting in BTG levels of 33-51 ng/ml.(ABSTRACT TRUNCATED AT 250 WORDS)

Failure of platelet markers to evaluate membrane biocompatibility during a combined hemodialysis-hemoperfusion treatment.

Plasma levels of the platelet markers beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) are among the most sophisticated indexes of biocompatibility available to evaluate new members for hemodialysis. This investigation was designed to determine the extent of platelet activation by measuring the alpha-granule release products, beta-TG and PF4; anticoagulation and thrombogenesis by monitoring plasma heparin; and fibrinopeptide A (FPA) and thromboxane B2 (TX B2) levels during treatment with a combined hemodialysis-hemoperfusion system. Both in vivo and in vitro results showed that the platelet markers had a pattern different from that generally observed during treatment with hemodialysis alone. This is due to the avidity of charcoal for the markers studied, which therefore cannot be used to evaluate the biocompatibility of the system.

A formula for correcting for the in vitro release of platelet beta-thromboglobulin.

The interpretation of platelet beta-thromboglobulin (BTG) and platelet factor 4 (PF4) levels as indicators of in vivo platelet activation is complicated by the artefactual release of these proteins in vitro. A formula was devised to correct for in vitro platelet activation and release of BTG. Blood was collected from normal volunteers by an ideal method and BTG and PF4 levels determined by radioimmunoassay; these were the reference values. Blood from normal volunteers was activated in vitro by standing at room temperature. The BTG and PF4 released was measured at different time intervals. The relationship between BTG and PF4 released was measured at different time intervals. The relationship between BTG and PF4 was measured mathematically best described by a second degree polynomial function. The true plasma BTG value was then calculated by correcting for in vitro release by the general formula: BTG corrected = BTG measured - BtG for PF4 measured + BtG for PF4 reference The validity of the correction formula was tested in 10 normal subjects and in patients with either recent myocardial infarction(n = 10), familial hypercholesterolaemia(n = 10) or arterial prostheses(n = 14). Correction was adequate in normal subjects if the plasma BTG levels did not exceed 260ng/ml. In patients with a thrombotic tendency, the formula overcorrected for in vitro release. This could be ascribed to increased in vivo PF4 levels in these patients, especially those with prostheses. The reference values for PF4 in these patients, and especially those with vascular prostheses, were also higher than normal. The PF4 measured in their plasma thus reflects both in vivo and in vitro released protein. The hypothesis on which the correction formula was based, is therefore not always applicable. It may be possible to improve the correction by establishing formulae for specific disease groups.

Role of alpha 2-adrenergic receptor in platelet activation during insulin-induced hypoglycemia in normal subjects.

The effects of alpha 2-adrenergic-receptor blocker mianserin on the responses of blood glucose, plasma beta-thromboglobulin (beta-TG), and various counterregulatory hormones to insulin-induced hypoglycemia were studied in nine healthy male subjects. The alpha 2-adrenoceptor-blocking action of mianserin was confirmed by its inhibitory effect on platelet activation in vitro. Mianserin was given orally 90 min before insulin injection; the same study without mianserin was performed on another day as the control study. The time courses of blood glucose and serum C-peptide (0, 20, 45, and 180 min after the insulin injection) were identical in both studies, indicating that mianserin has no effect on these parameters. However, a significant increase of beta-TG at 45 min after insulin injection was completely suppressed by the administration of mianserin (mean +/- SE, 68.5 +/- 6.0 vs. 28.8 +/- 7.6 ng/ml, n = 6, P less than .05). No significant differences were obtained between the two studies in the responses of plasma or serum catecholamines, cortisol, glucagon, growth hormone, thromboxane B2, and 6-ketoprostaglandin F1 alpha. These results suggest that epinephrine is responsible for some, if not all, of the beta-TG release from the platelets during insulin-induced hypoglycemia.

Reliability of a single beta-thromboglobulin measurement in a diabetic population: importance of PGE1 in anticoagulant mixture. Damad Study Group.

During the collection of samples for plasma beta-thromboglobulin (beta-TG) determination, it is well established that artificially high values can be observed due to in-vitro release. To estimate the reliability of a single beta-TG measurement, blood samples were collected simultaneously from both arms on two separate occasions in 56 diabetic patients selected for a clinical trial. From each arm, blood was taken into two tubes containing an anticoagulant mixture with (tube A) and without (tube B) PGE1. The overall mean value of B-TG in tube B was 1.14 times higher than in tube A (p less than 0.01). The markedly large between-arms variation accounted for the most part of within-subject variation in both tubes and was significantly greater in tube B than in tube A. Based on the difference between B-TG values from both arms, the number of subjects with artificially high B-TG values was significantly higher in tube B than in tube A on each occasion (overall rate: 28% and 14% respectively). Estimate of between-occasions variation showed that B-TG levels were relatively stable for each subject between two occasions in each tube. It is concluded that the use of PGE1 decreases falsely high B-TG levels, but a single measurement of B-TG does not provide a reliable estimate of the true B-TG value in vivo.

Formation of prostacyclin-sensitive platelet aggregates in human whole blood in vitro. Part I. The occurrence of the phenomenon in healthy male volunteers.

When unstirred citrated blood of young males was left to stand at 21 degrees C, the number of free platelets decreased as measured with a whole blood platelet counter. This decrease indicated spontaneous platelet aggregation (SPA) since it was accompanied by the time-dependent increase in platelet micro-aggregates and plasma beta-thromboglobulin while lactate dehydrogenase level did not change significantly. Addition of PGI2 to whole blood increased free platelet count and decreased the number of platelet micro-aggregates. The de-aggregatory effect of PGI2 (measured as a percentage increase in free platelet number) was correlated with the initial amount of platelet micro-aggregates. Blood storage enhanced SPA and de-aggregatory effect of prostacyclin. Immediately after blood collection de-aggregatory effect of prostacyclin was absent. Our results favour an assumption that prostacyclin-sensitive platelet aggregates in blood of young volunteers are formed in vitro rather than in vivo.

Human neutrophil degranulation during extracorporeal circulation.

Cardiopulmonary bypass, especially when prolonged, may result in hemostatic failure and pulmonary dysfunction, which has been attributed to changes in platelets and leukocytes, respectively. It has been well documented that contact of blood with synthetic surfaces causes platelet activation. In this report, we explore mechanisms of the activation of neutrophils during simulated in vitro extracorporeal circulation and document the release of neutrophil lactoferrin and elastase during clinical cardiopulmonary bypass (CCB). Inhibition in the simulated circuit by prostaglandin E1 (PGE1) and lidocaine suggests different mechanisms for release of neutrophil-specific proteins. During CCB with a bubble oxygenator it was observed that platelet counts fell to 42% +/- 2% of baseline. In addition, beta-thromboglobulin antigen (beta TG), a platelet-specific, alpha-granule protein marker reflecting the release reaction, increased from 0.15 +/- 0.05 to 0.84 +/- 0.11 microgram/mL. Neutrophil counts decreased to 67% +/- 7% of prebypass levels but then gradually rose as bypass continued. Both lactoferrin, a neutrophil-specific granule marker, and neutrophil elastase, an azurophilic granule marker, increased in plasma threefold to 1.66 +/- 0.33 micrograms/mL and 1.65 +/- 0.68 microgram/mL, respectively, just before bypass was stopped. When fresh heparinized human blood was recirculated within an extracorporeal membrane oxygenator bypass circuit for 120 minutes, plasma beta-TG rose to 5.13 micrograms/mL, lactoferrin increased from 0.13 +/- 0.04 to 1.62 +/- 0.22 micrograms/mL, and neutrophil elastase rose from 0.05 +/- 0.02 to 1.86 +/- 0.41 micrograms/mL. At 120 minutes, lidocaine (100 mumol/L), which inhibits neutrophil activation, delayed release of lactoferrin (1.33 +/- 0.26 micrograms/mL) and markedly inhibited release of elastase (0.24 +/- 0.05 microgram/mL) but did not inhibit release of beta-TG antigen (5.66 micrograms/mL at 120 minutes). PGE1 (0.3 mumol/L) inhibited significantly the release of beta-TG (0.31 microgram/mL) and elastase (0.52 +/- 0.11 microgram/mL) and attenuated the release of lactoferrin (1.57 +/- 0.45 micrograms/mL).