Tissue-type plasminogen activator (t-PA) and dilute blood clot lysis time in nephrotic patients

When compared to normal weight normolipidemic control subjects, dilute blood clot lysis time was found to be obviously (p less than 0.001) prolonged in hypertriglyceridemic patients without proteinuria and slightly (p less than 0.05) accelerated in hyperlipidemic nephrotic patients in spite of their very high levels of plasma fibrinogen. As a result the ratio plasma fibrinogen (mg/dl) per clot lysis time (minutes) was 1.241 +/- 0.08 (X +/- SEM) in control subjects, 0.574 +/- 0.07 in hypertriglyceridemic patients and 2.69 +/- 0.172 in nephrotic patients. This finding suggesting that a larger amount of fibrin is rather readily dispersed from dilute blood clots of nephrotic patients was associated with higher levels of plasma t-PA:Ag (9.45 ng/ml +/- 1.18 in nephrotic patients versus 5.8 ng/ml +/- 1.23 in controls before venous occlusion and respectively 33.1 ng/ml +/- 3.83 versus 20.3 +/- 3.40 in controls after venous occlusion). Plasminogen activator activity of the euglobulins as assessed by the bovine fibrin-agarose plate was significantly higher in nephrotic patients only after venous occlusion. Plasma samples of nephrotic patients exerted a more potent inhibition of fibrinolysis in a urokinase activated system. This effect was, however, mainly due to the high levels of alpha 2 macroglobulin in nephrotic plasma which apparently have little influence on dilute blood clot lysis time.     

Role of alpha 2-antiplasmin in fibrin-specific clot lysis with single-chain urokinase-type plasminogen activator in human plasma

The role of plasma alpha 2-antiplasmin (alpha 2-AP) in the fibrin-specificity of clot lysis by recombinant single-chain urokinase-type plasminogen activator (rscu-PA) and in the conversion of rscu-PA to its two-chain derivative (rtcu-PA, urokinase) was investigated in an in vitro human plasma clot lysis system. Fifty % lysis in 2 h of a 0.1 ml 125I-fibrin labeled human plasma clot immersed in 0.5 ml normal human plasma was obtained with 1.4 +/- 0.15 micrograms/ml rscu-PA (mean +/- SD, n = 8). This was associated with degradation of 23 +/- 7% of fibrinogen and generation of 0.20 +/- 0.09 micrograms/ml rtcu-PA. In alpha 2-AP-depleted plasma 50% clot lysis in 2 h required 2-fold less rscu-PA which was associated with 3-fold more extensive fibrinogen degradation and 2-fold more rtcu-PA generation. Fifty % lysis in 2 h, of a 0.1 ml alpha 2-AP-depleted plasma clot, submersed in 0.5 ml normal plasma, was obtained with 0.80 +/- 0.05 micrograms/ml rscu-PA (n = 3, p less than 0.001 vs normal clot) and was associated with 17 +/- 6% fibrinogen breakdown (p = 0.22 vs normal clot) and 0.08 +/- 0.02 micrograms/ml rtcu-PA generation (p less than 0.05 vs normal clot). In alpha 2-AP-depleted plasma the equipotent rscu-PA concentration was 4-fold lower than in normal plasma and was associated with 3-fold more fibrinogen degradation and a similar extent of rtcu-PA generation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of fish oil supplementation on platelet survival and ex vivo platelet function in hypercholesterolemic patients

Little is known about the effects of dietary supplementation on platelet survival with low doses of n-3 and n-6 fatty acids in patients with hypercholesterolemia. The effects of a 6-week intervention with fish oil capsules (daily intake: 216 mg eicosapentaenoic acid, 140 mg docosahexaenoic acid, 390 mg gamma-linolenic acid, and 3480 mg linoleic acid) on in vivo platelet survival (111 In-oxine labeled platelets) and on ex vivo markers of platelet activation were investigated in a placebo-controlled, double-blind study with 26 hypercholesterolemic patients. In vivo platelet survival increased in the fish oil group (T) from a mean of 159+/-14 hours to a mean of 164+/-12 hours (p=0.025), whereas it remained unchanged in the placebo (P) group (T vs. P; p=0.055). Ex vivo, thromboxane B2 decreased from a mean of 225+/-16 to 212+/-21 ng/mL (p=0.003) in T but did not change in P (T vs. P: p=0.002). Malondialdehyde formation was lowered significantly by fish oil supplementation from a mean of 5.49+/-1.3 to 5.12+/-1.05 nM/10(9) platelets, p=0.005, as compared with P (T vs. P; p=0.018). The trendwise decrease in 11-DH-thromboxane B2 plasma levels was not significant nor was the increase in platelet sensitivity to prostaglandin I2 by fish oil. Baseline platelet survival in patients with hyperlipoproteinemia type IIa was not different from those with hyperlipoproteinemia IIb and response to treatment in terms of platelet activation markers was not either. The changes in platelet activation parameters in T were associated with significant reductions in cholesterol (-2.9%), low density lipoprotein cholesterol (-3.5%), and triglycerides (-12.4%). Both ex vivo and in vivo platelet activation parameters exhibited signs of decreased activation by a 6-week diet supplemented with n-3 and n-6 fatty acids, which might be beneficial in reducing atherothrombotic risk, in patients with hyperlipoproteinemia type IIa and IIb.     

The effect of plasma fibrinogen levels on measures of fibrinolytic activity

It has been claimed that lysis time methods may be inappropriate as measures of fibrinolytic activity on the grounds that they are largely determined by plasma fibrinogen levels. The dilute blood clot lysis time (DBCLT) has therefore been measured in 103 subjects; fibrinolytic activity (expressed as 100/DBCLT) has been compared with the fibrin plate area (FPA) lysed by the euglobulin fraction, the latter method being, of course, virtually independent of plasma fibrinogen. Plasma fibrinogen levels have also been measured; they ranged from 167 to 416 mg%. Results have been analysed by the technique of multiple regression. The incorporation of plasma fibrinogen levels in the regression equation relating 100/DBCLT to FPA does not result in a significant increase in the proportion of the variance in 100/DBCLT that can be explained. It is concluded that there is no evidence that DBCLT is influenced by plasma fibrinogen levels within the physiological range.     

The anatomical distribution of plasma fibrinolytic activity in man during cardiac catheterisation.

Regional circulating plasma levels of fibrinolytic activity were assessed in 15 patients undergoing cardiac catheterisation. The euglobulin clot lysis time (ECLT) was longer in the abdominal aorta (AA) than the inferior vena cava (IVC), (median difference -17.5 min, p = 0.008). This was associated with higher inhibition of plasminogen activator activity (PAI) in the AA than IVC, -1.75 IU/ml, p = 0.002. In the venous circulation the ECLT was higher in the peripheral venous sample than in the IVC, -25.5 min, p = 0.003, with higher PAI peripherally than in the IVC, -1.9 IU/ml, p = 0.001. There were no differences in ECLT, PAI, PAI-1:Ag or t-PA:Ag throughout the arterial circulation. These results demonstrate higher fibrinolytic activity with lower inhibitor activity in the venous compared to the arterial circulation. Within the venous circulation fibrinolytic activity is lower peripherally with increased inhibitor activity.     

Correlation between progressive adsorption of plasminogen to blood clots and their sensitivity to lysis

The binding of plasminogen to preformed human plasma clots immersed in citrated human plasma was measured and correlated with the sensitivity of these clots to lysis with recombinant tissue-type plasminogen activator (rt-PA), recombinant single-chain urokinase-type plasminogen activator (rscu-PA) or two chain urokinase-type plasminogen activator (tcu-PA, urokinase). When 0.15 ml plasma clots were compressed mechanically to about 1% of their original weight, and immersed in 0.15 ml plasma, 131I-labeled native plasminogen (Glu-plasminogen) adsorbed progressively from the plasma milieu onto the clot; binding was 3 +/- 1% (n = 10) after 1 h, 7 +/- 1% after 12 h and 12 +/- 1% after 48 h. This was associated with an increased sensitivity of the clot to lysis; 50% clot lysis in 4 h was obtained with 65 +/- 5 ng/ml (n = 3) rt-PA before and 30 +/- 5 ng/ml (n = 3) after 48 h preincubation in plasma (p less than 0.01), with corresponding values of 660 +/- 55 ng/ml (n = 3) and 280 +/- 25 ng/ml (n = 3) for rscu-PA, (p less than 0.01), and 800 +/- 85 ng/ml (n = 3) and 270 +/- 35 ng/ml (n = 3) for urokinase (p less than 0.01). Additional binding of plasminogen and increased sensitivity to lysis were reduced or abolished when the clot was preincubated in plasminogen-depleted or in t-PA-depleted plasma, or when 20 mM 6-aminohexanoic acid or 2,000 KIU/ml aprotinin were added.(ABSTRACT TRUNCATED AT 250 WORDS)     

Fibrinolysis and coagulation in patients with infectious disease and sepsis

Sepsis is often associated with hemostatic dysfunction. This study aimed to relate changes in fibrinolysis and coagulation parameters to sepsis and sepsis outcome. Urokinase-type plasminogen activator (u-PA) antigen, tissue-type plasminogen activator (t-PA) antigen and activity, plasminogen activator inhibitor (PAI) type 1 antigen, PAI activity, antithrombin (AT) III activity, and protein C activity were measured in 24 patients suffering from sepsis or septic shock and the results were compared with those observed in 30 non-sepsis patients with severe infectious disease. The u-PA level was markedly increased in plasma of sepsis patients as compared to non-sepsis patients (11.5 +/- 9.4 versus 1.6 +/- 1.5 ng/ml, p less than 0.0001). PAI-1 antigen and t-PA activity showed a significant increase in sepsis patients (320 +/- 390 ng/ml versus 120 +/- 200 ng/ml, and 3.0 +/- 3.6 IU/ml versus 1.0 +/- 0.7 IU/ml, respectively, p less than 0.01). AT III was decreased in sepsis patients (58 +/- 28% in sepsis versus 79 +/- 26% in severe infectious disease, p less than 0.01) as was protein C (30 +/- 18% versus 58 +/- 27%, p less than 0.001). No significant difference was found for t-PA antigen nor for PAI activity. Nonsurvivors of sepsis were distinguished mainly by a high u-PA antigen level and increased t-PA activity. It is concluded that plasma u-PA antigen showed the strongest significant difference, among the parameters evaluated, between sepsis and severe infection. u-PA antigen may be of prognostic value in patients admitted to the medical intensive care unit for severe infectious disease.     

A history of early stent thrombosis is associated with prolonged clot lysis time

It has been demonstrated that formation of compact plasma fibrin clots resistant to plasmin-mediated lysis characterises patients following in-stent thrombosis (IST). The relationship between defective fibrinolysis, reflected as prolonged clot lysis time (CLT) and IST is unclear. We sought to investigate whether patients with acute and subacute IST have impaired fibrinolytic capacity. We studied 41 definite IST patients, including 15 with acute and 26 with subacute IST experienced 2-73 months prior to enrollment, versus 41 controls matched for demographics, cardiovascular risk factors, concomitant treatment and angiographic/stent parameters. CLT, reflecting lysis of a tissue factor-induced plasma clot by exogenous tissue plasminogen activator, together with plasminogen activator inhibitor-1 (PAI-1) antigen and activity, thrombin-activatable fibrinolysis inhibitor (TAFI) antigen and activity, thrombomodulin (TM), plasminogen and α2-antiplasmin (α2AP) were measured. There were no inter-group differences in angiographic parameters, indication to the first PCI, culprit vessel or a type of stent. Patients with IST had 11% longer CLT (p=0.005) and 13% higher PAI-1 antigen (p=0.04) compared to controls. There were positive correlations in both groups between CLT and PAI-1 antigen and TAFI activity (all p<0.001). Multiple regression analysis showed that CLT (odds ratio [OR]=1.04 per 1 minute, 95% CI 1.01-1.08, p=0.02) and platelet count (OR=1.01 per 1,000/μl, 95% CI 1.00-1.02, p=0.034) were independent predictors of IST (R(2)=0.28, p<0.05). Concluding, impaired fibrinolytic potential, that is in part determined by plasma PAI-1 antigen and TAFI activity, characterises patients with a history of acute and subacute IST, which might help identify patients at higher risk of IST.     

Poor fibrinolytic response to venous occlusion by different criteria in patients with deep vein thrombosis.

Five criteria for poor response to a 20 min venous occlusion test were applied to 58 patients 3 months or more after acute deep vein thrombosis (DVT). The criteria were arbitrarily defined as the last 5 percentiles of response distributions in an age- and sex-matched healthy control group of 51 subjects. The criteria were: 1. euglobulin clot lysis time after venous occlusion greater than or equal to 140 min; 2. t-PA activity after venous occlusion less than or equal to 0.04 IU/ml; 3. increase in t-PA antigen above resting value less than or equal to 2-fold; 4. ratio between t-PA antigen increase and resting PAI activity less than or equal to 0.5 ng/IU; 5. PAI activity after venous occlusion greater than or equal to 6 IU/ml. The last criterion of poor response was the only one that was significantly more frequently reached by patients than by controls: 28% (p less than 0.005) of all DVT patients and 35% (p less than 0.005) of the subgroup with idiopathic DVT (N = 34) were found to be poor responders. The percentage of poor responders according to the other four criteria was 7-11% in all patients and 9-15% in the subgroup with idiopathic DVT and thus was not significantly higher than in controls (5% by definition). It was concluded that residual PAI activity after venous occlusion might be a useful criterion for prospective studies on recurrence of DVT.     

Altered lipid composition and thromboxane A2 formation in platelets from patients affected by IIa hyperlipoproteinemia

Platelets from patients with familial hypercholesterolemia (type IIa hyperlipoproteinemia), a condition associated with high prevalence of atherosclerosis and of its thrombotic complications, are known to be hyperresponsive to aggregating stimuli and to synthesize increased amounts of thromboxane A2 (TxA2) in comparison to platelets from normal subjects. In order to search if these functional alterations are linked to a different platelet lipid composition, we studied a group of young patients affected by IIa hyperlipoproteinemia and a group of suitable controls with similar dietary habits. Both cholesterol and phospholipid content of platelets were higher in patients than in controls with a significant increase of cholesterol/phospholipid molar ratio (at least p less than 0.05). The percent contents of the main platelet phospholipid fractions were not altered, while an increase in saturated fatty acids, both unesterified and esterified in different lipid fractions, was observed. Moreover, an increased TxA2 production by platelets and a significantly increased number of megathrombocytes occur in patients with respect to controls (p less than 0.001). Our results indicates that platelets from patients with IIa hyperlipoproteinemia have an altered lipid composition which could explain, at least in part, the enhanced platelet reactivity reported in these patients.     

Effects of recombinant plasminogen activator inhibitor type 1 on fibrinolysis in vitro and in vivo

A biologically active recombinant PAI-1 (rPAI-1) was evaluated for its effects on clot lysis in vitro and in vivo. At concentrations of 0.5 to 10 micrograms/ml, the rPAI-1 significantly prolonged the time to lysis of rabbit euglobulin clots both in the presence and absence of exogenous tissue plasminogen activator. To examine the effects of PAI in vivo, we infused rPAI-1 into conscious rabbits after i.v. injection of homogenized fibrin clots, and assessed fibrinolysis by measuring the appearance of d-dimer fibrin degradation products (FDP). Plasma fibrinolytic activity, PAI activity and antigen were also measured in plasma samples taken during and after infusion of rPAI-1. In control rabbits, endogenous fibrinolytic activity resulted in a significant and continual generation of FDP, reaching 32.6 +/- 13.6 ng/ml 90 minutes after fibrin injection. Infusion of 1, 2, or 5 micrograms/kg/min rPAI-1 led to dose dependent increases in PAI activity and antigen, while FDP levels at 90 minutes were only 8.8 +/- 2.9, 5.7 +/- 2.4, and 0.3 +/- 0.3 ng/ml, respectively. Complete inhibition of fibrinolysis was observed with 10 micrograms/kg/min rPAI-1. These studies directly demonstrate that increases in PAI-1 impair the fibrinolytic system, and support indirect observations of an association between elevated levels of this protein and thromboembolic diseases.     

Thrombolytic properties of leukocytes from peripheral blood in healthy subjects and in patients with acute cerebral ischemia

Polymorphonuclear leukocytes are activated in acute ischemic stroke. Activated polymorphonuclear leukocytes may contribute to thrombolysis by proteolytic degradation of fibrin and by modification of the plasminogen system. We used an in vitro thrombolysis model to investigate (1) thrombolytic properties of leukocytes in young and healthy subjects, (2) to test the hypothesis of increased polymorphonuclear leukocyte-associated thrombolysis in patients with acute cerebral ischemia, and (3) to assess plasminogen-dependent and -independent thrombolytic properties of polymorphonuclear leukocyte elastase. Coincubation of polymorphonuclear leukocytes with fibrin clots led to increased thrombolysis, a process reaching statistical significance after 8 hours [1x10(7) polymorphonuclear leukocytes/mL; 12.8+/-1.9% (mean+/-SEM), spontaneous clot lysis: 7.3+/-0.7%]. Polymorphonuclear leukocytes inside clots caused more efficient thrombolysis than polymorphonuclear leukocytes in the incubation medium. Spontaneous and polymorphonuclear leukocyte-associated lysis tended to be lower in patients with acute cerebral ischemia (n=9, 24 hours, 9.5+/-1.8% and 12.9+/-2.2%) than in age- and sex-matched control subjects (n=8; 12.2+/-2.0% and 17.4+/-1.9%). In the presence of alpha(2)-antiplasmin, thrombolysis tended to be faster with elastase-digested plasminogen (miniplasminogen) than with native plasminogen. Purified polymorphonuclear leukocyte elastase itself had no thrombolytic effect. We conclude that the thrombolytic capacity of polymorphonuclear leukocytes from peripheral blood is small and slow and may have been overestimated in previous reports. Polymorphonuclear leukocyte thrombolytic activity may not be increased in acute cerebral ischemia. Miniplasminogen may be an interesting adjunct to plasminogen activators in acute stroke models.     

A modified quantitative whole blood clot lysis method for general laboratory analysis of fibrinolysis

Because fibrinolysis is now recognized as an important factor in hypercoagulable states, we have developed and characterized an easily performed, rapid, and quantitative screening test that assesses a patient's fibrinolytic activity. This modification of the dilute whole blood clotting time (DWBCT) counts the number of intact erythrocytes released from the clot formed in samples obtained before and after the application of a venous occlusion cuff. Samples were corrected for the plasma volume changes that occurred during venous occlusion. This test was performed on nine healthy volunteers. Specimens were diluted 1:1 with PBS, rapidly clotted with thrombin and incubated at 37 C. Starting thirty minutes after the thrombin was added and then at twenty minute intervals until 110 minutes, the number of RBCs released from the clot were counted using a Coulter S Plus counter. There were consistently more RBCs released at each time period after venous occlusion (p less than 0.001). Aliquots were also obtained for measuring PAI activity and TPA levels. PAI activity was lower post-cuff at every point (p less than 0.001). TPA level was higher at every point (p less than 0.001) post-cuff. The addition of exogenous TPA, activated protein C, or anti-PAI antibodies increased the amount of clot lysis; while the addition of anti-TPA antibodies and EACA each prevented the post-cuff increase. Unlike the euglobulin lysis time (ELT) this modified DWBCT (mDWBCT) measures the patients intact fibrinolytic system, including PAI and erythrocytes, in a quantitative fashion. Unlike either the ELT or the DWBCT the mDWBCT can be performed within two hours, so results are rapidly available for clinical decisions. These studies have demonstrated an easily performed, inexpensive, quantitative screening test of a patient's overall fibrinolytic system that reacts appropriately to pharmacologic manipulations.     

Hypofibrinolysis associated with vasculopathy in non insulin dependent diabetes mellitus

The pathogenesis of diabetic vasculopathy has been related to modifications in hemostasis and fibrinolysis. 50 non insulin dependent diabetes mellitus patients have been studied. Euglobulin clot lysis time, fibrin plate, tissue plasminogen activator (t-PA) antigen, plasminogen activator inhibitor (PAI) activity, Protein C and S, cholesterol, triglycerides and Hb A1c were determined in blood samples. Diabetic patients showed decreased fibrinolytic activity, as measured by ECLT, with clearly increased PAI levels. Fibrinolytic response to venous occlusion was lower than normal. Vascular complications were associated both with an even higher PAI activity and with a decreased fibrinolytic response to venous occlusion. Elevated PAI activity and decreased fibrinolytic response to stimulus may contribute to vascular disease in diabetes.     

Thrombolytic and pharmacokinetic properties of human tissue-type plasminogen activator variants, obtained by deletion and/or duplication of structural/functional domains, in a hamster pulmonary embolism model

A pulmonary embolism model in hamsters was used for the quantitative evaluation of the thrombolytic and pharmacokinetic properties of variants of tissue-type plasminogen activator (t-PA). A 25 microliters 125I-fibrin labeled human plasma clot was made in vitro and injected into the jugular vein of heparinized hamsters. The extent of thrombolysis within 90 min was determined as the difference between the radioactivity injected in the jugular vein and that recovered in the heart and lungs. Recombinant t-PA (home-made rt-PA or Activase) infused intravenously over 60 min caused dose-dependent progressive thrombolysis. The results of thrombolytic potency (clot lysis in percent versus dose administered in mg/kg) and of specific thrombolytic activity (clot lysis in percent versus steady state plasma level in microgram/ml) were fitted with an exponentially transformed sigmoidal function y = 100 c/(1 + e-a(ax-eh] and the maximal percent lysis (c), the dose or plasma level at which maximal rate of lysis is achieved (b) and the maximal rate of lysis (z = 1/4 ac.eb) were determined. With rt-PA, these parameters were c = 72 +/- 6% (mean +/- SEM), b = 0.19 +/- 0.08 mg/kg, z = 68 +/- 25% lysis per mg/kg, with corresponding values of 87 +/- 5%, 0.07 +/- 0.03 mg/kg and 150 +/- 38% lysis per mg/kg for Activase (p = NS). Deletion of the finger and growth factor domains in rt-PA (rt-PA-delta FE) was not associated with marked alteration of the thrombolytic potency (c = 90 +/- 30%, b = 0.34 +/- 0.35 mg/kg, and z = 54 +/- 14% per mg/kg), but was associated with a significant reduction of the specific thrombolytic activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of kringles derived from human plasminogen on fibrinolysis in vitro

Plasminogen kringle 1+2+3 (K1-3) containing lysine-binding sites inhibited the reaction of plasmin with alpha 2-plasmin inhibitor (alpha 2PI), in a rate assay using a synthetic chromogenic substrate, S-2251. However, K1-3 did not inhibit the reaction to any degree between alpha 2PI and mini-plasmin which lacked the kringle 1 to 4 portion of plasmin. These results suggest that K1-3 blocked the binding of alpha 2PI to the lysine-binding site of plasmin. In the urokinase (UK)-induced fibrinolysis, K1-3 shortened the human plasma clot lysis time at low concentration (0.5-6 microM), and prolonged the lysis time at a high concentration (20 microM). Similar results were obtained in the lysis time of a fibrin clot consisting of plasminogen, fibrinogen and alpha 2PI isolated from human plasma. The kringle 4 (K4) of human plasminogen did not accelerate human plasma clot lysis at any concentration (1.2-24.1 microM). Furthermore, in the tissue plasminogen activator (TPA)-induced fibrinolysis, K1-3 also shortened both the lysis time of human plasma clot and fibrin clot as observed in UK-induced fibrinolysis, but K4 did not. The above findings indicate that the reaction of alpha 2PI with the lysine-binding site of plasmin is involved in the inhibition of plasmin activity by alpha 2PI, and in the presence of an inhibitor of this reaction, the balance of coagulofibrinolytic activity in plasma will be shifted towards the fibrinolytic side.     

Factor XII, plasma prekallikrein, alpha 2-macroglobulin and C1-inhibitor levels in renal allograft recipients during immunosuppression with cyclosporin A--sequential measurements over four months in 17 patients

Factor XII clotting activity (F XII), plasma prekallikrein amidolytic activity (PK), alpha 2-Macroglobulin (alpha 2-M) and C1-Inhibitor (C1-Inh) antigens have been measured in 17 patients immediately before and sequentially for up to four months after kidney transplantation. Before transplantation mean F XII and PK levels were normal (99 +/- 27% and 102 +/- 21%, respectively, mean +/- S.D.) and alpha 2-M and C1-Inh levels were slightly elevated (115 +/- 55% and 129 +/- 32%, respectively, mean +/- S.D.). In the first two weeks after transplantation a significant decrease of F XII to 65 +/- 27%, of PK to 67 +/- 20% and of alpha 2-M to 88 +/- 42%, and a rise of C1-Inh to 201 +/- 44% (mean +/- S.D.) were observed (2 p less than 0.005). F XII levels four month after operation remained significantly (2 p less than 0.05) lower than preoperatively. PK and alpha 2-M values, however, were significantly higher (2 p less than 0.05) at four months as compared to the pretransplant period. Mean F XII levels in the 17 patients at various time points after transplantation correlated positively with PK, alpha 2-M and serum albumin and negatively with CyA level and dose and serum bilirubin. PK and alpha 2-M correlated positively with each other and albumin and negatively with creatinine, bilirubin and CyA (2 p less than 0.01). Whether CyA has a direct influence on production or consumption of F XII, PK, alpha 2-M and C1-Inh, or whether the changes merely reflect altered protein metabolism awaits further study.     

Influence of factor XIIIa activity on human whole blood clot lysis in vitro

We studied the influence of Factor XIIIa (F XIIIa) activity on the lysis rate of fresh whole human blood clots, without using anticoagulants. Clotting was induced by exogenous thrombin, lysis by tissue-type Plasminogen Activator (t-PA) added before clotting. After various periods of time, lysis rates were determined by measuring the radioactivity in the supernatant of the clot originating from 125I-Fibrinogen added before clotting. Lysis rates were determined in the presence of endogenous F XIIIa and compared with those obtained after specific inhibition of F XIIIa activity. We used an IgG fraction of an antiserum quenching the F XIIIa activity. Addition of increasing amounts of the antibodies to normal blood resulted in a dramatic increase in clot lysis rate, concomitant with loss of F XIII activity. Lysis of blood clots from a patient with a congenital, homozygous, functional alpha 2-Antiplasmin (alpha 2-AP) deficiency (alpha 2-AP-Enschede) was not or slightly increased by the anti F XIII antibodies indicating that fibrin-fibrin crosslinking per se does not contribute essentially to resistance of the blood clot against fibrinolysis. Both active alpha 2-AP and F XIIIa are required for the major part of the F XIII-dependent resistance of whole blood clots against lysis.     

Diurnal variation of the fibrinolytic system

To elucidate which component(s) of the fibrinolytic system is (are) responsible for the diurnal variation of fibrinolytic activity we have studied several parameters of this system in 8 healthy male volunteers during a period of 24 h. Blood was collected at 8 a.m., 10 a.m., 12 a.m., 4 p.m., 8 p.m. and 8 a.m. next morning. The following tests were performed: euglobulin clot lysis time (ECLT), fibrinolytic activity of euglobulins on fibrin plates in the presence and absence of blocking antibodies to tissue-type plasminogen activator (t-PA) and/or urokinase (u-PA), overall plasminogen activator inhibitor (PAI) activity, antigen levels of t-PA, u-PA and PAI-1 and zymography of the euglobulin fraction after SDS-PAGE. From 8-10 a.m. to 4-8 p.m., total fibrinolytic activity increased by 113% (p less than 0.01) or 71% (p less than 0.01) when measured by ECLT or by fibrin plate assay, respectively. The immunoquenching experiments showed that this increase was entirely due to t-PA related activity whereas u-PA activity and t-PA/u-PA independent activity remained constant during the day. Average antigen levels of u-PA and t-PA in the afternoon were 6% and 25% lower than those measured in the morning. During this period, overall PAI activity and PAI-1 antigen decreased by 31% (p less than 0.01) and 52% (p less than 0.01) respectively. Electrophoretic-zymographic analysis of the euglobulins revealed that throughout the day the majority of t-PA was present in the form of the 110 kDa t-PA/PAI-1 complex. The intensity of this complex was lowest in the afternoon.(ABSTRACT TRUNCATED AT 250 WORDS)     

Platelet function and antithrombins in hyperlipoproteinemia type IIa

Platelet function was studied in 35 patients with type IIa hyperlipoproteinemia and 22 normal controls. Platelet aggregation by ADP in concentrations ranging from 2×10^?6 to 2×10^?5 M, and adrenaline 10^?6 M were similar in both groups. In contrast, aggregation induced by collagen and secondary aggregation elicited by bovine factor VIII were significantly lower in hypercholesterolemic patients, who also disclosed an increased plasmatic antiheparin activity. This plasmatic antiheparin activity was inversely correlated to plasmatic cholesterol concentration, (r = ?0.50, p<0.01). PF3 and PF4 were normal in patients with type IIa hyperlipoproteinemia. AntiFXa, antithrombin III, and heparin cofactor were also determined. Heparin cofactor was found to be significantly lower in hypercholesterolemic patients than in controls, and inversely correlated to plasmatic cholesterol concentration (r = ?0.51, p<0.05). These observations suggest that the thrombotic tendency in hypercholesterolemia may be related to a decrease of plasmatic heparin cofactor.