Platelet function (as production of thromboxane B2 by platelets stimulated with collagen, and plasma beta-thromboglobulin) and thrombin activity (as plasma fibrinopeptide A) were investigated in eight young (mean age 27 +/- 3 SE years) male patients in which type 1 diabetes mellitus had been diagnosed 2 to 6 months previously. They were all in excellent stable metabolic control (mean HbA1c 5.6 +/- 0.4 SE %) and free from any complications. The haemostatic variables were assessed at rest and after cycloergometric exercise to exhaustion. When compared to age- and sex-matched healthy controls, patients showed higher beta-thromboglobulin, fibrinopeptide A and thromboxane B2 at rest. After exercise, plasma beta-TG increased only in the controls. Platelet and thrombin activation are present in the very early stages of type 1 diabetes mellitus, in the absence of any complications.

Two automatic coagulometers--ACL 810 (Instrumentation Laboratory), a laser-nephelometric centrifugal analyzer, and KoaguLab 40 A (Ortho Diagnostics), an optical automatic coagulometer--were compared with the manual tilt-tube method for the performance of prothrombin time (PT). Seven ISI- (International Sensitivity Index) calibrated commercial thromboplastin reagents were used for duplicate determinations in 30 normal subjects, 30 patients with liver disease, and 30 patients receiving stabilized oral anticoagulation. Clotting times were longer with the manual method than with ACL 810 and, to a lesser extent, with KoaguLab 40 A. Average imprecision of duplicate determinations (CV) was less than 1% with ACL 810; KoaguLab 40 A and the manual method had similarly higher imprecisions (2.8% and 2.7%). Differences in origin and slope of the regression curves of clotting times obtained with the coagulometers over the tilt-tube method were observed with all the reagents tested. Transformation of clotting times to PT ratios did not eliminate the bias resulting from the different clot-detection methods. A higher percentage of patients with liver disease had abnormal PT ratios when their plasma was tested with the coagulometers than with the manual method. Transformation of PT ratios to International Normalized Ratios effectively eliminated the bias resulting from the different thromboplastin reagents but had no effect on the bias resulting from the different clot-detection methods. A significant proportion of patients appeared excessively anticoagulated (INR greater than 4.5) with the coagulometers but not with the manual method. These results highlight the need for standardization of both instrumentations and reagents to improve monitoring of oral anticoagulant treatment.

Iloprost is a chemically stable analogue of prostacyclin, with similar vasodilator and anti-platelet actions. Platelet sensitivity to the inhibitory action of Iloprost has been tested in vitro. Platelet-rich plasma from six healthy subjects and from six patients with type 1 diabetes mellitus was incubated with different concentrations of Iloprost, and then stimulated with ADP (at threshold aggregating concentration) and collagen 4 micrograms/ml. The half-maximal inhibitory concentration (IC50) of Iloprost was calculated and no differences were found between patients and controls. The results of this study suggest that diabetic patients without complications do not differ from healthy subjects in their platelet sensitivity to Iloprost.

Habitual smoking is one of the best established risk factors for cardiovascular disease. The pathogenesis of smoke-induced damage is not so well clarified, but it probably includes--among some other aspects--an activation of the hemostatic system. Recently it has been shown that smoking a single cigarette can significantly decrease the coronary blood flow in coronary patients as well as in normal subjects. We tested the hypothesis that the acute effects of smoke are mediated by the hemostatic system. Seven healthy male volunteers, aged 20-40 years (mean 32 +/- 6 years), entered the study. All were habitual smokers, but had abstained from smoking in the 12 hours preceding the test. After lying in absolute rest for about 30 minutes, each subject smoked a cigarette containing 1.2 mg of nicotine. Immediately before and after smoking, blood was drawn by clear venipuncture for the evaluation of the following hemostatic variables: collagen-induced platelet aggregation by the method of Born; thromboxane B2 (TxB2) production by platelets stimulated with collagen, radioimmunoassay (RIA); plasma beta thromboglobulin (TG) (RIA); plasma fibrinopeptide A (FPA) (RIA); plasma fibrinolytic activity in the euglobulin fraction (NEF) (fibrin plate method). The following results, respectively before and after smoking, were observed: collagen-induced platelet aggregation 55 +/- 3 vs. 57 +/- 6%; TxB2 100.5 +/- 5.9 vs. 90.3 +/- 9.0 ng/10(8) platelets; plasma beta-TG 20.8 +/- 2.2 vs. 19.2 +/- 2.3 ng/ml; plasma FPA 2.3 +/- 0.3 vs. 2.2 +/- 0.1 ng/ml; NEF, lysis diameter 16.8 +/- 1.6 vs. 16.7 +/- 1.7 mm; NEF + C1 inhibitor lysis diameter 8.8 +/- 0.7 vs. 9.1 +/- 0.7 mm.(ABSTRACT TRUNCATED AT 250 WORDS)

It is current clinical practice to monitor heparin therapy by maintainig the activated partial thromboplastin time (APTT) at 1.5-2.5 × control normal pool. There is however controversy regarding the choice of reagents with respect to their heparin sensitivity. Choice of instrumentation is also known to affect the results. We have compared two automatic coagulometers, the ACL (Instrumentation Laboratory), a laser-nephelometric centrifugal analyzer and the KOAGULAB 40A (Ortho Diagnostics), an optical automatic coagulometer, with the tilt tube technique for the performance of APTT. Seven commercial reagents have been used for APTT replicate determinations in 30 normals, 30 liver disease patients and 30 patients on heparin therapy (20-40,000 IU daily). The overall observed imprecision (C. V.)was 1.8% for ACL, 3.0% for KOAGULAB 40A and 2.3% for tilt tube technique. The F test for the two-way interaction of ratios was statistically significant (p<0.001) for the large majority of reagent/ technique combinations in normals, liver disease and heparin treated patients. The percentage of patients adequately, inadequately and excessively heparinazed obtained with the two automatic instruments and with the tilt tube technique were not sig nificantly different when using the same reagent. However, major differences were observed when comparing the different reagents. For instance, inadequately heparinized patients were 90% according to one reagent (Cephotest, Nyegaard) as opposed to 17% according to another reagent (APTT, Instrumentation Laboratory). These results stress the need for a standardized APTT reagent to provide effective laboratory monitoring of heparin treatment. .

The prothrombin time (PT) is the most widely used assay to monitor oral anticoagulation (OA). Although it has been established that both thromboplastin and instrumentation significantly affect the results, major standardization attempts have been devoted to the calibration of reagents rather than of instruments. To provide safe laboratory monitoring of OA, an International Sen sitivity Index (ISI) for thromboplastin has been introduced. We have compared two automatic coagulometers, the ACL (Instrumentation Laboratory), a laser-nephelometer centrifugal analyzer which measures the intensity of the light scattered by a plasma sample before, during and after clot formation and the KOAGULAB 40A (Ortho Diagnostics), an optical automatic coagulometer, with the till tube technique for the performance ofPT. Five calibrated commercial thromboplastins have been used for replicate determinations in 30 normals, 30 liver disease patients and 30 patients on stabilized OA. The overall observed imprecision (C.V.) was 1.1% for ACL, 2.9% for the KOAGULAB 40A and 3.0% for the till tube technique. The F test for the two-way interaction of ratios was statistically significant (p< O.OOl) for the large majority of reagent/technique combinations in normals and in liver disease patients. Internatio nal normalized ratios were also significantly different (p< O.OOl) in most instances in patients on OA. Inadequate anticoagulation (INR<2.0)was observed in 18% of patients with the ACL , in 31% with the KOAGULAB 40A and in 33% with the till tube technique. Excessive anticoagulation (INR> 4.5) was observed in 19% of the patients with the ACL, in 7% with KOAGULAB 40A and in 3% with the till tube tech nique. These data highlight the need for standardization of both instrumentations and reagents to improve monitoring of OA.

Protein S (PS), the cofactor of activated protein C (PC), circulates in plasma as free PS (active) and in complex with C4b-binding protein (inactive). We have followed the changes of total PS, free PS, PS activity and C4b-binding protein in 5 adult patients with acute lymphoblastic leukemia treated with 20,000 IU/sqm L-asparaginase (L-ASE) administered three times weekly for two weeks (6 doses) and compared them to the changes of PC antigen and activity, antithrombin III and fibrinogen. The table shows the mean values of these parameters (% of pretreatment values) observed after three doses of L-ASE (A), after completion of treat ment (B) and one week after the end of treatment (C). These data suggest that at variance with PC and similar to fibri nogen and antithrombin III, total PS antigen is still reduced af ter one week from the end of L-ASE treatment. The reduction of total PS reflects that of free PS, as predicted by mass action. On the contrary, PS anticoagulant activity is decreased to a higher extent than total PS antigen. Acquired PS deficiency might play a contributory role to the development of thrombotic comply cations in patients receiving L-ASE. Supported in part by CNR grant n 85.02200.44

Platelet function and thrombin activity were investigated in 12 hospitalized patients (7 men and 5 women, mean age 53 years) who had had transient cerebral ischemic attacks in the previous 2-12 weeks. Each patient was given an extensive clinical and instrumental evaluation, including Doppler sonography of the cervical and lower limb vessels, cerebral angiography, and head computed tomography scan, after which relevant atherosclerotic disease was excluded. The controls consisted of 12 subjects hospitalized for nonvascular neurologic problems and matched for age, sex, and risk factors to the transient ischemic attack patients. Collagen-induced platelet thromboxane B2 production, plasma beta-thromboglobulin, and fibrinopeptide A were significantly higher in the patients than the controls. Platelet aggregability by collagen was the same in the 2 groups. Platelet hyperfunction and enhanced thrombin activity are present in patients some weeks after the acute episode, suggesting that the hemostatic system has a primary pathogenetic role.