Evaluation of back pain secondary to spinal epidural hematoma associated with aspirin intake and a partial platelet glycoprotein Ia/IIa deficiency.

We report a case of spontaneous spinal epidural hemorrhage with three unusual features: (1) the hemorrhage was associated with aspirin ingestion and a reduced level of platelet glycoprotein Ia/IIa; (2) the patient presented with typical severe back pain but without neurologic dysfunction; and (3) the patient initially recovered without surgical decompression but suffered from recurrent epidural hematoma.

Fibrinogen Zurich I: impaired release of fibrinopeptide A.

Fibrinogen Zurich I is characterized by an abnormal fibrin monomer polymerization. It consists of two fractions of molecules, one with a normal aggregation and one not aggregating at all and interfering with the aggregation of the normal population. Using a radioimmunoassay for fibrinopeptide A, only approximately half of the expected fibrinopeptide A could be recovered after thrombin or Defibrase proteolysis. The defective fibrinopeptide A release could be confirmed by measurement of the N-terminal Gly/Tyr ratio. It is likely that the abnormal fibrin monomer aggregation of the abnormal fraction of fibrinogen Zurich I is due to the defective fibrinopeptide A release of this fraction.

Sialic acid dependent polypeptide chain heterogeneity of human fibrinogen demonstrated by two-dimensional electrophoresis.

Human fibrinogen was compared with asialofibrinogen by two-dimensional electrophoresis to evaluate the contribution of sialic acid to the heterogeneity of the gamma- and B beta-polypeptide chains. Reduced fibrinogen showed three major variants for both the gamma- and B beta-chains. In addition two minor gamma-bands with a more acidic isoelectric point than the normal gamma-chains were observed. Electrophoresis in the second dimension (SDS) suggests that these most acidic bands are gamma-chain-variants with a higher molecular weight. In asialofibrinogen only two predominant variants with more alkaline isoelectric points were present in each chain type. It is concluded that enzymatic removal of sialic acid partially reduced the heterogeneity of the gamma- and B beta-polypeptide chains of human fibrinogen, but additional sources producing charge heterogeneity must be sought.

Blood coagulation after long distance running: antithrombin III prevents fibrin formation.

Physical exercise causes shortening of activated partial thromboplastin time (aPTT) and euglobulin clot lysis time. To investigate whether this activation of coagulation and fibrinolysis leads to in vivo thrombin or plasmin action after long distance running, 19 well trained male runners (36-65 years) were examined 5 to 53 min after termination of a 100 km race and 5 days later after at least 1 day without physical exercise. Compared to the control examination aPTT was decreased (30.2 +/- 2.8 vs 35.3 +/- 3.0 sec) and the following parameters were increased after the race: beta thromboglobulin (40 +/- 16 vs 23 +/- 7 ng/ml), thrombin-antithrombin III (TAT) complexes (5.5 +/- 3.4 vs 2.3 +/- 0.7 microgram/l), the fibrin(ogen) degradation products fragment E (57 +/- 15 vs 35 +/- 7 ng/ml) and B beta 15-42 (8.5 +/- 2.5 vs 6.5 +/- 2.5 ng/ml) (all p values less than 0.001). Platelet count, platelet factor 4, fibrinoepetide A (FPA) and haematocrit did not change significantly. Increased TAT complexes and unchanged FPA suggest that the generated thrombin was fully inactivated by antithrombin III (AT III) and did therefore not give rise to fibrin formation. The small increase of fibrin(ogen) degradation products indicates a minor in vivo activity of the fibrinolytic system. This investigation demonstrates the importance of AT III in the regulation of haemostasis in activated blood coagulation.

Prothrombin fragment 1 + 2, thrombin-antithrombin III-complexes and fibrinopeptide A in spontaneously clotting whole blood in vitro. Effects of heparin addition and antithrombin III deficiency.

Previous in vitro studies using spontaneously clotting whole blood revealed thrombin formation and high fibrinopeptide A (FPA) concentrations measured during incubation time. This occurred in spite of normal concentrations of thrombin antagonists present in the blood of the healthy subjects examined. However, there are several reports showing that in vivo increased thrombin-antithrombin III-complex (TAT) concentrations and relatively low FPA concentrations may occur e.g. in patients with (pre)thrombotic disorders. These in vivo findings indicate more effective thrombin inhibition by antithrombin III, with almost no fibrin formation. To find an explanation for the differences observed in vitro and in vivo, we extended the in vitro studies by measuring concentrations of prothrombin fragment 1 + 2 (F1 + 2), TAT and FPA at several time points until 30 min. Our goal was to test whether thrombin at least initially is neutralized by antithrombin III, resulting in a lack of fibrin formation, either in the absence or in the presence of heparin (0.2 and 0.5 U/ml whole blood, respectively). In the absence of heparin a simultaneous increase in the concentrations of F1 + 2, TAT and FPA was observed. Thrombin was only partially neutralized by antithrombin III and large amounts of fibrin were formed. The addition of heparin virtually suppressed thrombin formation since the F1 + 2 concentration remained low. Moreover, the small amounts of thrombin formed were neutralized by antithrombin III to a greater extent than in the absence of heparin.(ABSTRACT TRUNCATED AT 250 WORDS)

Excessive deposition of fibrin, platelets and platelet thrombi on vascular subendothelium during contraceptive drug treatment.

We investigated the effect of oral contraceptives on thrombogenesis induced by subendothelium of rabbit aorta (SE), exposed to flowing non-anticoagulated blood in an annular flow chamber. Six healthy women on sequential contraceptive drugs (0.05 mg aethinylestradiol/day, 0.125 mg desogestrel/day) were compared with 6 women without hormonal contraception and 6 men. On contraceptive drug treatment, blood values were significantly increased for fibrinogen (2.6 +/- 0.2 vs 1.9 +/- 0.1 g/l) and fibrinopeptide A (3.9 +/- 0.9 vs 0.9 +/- 0.1 ng/ml), whereas antithrombin III was decreased (81 +/- 4 vs 97 +/- 6%). Fibrin deposition on vascular subendothelium was more than four-fold increased when measured morphologically (63.4 +/- 2.5 vs 14.6 +/- 6.8% coverage of SE surface with fibrin) as well as immunologically (29.3 +/- 2.2 vs 4.5 +/- 1.9 micrograms fibrin/cm2 of SE). Thrombus volumes were more than two-fold increased in women with contraceptives (9.0 +/- 1.4 vs 3.7 +/- 1.0 micron 3/micron 2). Our study shows that during contraceptive drug treatment the exposure of flowing blood to vascular subendothelium leads to excessive deposition of fibrin and platelet thrombi. Measurement of blood interactions with subendothelium might be of predictive value in hypercoagulable states such as contraceptive treatment.

Relationship between fibrinopeptide A and fibrinogen/fibrin fragment E in thromboembolism, DIC and various non-thromboembolic diseases.

Increased fibrinopeptide A (FPA) levels have been reported in various non-thrombotic disorders, including cancer, acute myocardial infarction, liver cirrhosis and collagen vascular diseases. To investigate the significance of these findings, the present study combined the radioimmunoassay of FPA with that of fibrinogen/fibrin degradation fragment E (FgE) in the aforementioned disorders and compared the results with those observed in healthy subjects as well as in patients with thromboembolism and overt disseminated intravascular coagulation (DIC). Mean FPA and FgE in malignancy were 6.3 and 305 ng/ml, in myocardial infarction 5.6 and 98 ng/ml, in liver cirrhosis 2.7 and 132 ng/ml and in collagen vascular diseases 5.6 and 142 ng/ml. All these values were significantly higher than in healthy controls (mean FPA 1.6 ng/ml, mean FgE 49 ng/ml) but significantly lower than in thromboembolism (mean FPA 10.7 ng/ml, mean FgE 639 ng/ml). and DIC (mean FPA 22.0 ng/ml, mean FgE 1041 ng/ml). The overall correlation between FPA and FgE was highly significant. However, different disorders showed peculiar patterns in FPA, FgE and fibrinogen levels. In malignancy, a definite increase of FPA, FgE and plasma fibrinogen levels was observed. This finding probably indicates a compensated state of (intra- or extravascular) fibrin formation and lysis. Acute myocardial infarction was characterized by a high FPA to FgE ratio, which is interpreted to reflect acute thrombin generation and fibrin formation. FPA in cirrhosis was only marginally elevated with most single values within the normal range, indicating that intravascular coagulation was infrequent and unimportant in quantitative terms.

Isolation of human fibrinogen and its derivatives by affinity chromatography on Gly-Pro-Arg-Pro-Lys-Fractogel.

With an immobilized synthetic pentapeptide GlyProArgProLys comprising the N-terminal sequence GlyProArg of the alpha-chain of fibrin, a new affinity method for the quantitative isolation of fibrinogen out of anticoagulated plasma was developed. The method proved to be superior to all known isolation methods in respect to ease of use and yield, since fibrinogen could be isolated in one step out of plasma with a recovery of more than 95% when compared to the immunologically measurable amounts of fibrinogen. Moreover the amounts of contaminating proteins such as fibronectin, factor XIII or plasminogen were negligible and the purity of the isolated fibrinogen was higher than 95% as measured by polyacrylamide gel electrophoresis. The clottability was 90% and more. Another advantage of this affinity purification method is the possibility to isolate fibrinogen quantitatively out of small plasma samples (less than 5 ml). Further, abnormal fibrinogen molecules, provided their complementary binding site for GlyProArg is preserved, may also be quantitatively isolated independent of any solubility differences as compared to normal fibrinogen. In addition fibrin(ogen) fragments originating from plasmic digestion can be separated on the basis of their affinity to GlyProArg. The described affinity gel can be used more than 50 times without any loss of capacity.

Whole blood clot lysis: enhanced by exposure to autologous but not to homologous plasma.

A new phenomenon is described: Whole blood clots lyse faster in the plasma of the same donor than in another donor's plasma. We have confirmed this finding in 68 healthy volunteers by a standardized, pair-wise analysis and have found a mean difference in clot weights of 8.8 +/- 0.99% (SEM, p < 0.0001) after 6 h of urokinase-induced (200 U/ml) clot lysis. No difference was found in a group of 7 pairs of identical twins. Further analysis revealed that increasing concentrations of platelets in the plasma reduced the difference significantly but did not abolish it. A 1:1 mixture of autologous with homologous plasma reduced the autologous advantage by almost 50%, thus making an inhibitor unlikely. The absence of cellular components in clots of platelet-poor plasma resulted in the loss of the advantage after 2 h of lysis, but not in the early phase. We conclude that there is a clear advantage of autologous over homologous clot lysis. Potential mechanisms are discussed and include an increased affinity of enzymes for their substrates in a given individual.

The suppression of the coagulation of nonanticoagulated whole blood in vitro by human umbilical endothelial cells cultivated on microcarriers is not dependent on protein C activation.

Human umbilical vein endothelial cells (HUVEC) were cultivated on globular microcarriers in order to improve the endothelial cell surface to blood-volume ratio over the conventional flat bed cultures. HUVEC-beads were tested for their modulation of blood coagulation using a combination of two steps: HUVEC-beads were added into the syringe used for the venipuncture, in order to achieve immediate contact between cells and blood, and no anticoagulant was used during the incubation time of HUVEC-beads with whole blood. The coagulation initiation produced by venipuncture was almost completely suppressed as judged by thrombin measurements over a period of 60 min. The activated partial thromboplastin time showed a prolongation by a factor > 3. Direct measurements of activated protein C (APC) were negative. Moreover, inhibition of APC-generation with a monoclonal anti-human protein C antibody did not affect the anticoagulant properties of endothelial cells. Therefore the anticoagulant properties exerted by HUVEC-beads are not dependent on APC.

Fibrinogen: agreement of experimental and calculated hydrodynamic data with electron-microscopic models.

There is no general agreement on the size and shape of the fibrinogen molecule. We have studied the diffusion of the fibrinogen in solution by means of dynamic light scattering, nanosecond fluorescence depolarization and analytical ultracentrifugation. The results obtained under physiological concentration, pH an ionic strength are DT = 2.0 X 10(7) cm2sec-1, DR perpendicular = 40'000 sec 1. Nanosecond fluorescence depolarization yielded DR parallel = 1.6 x 10(6) sec-1. Tentatively this value is interpreted as DR parallel, namely rotational diffusion about the major axis of the molecule. The sedimentation coefficient is 8.1 S. The hydrodynamic parameters derived from our measurements were compared with those calculated on the basis of the models proposed by Hall and Slayter, Hudry-Clergeon and Marguerie et al., Bachmann and Lederer, and Köppel. The agreement is poor even if the degree of hydration is varied within wide limits. However, satisfactory agreement can be achieved by assuming a flexible molecule of about 900 A length corresponding to two end-to-end bound trinodular structures of the Hall and Slayter type, with a nodule diameter of about 45 A. Experimental evidence indicates that the discrepancies between the different models might be due to different techniques of sample preparation leading to different conformations of the molecule.

The influence of extracorporeal circulation on erythrocytes and flow properties of blood.

The influence of extracorporeal circulation on red blood cells and flow properties of blood was studied in 10 patients undergoing aorta-coronary bypass grafting. Blood samples were drawn on admission, under general anesthesia before the operation, during extracorporeal circulation, immediately after extracorporeal circulation, and 24 hours after extracorporeal circulation. Echinocytes were found during and shortly after extracorporeal circulation, but disappeared within 24 hours. Washing the cells in buffer restored the normal discocytic shape, which indicated that a plasma factor was responsible. Red cell membrane lipids were not affected. Analysis of the membrane proteins revealed a decrease of ankyrin after extracorporeal circulation, which was prevented by protease inhibitors during preparation. This suggests an increased proteolytic activity of the plasma after extracorporeal circulation. Red cell deformability was not altered. Plasma viscosity and hematocrit were markedly reduced by hemodilution with the priming solution. Their low levels resulted in a low blood viscosity during extracorporeal circulation, which was even lower at 26 degrees C than before or after the operation at 37 degrees C. We conclude that the red cell is affected by extracorporeal circulation. The flow properties of blood, however, are not impaired, but are improved by hemodilution.

Hypofibrinogenaemia in metastatic carcinoma of the prostate: suppression of systemic fibrinolysis by haparin.

In a patient with generalized prostatic carcinoma and hypofibrinogenaemia heparin infusion on four occasions abolished systemic fibrinolysis as determined by euglobulin clot lysis time, bovine fibrin plate assay, thromboelastography, and immunoelectrophoretic demonstration of fibrinogen split products. The pathogenesis of hypofibrinogenaemia in cases of prostatic carcinoma and the possibility of a direct heparin effect on fibrinolysis are discussed. Despite the lack of histological evidence for intravascular coagulation, the findings are considered additional evidence for the view that fibrinolysis in this syndrome may be secondary to intravascular coagulation.

Clotting factor v activity in plasma cryoproteins.

Plasma cryoprecipitates containing considerable quantities of clotting factor V were observed in five patients. The unusual factor V activity was not associated with significant amounts of cryofibrinogen. Clinically, the symptoms were not distinguishable from those of other cryopathies. The abnormality did not lead to a deficiency of factor V in the circulating blood nor to a haemorrhagic diathesis.

Plasma volume and proteins in voluntary hyperventilation.

In a controlled study the changes of the plasma volume and plasma proteins induced by voluntary hyperventilation (HV) were investigated in nine splenectomized volunteers. The plasma volume changes were calculated from the changes of the hemoglobin and hematocrit. After 20 min of HV in supine position, which lead to a decrease of the venous CO2 partial pressure by 19 Torr and to an increase of plasma epinephrine and norepinephrine levels, the plasma volume was reduced by 12.9%. The intravascular masses of total protein, albumin, and several other proteins decreased during HV but a similar decrease of these proteins was also observed during the control study (C), i.e., rest in supine position without HV. The differences between changes during HV and C were not significant, indicating that the loss of protein was not due to HV. It is concluded that acute HV leads to a rapidly reversible loss of a virtually protein-free solution from the vascular space. The red cell compartment participated in fluid shifts in that the mean red cell volume decreased by 2.2% (P less than 0.02 compared with C). Comparison with earlier work shows that addition of erythrocytes from the normal spleen does not play a part in the HV-induced increase of hemoglobin and hematocrit.

Increased thrombin-antithrombin III complexes after 1 h of physical exercise.

Concentrations of thrombin-antithrombin III (TAT) complexes in plasma were previously reported to be increased after a 100-km run, while fibrinopeptide A (FPA) concentration remained unchanged. Thus, antithrombin III appears to neutralize thrombin generated during running and prevents fibrin formation. To determine the clinical relevance of these findings, we compared the effects of exhaustive running (1 h, n = 10) on the plasma concentrations of prothrombin fragments F1 and F2, TAT, FPA, and beta-thromboglobulin with the effects of recreational jogging (1 h, n = 10) and exhaustive bicycling on an ergometer (1 h, n = 8). Prothrombin fragments F1 and F2 and TAT concentrations increased significantly in each group. The most significant increase in TAT concentration was measured in the running group (from 1.72 +/- 0.49 to 3.61 +/- 1.03 ng/ml, P < 0.001). The best correlation was found between the postexercise TAT and lactate concentrations (r = 0.62, n = 28, P < 0.001). Mean FPA concentrations after exercise did not exceed normal values in any of the three groups analyzed. An increase in beta-thromboglobulin concentration was measured in the running and in the cycling group. Thus, thrombin is formed, in particular, when associated with anaerobic metabolism, and platelets are activated during high-intensity exercise.

Hyperventilation-induced changes of the blood picture.

In a controlled study of 11 male volunteers the following changes (means +/- SD) were observed in venous blood during (D) and 75 min after (A) a period of 20 min of voluntary hyperventilation in comparison with before (B) hyperventilation (P values referring to the difference between D and B) erythrocyte count 5.18 +/- 0.17 X 10(6) (B), 5.70 +/- 0.21 X 10(6) (D) (P less than 0.001), and 5.18 +/- 0.16 X 10(6)/microliter (A); hemoglobin 15.7 +/- 0.6 (B), 17.2 +/- 0.7 (D) (P less than 0.001), and 15.8 +/- 0.6 g/dl (A); centrifuged hematocrit 46.6 +/- 1.0 (B), 50.4 +/- 1.7 (D) (P less than 0.001), and 47.0 +/- 1.8% (A). The platelets increased from 159 +/- 30 X 10(3) (B) to 205 +/- 40 X 10(3) (D) (P less than 0.001) and returned to 157 +/- 26 X 10(3)/microliter (A). The leukocytes (WBC) were 4,210 +/- 630 (B), 6,220 +/- 1,660 (D) (P less than 0.001), and 6,190 +/- 1,870/microliter (A) (P less than 0.002, as compared with B). The rise of WBC during hyperventilation was mainly due to a 83% increase of lymphocytes, whereas a 93% increase of neutrophil leukocytes accounted for the increased WBC 75 min posthyperventilation. The increase of the ratio of band forms to segmented neutrophils from 9 (B) to 19% (A) (P less than 0.01) indicates that band forms were released from the bone marrow. The results show that WBC and platelets can be mobilized by hyperventilation by as yet unidentified mechanisms.

Coagulation and fibrinolysis in acute mountain sickness and beginning pulmonary edema.

To examine whether intravascular coagulation and/or decreased fibrinolysis precedes high-altitude pulmonary edema (HAPE) we examined 25 male mountaineers (median age 40 yr) at low altitude (550 m) and after 6, 18, and 42 h at an altitude of 4,559 m, which was climbed in 24 h. In 14 subjects, 2 of whom showed radiological evidence of HAPE after 42 h, symptoms of acute mountain sickness (AMS) were mild or absent. Eleven subjects suffered from AMS, six of whom developed radiologically documented HAPE after 18 or 42 h. In the absence of AMS there were no significant changes at high altitude, with the exception of a decrease in bleeding time from 246 +/- 18 to 212 +/- 13 (SE) (P less than 0.05). In AMS, partial thromboplastine time decreased from 34.2 +/- 0.8 to 31.1 +/- 0.5 s (P less than 0.001) and factor VIII procoagulant activity and von Willebrand factor antigen were increased by 57 +/- 12 and 70 +/- 13%, respectively (P less than 0.001), whereas there were no significant changes in beta-thromboglobulin (BTG), fibrinopeptide A (FPA), and fibrin fragment B beta 15-42. In subjects with HAPE, BTG, FPA, and B beta 15-42 were normal before and in beginning HAPE. Preceding HAPE, euglobulin clot lysis time declined at high compared with low altitude from 289 +/- 48 to 201 +/- 42 min without venous occlusion (VO) and from 107 +/- 36 to 86 +/- 31 min after VO (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Contact phase of blood coagulation is not activated in edema of high altitude.

To examine whether bradykinin generated by the activation of the contact phase of blood coagulation is involved in the pathogenesis of edema occurring after acute exposure to high altitude, 15 mountaineers were examined at 490 m and 1, 3, and 5 days after arrival at 4,559 m. The clotting activity levels of factor XII, factor XI, plasma prekallikrein, and high-molecular-weight kininogen (HMWK) were measured, and plasma kallikrein-induced proteolytic cleavage of HMWK was assessed by ligand blotting by use of radiolabeled factor XI. After an ascent on foot from 1,170 to 4,559 m in 3 days, three subjects developed high-altitude pulmonary edema, and four subjects presented facial edema. There was no evidence for activation of the contact system in any subject as demonstrated by the lack of proteolytic cleavage of HMWK at high altitude. The absence of contact system activation was further supported by stable plasma levels of the individual factors of contact activation. Therefore, we conclude that bradykinin generated by plasma kallikrein-induced cleavage of HMWK is not involved in the pathogenesis of edema due to acute exposure to high altitude.

Erythrocyte adenosine triphosphate depletion during voluntary hyperventilation.

Chronic hypophosphatemia in humans is associated with a slow depletion of adenosine triphosphate (ATP) and 2,3-diphosphoglycerate (2,3-DPG) in erythrocytes, combined with shape alteration, impaired deformability, and viability of the cells. Likewise, incubation of erythrocytes in alkaline solution is associated with ATP depletion. Since in hyperventilation both hypophosphatemia and alkalosis are present, we have investigated red cell organic phosphates, shape, deformability, and osmotic fragility before, during, and after 20 min of voluntary hyperventilation. On the average, red cell ATP decreased by 42%, the blood pH increased by 0.2 units, and plasma inorganic phosphorus decreased by 46% compared with the initial values. Red cell 2,3-DPG, shape, deformability, and osmotic fragility remained unchanged. After the end of hyperventilation ATP increased rapidly to control values in parallel with the normalization of the blood pH, whereas inorganic plasma phosphorus remained at the low level observed during hyperventilation. It is concluded that the combined effects of hypophosphatemia and alkalosis in acute hyperventilation lead to an isolated fall of red cell ATP, which occurs as rapid as after total inhibition of red cell glycolysis in vitro.