[Haemostaseologic effects of fractionated snake venoms]. / Wirkungen von Schlangengiftfraktionen auf das Gerinnungssystem.

Comparative investigation concerning gelfiltration as well as haemostaseologic analysis of venoms and venom fractions of some snakes (elapidae and viperidae) have shown that in elapidae an inhibition of coagulation is dominant whilst in viperidae the stimulation of coagulation is of importance. Our investigations produce a basis to select substances for activation of coagulation and substances for inhibition of coagulation. Under pharmacological viewpoints the data may produce information to use snake fractions for anticoagulation or for procoagulant therapy in bleeding tendency.

[Mutation in factor II and factor V gene in patients with peripheral arterial occlusive disease]. / Chronisch-arterielle Verschlusskrankheit der Beine im Stadium IIb-IV: Molekulargenetische Diagnostik von Mutationen im Faktor-II-und Faktor-V-Gen.

Mutations in factor-V- and factor-II-genes are correlated with an increased risk for venous thrombosis according to the literature. The significance of the mutations in factor- II- and factor-V-genes for the development of the peripheral arterial occlusive disease is not known. Therefore, we investigated the presence of these mutations in 152 patients with documented peripheral arterial occlusive disease and 318 controls without peripheral arterial occlusive disease with polymerase chain reaction (PCR). There was no association between factor-II-mutation and peripheral arterial occlusive disease. The factor-V-mutation, however, was increased in patients with peripheral arterial occlusive disease double fold (12 positive cases in 318 controls, 12 positive cases in 152 patients with peripheral arterial occlusive disease). The significance level was reached (p = 0.05) in statistical analysis but the result did not fall below the significance level as necessary to reach statistical significance (odds ratio 2.19). Nevertheless, from these data we have to discuss a biological relevance of factor-V-mutation in the pathogenesis of peripheral arterial occlusive disease.

[Chromogenic substrate as antidote against the thrombin inhibitor Melagatran?]. / Chromogenes Substrat als Antidot gegen den Thrombininhibitor Melagatran?

It could be shown in vitro that a chromogenic substrate (Chromozym TH, Roche Mannheim) acts at least partially as antidote against the new thrombin inhibitor Melagatran (AstraZeneca, Mölndal, Sweden). It is discussed that this antidote effect of a chromogenic substrate might be due to a substrate competition of fibrinogen, thrombin inhibitor, and chromogenic substrate for thrombin. Further animal experiments will clarify whether this in vitro observation is of practical relevance in vivo, too.

Thrombin as a survival factor for cancer cells: thrombin activation in malignant effusions in vivo and inhibition of idarubicin-induced cell death in vitro.

OBJECTIVES: The aim of the experiments shown here, is to demonstrate exemplarily that thrombin can be a survival factor for malignant cells. METHODS: Activation of the coagulation system has been examined in patients with acute myeloid leukemia (AML) and non-Hodgkin lymphoma (NHL) before and after chemotherapy as well as in malignant effusions of heavily pretreated patients with solid tumors. Thrombin receptor expression (PAR-I) has been examined on HL-60 cells; the effect ofthrombin on the proliferation of the cells and inhibition of apoptosis induction by idarubicin has been shown. RESULTS: Using fibrinopeptide A as an indirect parameter for thrombin activation, we found elevated levels in patients with AML and NHL before and a significant 2-fold increase after chemotherapy (p < 0.02 for the AML group; p < 0.0006 for the NHL group). Apparently, this does not only affect patients with hematological diseases, but also with solid tumors. In order to find out if the tumor cells directly activate thrombin, we examined malignant effusions of patients with different solid tumors. Comparing prothrombin fragment 1 + 2 in ascites and pleural effusions with the patients' serum levels, we found it significantly increased in all cases (mean of 1.96 +/- 0.5 nmol/l in the serum vs. 12.1 +/- 3.6 nmol/l in effusions; p < 0.001). The majority of patients presented elevated serum levels. Additionally, we incubated HL-60 cells (human promyelocytic leukemia) with thrombin prior to treatment with idarubicin. Expression of thrombin receptor (PAR-1) could be verified by FACS-analysis using a monoclonal antibody. HL-60 cells responded with increased proliferation to thrombin exposure with concentrations between 0.3 and 3 U/ml. This effect could be abolished by the addition of hirudin, demonstrating thrombin specificity. In these concentrations, thrombin was able to abrogate the induction of apoptosis by idarubicin completely (p < 0.005). CONCLUSIONS: Here we give evidence for the role of thrombin as a resistance factor for tumor cells towards chemotherapy. In the light of the fact that thrombin is regularly activated in cancer patients, these findings indicate that thrombin is a clinically relevant cellular resistance factor. A number of pre-clinical and clinical studies imply that inhibition of the coagulation system, e.g. by low-molecular weight heparins or warfarin, increases the effect of chemotherapy.

[Effect of exercise tolerance test on hemostasis in patients with and without coronary heart disease]. / Der Einfluss der Fahrradergometrie auf die Hämostase bei Patienten mit und ohne koronare Herzerkrankung.

BACKGROUND: Physical exercise leads to an elevated coagulation activity with a possibly disturbed hemostatic balance. Therefore patients with coronary heart disease have a potentially increased risk of thromboembolic events after a bicycle exercise tolerance test, that is frequently performed for diagnostic reasons. PATIENTS AND METHODS: Patients with angiographically known coronary heart disease (Group I: n = 49; age 59 years; male = 42, female = 7) were investigated in comparison to a healthy cohort (Group 2: n = 51; age 53 years; male = 44, female = 7) to study the influence of a standardized exercise tolerance test on hemostatic variables. Blood samples were taken before and after exercise. RESULTS: No significant changes were found for any investigated parameter between both groups. However, 3 parameters did change significantly within the groups: factor VIII rose in Group 1 from 132 to 156% and in Group 2 from 106 to 136% and the von Willebrand factor rose in Group 1 from 230 to 249% and in Group 2 from 228 to 247%. An elevated fibrinolytic potential was found with an increase of plasminogen-alpha 2-antiplasmin in Group 1 from 251 to 401 micrograms/l and in Group 2 from 247 to 350 micrograms/l. CONCLUSION: The findings underline the clinical presumption that exercise tolerance test does not increase the risk for thromboembolic complications in patients with coronary heart disease in comparison to patients without coronary heart disease, as long as the exercise tolerance test is performed in a standardized way and under aerobe conditions.

Hyperthermic fibrinolysis with rt-PA: in vitro results.

PURPOSE: To investigate the influence of hyperthermia up to 45 degrees C on fibrinolysis with recombinant tissue-type plasminogen activator (rt-PA). METHODS: Standardized fibrin clots were incubated in a water bath for 5 hr with either rt-PA (test group) or 0.9% sodium chloride (control group) and blood plasma at temperatures of 30-45 degrees C. Concentrations of D-dimer and time to complete clot lysis were measured. RESULTS: The activity of fibrinolysis with rt-PA rose with increasing temperature: time to lysis approximately halved from 30 degrees C to 40 degrees C and the concentration of D-dimer tripled. In the control group clot size did not change. CONCLUSIONS: Activity of rt-PA-induced fibrinolysis rises distinctly with higher temperatures. Since even healthy subjects show a physiologic decline in body temperature in the extremities, in patients with occlusive arterial disease decreased activity of fibrinolysis with rt-PA can be expected. Controlled hyperthermia may improve fibrinolysis with rt-PA and should be investigated in vivo.

Anticoagulant fucoidan fractions from Fucus vesiculosus induce platelet activation in vitro.

Anticoagulant fucoidan fractions of different molecular weight and sulfate content were prepared and investigated for their effects on platelet function in vitro. The fucoidan fractions were incubated with human platelet rich plasma (PRP) at concentrations of 5, 10 and 50 micrograms/ml. Platelet activation was subsequently studied by a standard aggregation assay and flow cytometric determination of the activation dependent platelet-surface markers CD62p (P-selectin, GMP-140) and CD63 (GP53). All fucoidan fractions induced irreversible platelet aggregation in a dose-dependent manner. Comparing fractions of identical molecular weight (100 kDa) the low sulfate content fucoidan FF5 (S = 7.6%) exerted a significantly greater effect than the highly sulfated fucoidan FF7 (S = 10.2%) over the whole concentration range (n = 5, P < 0.05). Among fractions of identical sulfate content fucoidan-induced platelet aggregation was also found to depend on the molecular weight of the fucoidan. At concentrations of 10 and 50 micrograms/ml the high molecular weight fraction FF7/1 (150 kDa) showed a significantly greater effect than the 50 kDa fraction FF7/3 (24.8 +/- 6.7 vs. 7.0 +/- 3.5 and 54.6 +/- 13.5 vs. 15.0 +/- 9.0%, respectively; mean +/- SD, n = 5, P < 0.05). The molecular weight dependence of the fucoidan effect was also reflected by the flow cytometric data. Coincubation of FF7/1 and FF7/3 (10 micrograms/ml) with PRP increased the number of CD62p and CD63 positive platelets by 9.0 +/- 3.3 vs. 2 +/- 1.9 and 7.1 +/- 2.4 vs. 3.2 +/- 2.6% over control values, respectively (n = 5, P < 0.05). In conclusion, our results show that the low molecular weight fucoidan FF7/3 combines potent anticoagulant and fibrinolytic properties with only minor platelet activating effects and is therefore a suitable substance for further pharmacological studies.

[The thrombophilic status of patients with inflammatory diseases]. / Thrombophiler Status bei Patienten mit entzündlichen Erkrankungen.

BASIC PROBLEM AND OBJECTIVE OF STUDY: Inflammatory reactions are taken to be nonspecific defensive measures of the organism and are associated with complex changes at cellular and humoral level. Activation of blood coagulation plays an important part in this, especially as it is accompanied by an increased risk of thromboembolism. It was the aim of this investigation to assess this risk by measuring sensitive markers of coagulation activation. PATIENTS AND METHODS: Biochemical markers of coagulation activation (prothrombin-fragment F1 + 2 and thrombin-antithrombin III complex [TAT]) and fibrin formation (D-dimer) were measured in 130 patients (61 men, 69 women; mean age 56.9 [20-89] years). 44 had pneumonia, 44 bronchitis and 42 urinary tract infections. A healthy control group for comparison consisted of 11 men and 15 women (mean age 48.7 [23-79] years). RESULTS: F1 + 2, TAT and D-dimer were significantly increased, compared with the controls, in all three patient groups (P < 0.01). The greatest rises occurred in the patients with pneumonia: F1 + 2: median 1.2 vs 0.6 nmol/I, TAT: 6.2 vs 2.1 micrograms/l and D-dimer 2476 vs 223 ng/ml. CONCLUSION: These findings underline the importance of consistent thrombosis prophylaxis in patients with inflammatory disease, especially those at an increased risk.

[A modified technic for local rt-PA catheter-mediated lysis]. / Modifizierte Technik der lokalen rt-PA-Katheterlyse.

PURPOSE: To develop an economic and efficient concept for more time-saving local rt-PA thrombolysis therapy. METHOD: 40 patients with peripheral vascular occlusive disease stage IIb-III according to the Fontaine classification and with angiographically proven occluded segments of pelvic and lower limb arteries were treated by a modified concept of local rt-PA catheter thrombolysis. Via a thin guide wire the catheter for thrombolysis is slowly advanced through the thrombus without fluoroscopic control, outside the room in which angiography is performed. In 24 cases a short-term lysis and in 16 cases a long-term lysis was carried out. RESULTS: The initial success rate was 75%, the patency rate in six months' follow-up was 66.7%. The ankle-brachial index decreased from 0.4 +/- 0.3 to 0.8 +/- 0.2 on the average. There were no relevant clinical complications. The average occupancy time of the angiography room or table was 60 +/- 52 min, the average time of fluoroscopy was 17 +/- 13 min. CONCLUSION: In modified local rt-PA thrombolysis, short-term lysis and long-term lysis were mostly performed outside the angiography room, so that the exposure to radiation and there fore the radiation dose were reduced for both the patient and the attending staff. The angiography room is thus available for other patients and can therefore be used more efficiently.

The use of prothrombin fragment F1+2 to monitor the effect of oral anticoagulation.

In comparison to a control group of 47 healthy, non-anticoagulated persons, 164 patients under stable oral anticoagulation therapy showed significant reduction in the plasma level of the prothrombin fragment F1+2 (p < 0.0005). Even with low intensity anticoagulation (INR < 2.0), F1+2 levels were reduced to within the normal range (0.32 - 1.2 nM/l) in all patients. The reduction in the plasma level of prothrombin fragment F1+2 is directly dependent on the intensity of oral anticoagulation therapy and provides a means of monitoring the anticoagulation effect achieved. Clinical studies are required to assess the practicality of using F1+2 for monitoring anticoagulation and in particular to assess the usefulness of low levels in predicting bleeding risk and high levels in predicting thrombotic risk during treatment.

[Oral contraception in protein C deficiency]. / Orale Kontrazeption bei Protein-C-Mangel.

PIP: The question was posed whether a young patient with familial protein C deficiency with previous thromboembolic complication and under prophylactic oral anticoagulation therapy could be recommended an oral contraceptive. Basically, familial protein C deficiency per se does not require oral anticoagulation therapy. However, when risk factors such as confinement to bed, operation, contraception, and childbirth enter, anticoagulation is indicated. In the event severe thromboembolism is in the anamnesis, it has to be resolved whether oral contraceptives are necessary or other contraception is possible. Usually, oral anticoagulants are halted 6 months or 1 year after a leg vein thrombosis as long as no severe venous thrombotic residues are at hand. However, OCs are continued, which could result in a conflict situation. The analysis of prothrombin fragments F1+2 can ascertain whether hypercoagulability is present independent of or in connection with anticoagulation, which can be read on a pathological rise of the prothrombin fragments F1+2 in the plasma. If such a finding is obtained, the taking of OCs would be critically dealt with despite anticoagulation. If in the phase of discontinuation of oral anticoagulation the prothrombin fragment F1+2 is lowered or is normal, there is no danger of thrombosis, thus the oral anticoagulation can be halted. However, if the prothrombin fragment F1+2 rises pronouncedly, then there is an urgent need for renewed introduction of oral anticoagulation, which cannot be stopped in such a patient. These aspects have to be considered also in relation to the question of compatibility of OCs with oral anticoagulation.^ieng