Two different mechanisms in patients with venous thrombosis and defective fibrinolysis: low concentration of plasminogen activator or increased concentration of plasminogen activator inhibitor.

Fibrinolytic components after venous occlusion and concentrations of tissue plasminogen activator inhibitor were studied in 100 consecutive patients with confirmed recurrent deep vein thrombosis or pulmonary embolism. After 20 minutes of venous occlusion the fibrinolytic response was decreased in 33 patients, as measured both amidolytically with S-2251 and on fibrin plates. Two different mechanisms responsible for the poor fibrinolytic response could be distinguished. Twenty two of the patients in whom the response was poor released normal amounts of tissue plasminogen activator antigen, as assayed by immunoradiometric assay, but had appreciably increased concentrations of tissue plasminogen activator inhibitor. The 11 other patients in whom the response was poor had both low tissue plasminogen activator activities and low tissue plasminogen activator antigen concentrations but normal concentrations of tissue plasminogen activator inhibitor. The results show not only that defective synthesis or release of tissue plasminogen activator may be important in the pathogenesis of venous thrombosis but also that a large group of patients with thrombosis have an increased concentration of the inhibitor to tissue plasminogen activator.     

Effect of moderate dose of alcohol with evening meal on fibrinolytic factors.

OBJECTIVES--To evaluate the effects of moderate consumption of alcoholic beverages on the fibrinolytic system and to assess whether these effects could help explain the relation between moderate alcohol consumption and reduced coronary heart disease. DESIGN--Four treatments were allocated in a randomised controlled order on four days over a period of 11 days. SETTING--Metabolic ward of research institute. SUBJECTS--Eight white healthy middle aged men. INTERVENTIONS--Subjects were provided with food for the 11 days. On the four study days mineral water or 40 g of alcohol in the form of beer, wine, or spirits was consumed at dinner early in the evening. MAIN OUTCOME MEASURES--Plasminogen activator inhibitor activity, tissue type plasminogen activator antigen, and tissue type plasminogen activator activity one hour before and one, three, five, nine, and 13 hours after dinner with mineral water or alcoholic beverages. RESULTS--After dinner with alcohol plasminogen activator inhibitor activity rose from 53 (SD 19)% to a maximum of 667 (283%) five hours after dinner (P < 0.001). Tissue type plasminogen activator antigen levels rose from 5.3 (2.2) micrograms/l to a maximum of 10.8 (3.8) micrograms/l nine hours after dinner with alcohol (P < 0.001). Plasminogen activator activity was reduced in the postprandial period (from 1387 (483) IU/l to 323 (288) IU/l five hours after eating; P < 0.001) but was higher than normal early the next morning (1516 (809) IU/l after alcohol, 779 (516) IU/l after water; P = 0.04). CONCLUSION--Moderate alcohol consumption with dinner affects plasminogen activator inhibitor activity, plasminogen activator antigen level, and tissue type plasminogen activator activity temporarily. The effects observed in the early morning are consistent with a decrease in risk of coronary heart disease in moderate drinkers.     

Fibrinolytic capacity of arm and leg veins after femoral shaft fracture and acute myocardial infarction.

The local fibrinolytic activity generated in the leg and arm veins during venous occlusion (fibrinolytic capacity) and the systemic fibrinolytic activity were measured at intervals in 11 patients after fracture of the femoral shaft and in 11 patients after acute myocardial infarction. In both groups the fibrinolytic capacity of the leg veins and the systemic fibrinolytic activity were significantly reduced two days after the onset of tissue injury. The fibrinolytic capacity of the arm veins was not altered. These results provide a possible explanation for the predilection of venous thrombosis for the leg veins after accidental trauma and acute myocardial infarction.     

Decreased fibrinolytic activity in porcine-to-primate cardiac xenotransplantation.

BACKGROUND: One major barrier to successful xenotransplantation is acute vascular rejection, a process pathologically characterized by microvascular thrombosis and diffuse fibrin deposition in transplant blood vessels. This pathologic picture may result from a disturbance in the coagulant or fibrinolytic pathways that regulate normal vascular patency. This study evaluated the regulation of fibrinolytic activity defined by tissue plasminogen activator and plasminogen activator inhibitor-1 as it may exist in the setting of acute vascular rejection. MATERIALS AND METHODS, RESULTS: Serial biopsies from cardiac xenotransplants evaluated by immunofluorescence microscopy demonstrated progressive decreases in tissue plasminogen activator and increases in plasminogen activator inhibitor-1. In vitro studies measuring fibrinolytic activity of cell culture medium from porcine aortic endothelial cells stimulated with human serum or autologous porcine serum revealed that human serum triggered as much as 93% increase in antifibrinolytic activity. CONCLUSIONS: These findings demonstrate that porcine vascular endothelial cells change toward an antifibrinolytic state following stimulation with human xenoreactive antibodies and complement. The shift is at least partly explained by an increased ratio of plasminogen activator inhibitor-1 to tissue plasminogen activator, and is at least in part mediated by the activation of complement. This increased antifibrinolytic activity may contribute to the thrombotic diathesis seen in acute vascular rejection in pig-to-primate xenografts.     

Influence of beta-sitosterol on the fibrinolytic potential in rabbits

Long-term treatment of rabbits with beta-sitosterol (40 mg/kg over 3 months) caused an increased fibrinolytic activity in blood, an increased fibrinolytic capacity and an enhanced plasminogen activator activity in tissue of lungs and kidneys. The 3-months lasting beta-sitosterol administration did not influence the content of plasminogen activator inhibitor, plasminogen, alpha 2-antiplasmin, antithrombin III and fibrinogen.     

Fibrinolytic activity of prostacyclin and iloprost in patients with peripheral arterial disease

We studied the effects of prostacyclin (PGI2) and its stable analog, iloprost, on blood fibrinolytic activity in 33 patients with peripheral arterial disease. Ten subjects (group A) received three 5-hour infusions of iloprost on three consecutive days. The remaining 23 patients received three different 5-hour infusions (placebo, iloprost 2 ng/kg/min, PGI2 5 ng/kg/min). Tissue plasminogen activator (t-PA), total plasma fibrinolytic activity and euglobulin clot lysis time (ECLT) were determined in patients before and after each infusion, both in freely flowing blood samples and following 10 min venous occlusion. In patients of group A, ECLT at rest was significantly shortened after all three iloprost infusions (on average by about 5-11%). First and third infusions produced also shortening of ECLT after venostasis (by 21 and 32%). Statistically significant rise in t-PA activity (by about 68% on average) accompanied only the first infusion. In patients of the group B iloprost provoked significant fall in ECLT at rest (by about 19% on average) only. PGI2 shortened ECLT both at rest and after venous occlusion (by about 17% and 20% on average, respectively) and led to a rise in t-PA activity after venous occlusion by about 33% on average. Our results indicate that prostacyclin and its stable analog, iloprost, enhance fibrinolytic activity in man by releasing or facilitating the release of tissue plasminogen activator from the vessel wall.     

Fibrinolytic activity of prostacyclin and iloprost in patients with peripheral arterial disease

We studied the effect of prostacyclin /PGI₂/ and its stable analog, iloprost, on blood fibrinolytic activity in 33 patients with peripheral arterial disease. Ten subjects /group A/ received three 5-hour infusions of iloprost on three consecutive days. The remaining 23 patients received three different 5-hour infusions /placebo, iloprost 2 ng/kg/min, PGI₂ 5 ng/kg/min/. Tissue plasminogen activator /t-PA/, total plasma fibrinolytic activity and euglobulin clot lysis time /ECLT/ were determined in patients before and after each infusion, both in freely flowing blood samples and following 10 min venous occlusion. In patients of group A, ECLT at rest was significantly shortened after all three iloprost infusions /on average by about 5–11%/. First and third infusions produced also shortening of ECLT after venostasis /by 21 and 32%/. Statistically significant rise in t-PA activity /by about 68% on average/ accompanied only the first infusion. In patients of the group B iloprost provoked significant fall in ECLT at rest /by about 19% on average/ only. PGI₂ shortened ECLT both at rest and after venous occlusion /by about 17% and 20% on average, respectively/ and led to a rise in t-PA activity after venous occlusion by about 33% on average. Our results indicate that prostacyclin and its stable analog, iloprost, enhance fibrinolytic activity in man by releasing or facilitating the release of tissue plasminogen activator from the vessel wall.     

Position and developmental trends in fibrinolytic therapy

Increasing insight into the mechanism of fibrinolysis and particularly into the formation and release of plasminogen activator has led to more effective thrombolytic therapy. The understanding of the mechanism of thrombolysis has provided the possibility to improve the therapeutic effects of the fibrinolytic agents streptokinase and urokinase. Further advances in thrombolytic therapy are expected by the use of the plasminogen activator from tissue endothelium and pro-urokinase. Acylation of fibrinolytic enzymes will lead to beneficial effects (depot effect, protection from intrinsic inhibitors). Due to the extensive research into substances with fibrinolytic and thrombolytic effects a new generation of activators of fibrinolysis is expected that interfere with the biosynthesis and release of plasminogen activator of the vessel wall and that are suited for treatment of hypofibrinolytic states.     

Plasma levels of t-PA and PAI-1 correlate with the formation of experimental post-surgical peritoneal adhesions

This study has evaluated whether systemic changes of plasminogen activator (t-PA) and plasminogen activator inhibitor-1 (PAI-1) parallel the adhesions development and whether they could be used as predictors of adhesion risk. This has been studied in an animal model of post-surgical peritoneal adhesion by monitoring for 10 days the plasma and tissue levels of t-PA and PAI-1. The results showed that both tissular and plasmatic levels of t-PA were decreased in concomitance with the development of peritoneal adhesions. In contrast, PAI-1 was found increased into the tissue and into the plasma samples of the rats taken at 5 and 10 days time points. Inflammatory mediators such as ICAM-1, VCAM-1, and IL-6 within the peritoneal lavage fluid also correlated with the adhesion formation process. In conclusion, post-surgical peritoneal adhesions provide alterations of local inflammatory components and local and systemic fibrinolytic components, possibly with PAI-1 quenching t-PA. This may have potential for the identification of high-risk patients.     

Fibrinolytic activity of heart tissues and aorta of rats undergoing electroshock

In the first minute of shock caused by electric current passing through the rats parietal lobe the increase of fibrinolytic activity of euglobulin fraction of blood occurs. During the investigation of the right heart ventriculus tissue and aorta by histochemical method of Todd in our modification the decrease of fibrinolytic activity in wall of aorta (on 80%) and in tissue of heart ventriculus (on 30%) has been revealed in tentative animals as compared with control ones. It is suggested that the entering of plasminogen activator from vessel wall into the blood flow plays an important role in activation of fibrinolytic system in generalized reaction of organism to strong stress action.     

Effect of tissue-type plasminogen activator and its inhibitor in haemostasis system function in norm and pathology

The paper is dedicated to the study of structure and physiological functions of tissue type plasminogen activator and its inhibitor PAI-1, changes of these parametres in normal and pathology conditions. The interrelation of the coagulation and the fibrinolytic system during the range of pathologies was studied. It was demonstrated that simultaneous analysis of the coagulation and fibrinolytic systems analysis allow to diagnose the thrombotic complications.     

Effect of Influenza Virus Proteins Modulate Hemostasis in vitro and in vivo

We studied the effect of influenza virus proteins—hemagglutinin, neuraminidase, nucleoprotein, and membrane protein—on hemostasis in vitro and in vivo. The results demonstrated that envelope proteins hemagglutinin and neuraminidase enhanced the fibrinolytic and anticoagulant activities of blood plasma and the activity of human tissue plasminogen activator. The membrane protein proved to have the highest activity among the core proteins of influenza virus; in contrast to hemagglutinin and neuraminidase, it inhibited fibrinolysis, increased the coagulant activity of blood plasma, and decreased the activity of human tissue plasminogen activator. The combined action of hemagglutinin and neuraminidase increased the plasma fibrinolytic and anticoagulant activities to a greater extent than the individual action of each agent. The combined action of hemagglutinin and membrane protein also increased the plasma fibrinolytic and anticoagulant activities, although to a lesser extent than the action of hemagglutinin alone. These data indicate that viral proteins are physiologically active and can cause influenza-specific disturbances of hemostasis.     

凝血系统相关基因突变及表达异常与高血凝

摘要高血凝是动脉粥样硬化(As)的危险因子,在As的发展中具有重要作用。凝血系统、抗凝系统、纤溶系统及其它相关基因的突变及表达异常导致高血凝的产生。凝血系统的凝血因子V基因、凝血酶原基因、组织因子基因,抗凝系统的血栓调节蛋白基因、抗凝血酶Ⅲ基因,纤溶系统的纤溶酶原激活物抑制剂-1基因,均与高血凝密切相关。     

华广虻(Tabanusamaenus Walker)溶纤活性蛋白的纯化及生物活性分析

经过 ７５％ 饱和度硫酸铵沉淀、 Ｓｅｐｈａｄｅｘ Ｇ ７５ 凝胶过滤层析、 Ｌｙｓ Ｓｅｐｈａｒｏｓｅ ４ Ｂ 亲和层析和电泳制备洗脱,从华广虻（ Ｔａｂａｎｕｓ ａｍ ａｅｎｕｓ Ｗ ａｌｋｅｒ）腹部组织匀浆液中分离纯化出分子量约为 ６７ｋ Ｄ 的溶纤活性蛋白 Ｔ Ａ Ｆ Ｐ经纤维蛋白平板测定表明, Ｔ Ａ Ｆ Ｐ 只具有纤溶酶作用,不具有激活纤溶酶原的作用;但 Ｔ Ａ Ｆ Ｐ 能分解纤溶酶原激活剂的生色底物—— Ｃｈｒｏｍ ｏｚｙｍ Ｕ Ｋ 及 Ｓ ２２８８还能水解胰蛋白酶专一底物 Ｂｚ Ｐｈｅ Ｖａｌ Ａｒｇ Ｎ Ａ 及 Ｃ Ｂ Ｚ Ｇｌｙ Ｐｒｏ Ａｒｇ Ｎ Ａ,表明 Ｔ Ａ Ｆ Ｐ具有类胰蛋白酶活性,专一水解精氨酸形成的酰胺键（或肽键） Ｔ Ａ Ｆ Ｐ无胰凝乳蛋白酶活性      

Functional effects of asparagine-linked oligosaccharide on natural and variant human tissue-type plasminogen activator

The role of Asn-linked oligosaccharide in the functional properties of both human tissue-type plasminogen activator (t-PA) and a genetic variant of t-PA was studied. Nonglycosylated and glycosylated wild-type t-PA were produced in mammalian cells which express recombinant t-PA. These proteins were compared in fibrin binding and 125I-labeled fibrin clot lysis assays, using purified components. The nonglycosylated form showed higher fibrin binding, as well as higher fibrinolytic potency than the glycosylated form. Subsequently, prevention of glycosylation of a t-PA variant which lacked the finger and epidermal growth factor domains (delta FE), was carried out in an attempt to enhance its fibrinolytic activity. Glycosylation was prevented by changing Asn to Gln; at Asn-117 to produce delta FE1X t-PA, and at Asn-117, -184, and -448 to produce delta FE3X t-PA. All variants were similar to wild-type t-PA in their catalytic dependence on fibrinogen fragments, fibrinolytic activity in fibrin autography analysis, and plasminogen activator activity. In a clot lysis assay, using citrated human plasma, the fibrinolytic potency of the variants were comparable to that of wild-type t-PA at activator concentrations of 17-51 nM (approximately 1-3 micrograms/ml). At 0.5-5.1 nM (approximately 0.03-0.3 micrograms/ml), however, the variant proteins had lower fibrinolytic potency than wild-type t-PA. Fifty percent lysis in 1.5 h for wild-type, delta FE, delta FE1X, and delta FE3X t-PA, required 2.5, 10, 7.5, and 5.5 nM t-PA, respectively. The fibrinogenolytic activity in human plasma was measured for wild-type, delta FE, delta FE1X, and delta FE3X t-PA, and showed significant fibrinogen depletion after 3 h of incubation at 51 nM, decreasing to 11, 11, 50, and 72% of basal levels, respectively. These data indicate that partial or total nonglycosylated t-PA variants have a higher fibrinolytic versus fibrinogenolytic ratio than their fully glycosylated counterparts.     

Enhanced expression of genes involved in coagulation and fibrinolysis in murine arthritis

We have analyzed the pattern of procoagulant and fibrinolytic gene expression in affected joints during the course of arthritis in two murine models. In both models, we found an increased expression of tissue factor, tissue factor pathway inhibitor, urokinase plasminogen activator, and plasminogen activator inhibitor 1, as well as thrombin receptor. The observed pattern of gene expression tended to favor procoagulant activity, and this pattern was confirmed by functional assays. These alterations would account for persistence of fibrin within the inflamed joint, as is seen in rheumatoid arthritis.     

Fibrinolytic components in nasal mucosa and nasal secretion

 We evaluated a possible role for fibrinolytic components in nasal secretion by tissue localization with immunohistochemical techniques and by measuring their antigen concentrations in nasal discharge by means of ELISA and fibrin autography. Nasal mucosa was obtained surgically from the inferior turbinate. Urokinase-type plasminogen activator (u-PA) specific staining was observed in pseudostratified ciliated epithelium and was predominant in mucous cells of the seromucinous gland, while serous cells were almost devoid of stain. The pattern of staining of plasminogen activator inhibitor-2 was similar to that of u-PA. In contrast, plasminogen activator inhibitor-1(PAI-1) immunoreactive material was localized exclusively in serous cells of seromucinous glands. Positive staining for tissue-type plasminogen activator (t-PA) was observed in endothelial cells and basal cells, which differentiate into either ciliated or goblet cells. Nasal secretions were partially fractionated by immunospecific antibody-immobilized Sepharose. Subsequent fibrin autography patterns indicated the presence of u-PA, PAI-1, and t-PA. After methacholine provocation, the level of t-PA increased transiently but decreased rapidly with subsequent challenges. These differential stainings of fibrinolytic components and the existence of PAs and PAI-1 in the nasal discharge suggest that the fibrinolytic system may play a role in the movement and fluidity of nasal secretion. Accepted: 25 May 1998     

Normal tissue plasminogen activator and plasminogen activator inhibitor activity in plasma from patients with type 1 diabetes mellitus.

The fibrinolytic system was investigated in 38 patients (21 males and 17 females) affected by type 1 diabetes mellitus (18 free from complications, 10 with retinopathy, and 10 with autonomic neuropathy) and in 8 healthy controls. Two separate fibrinolysis-stimulating tests were done: standardized venous occlusion and 1-desamino-8-D-arginine vasopressin infusion. Plasma tissue plasminogen activator antigen and activity and plasma plasminogen activator inhibitor activity were measured. All the patients were in good metabolic control (mean HbA1c 7.4%, range 6.1-8.0%). No significant differences were observed either between the diabetic patients and the control subjects, nor among the subgroups of diabetic patients. The fibrinolytic system is probably not involved in type 1 diabetes mellitus.     

Endothelial cell annexin A2 regulates polyubiquitination and degradation of its binding partner S100A10/p11

The annexin A2 (A2) heterotetramer, consisting of two copies of A2 and two copies of S100A10/p11, promotes fibrinolytic activity on the surface of vascular endothelial cells by assembling plasminogen and tissue plasminogen activator (tPA) and accelerating the generation of plasmin. In humans, overexpression of A2 by acute promyelocytic leukemia cells is associated with excessive fibrinolysis and hemorrhage, whereas anti-A2 autoantibodies appear to accentuate the risk of thrombosis in patients with anti-phospholipid syndrome. Complete deficiency of A2 in mice leads to a lack of tPA cofactor activity, accumulation of intravascular fibrin, and failure to clear arterial thrombi. Within the endothelial cell, p11 is required for Src kinase-mediated tyrosine phosphorylation of A2, which signals translocation of both proteins to the cell surface. Here we show that p11 is expressed at very low levels in the absence of A2 both in vitro and in vivo. We demonstrate further that unpartnered p11 becomes polyubiquitinated and degraded via a proteasome-dependent mechanism. A2 stabilizes intracellular p11 through direct binding, thus masking an autonomous p11 polyubiquitination signal that triggers proteasomal degradation. This interaction requires both the p11-binding N-terminal domain of A2 and the C-terminal domain of p11. This mechanism prevents accumulation of free p11 in the endothelial cell and suggests that regulation of tPA-dependent cell surface fibrinolytic activity is precisely tuned to the intracellular level of p11.     

Fibrinolysis mediated by tissue plasminogen activator. Disclosure of a kinetic transition

The rate of 'Glu'-plasminogen activation by tissue plasminogen activator was repeatedly determined during a fibrinolytic process. The process was found to proceed via two distinct phases. The kinetics of each phase obeyed Michaelis-Menten equation: First phase; kcat about 0.17 s-1 and Km about 1 microM, second phase; kcat about 0.13 s-1 and Km about 0.06 microM. Practically identical results were obtained with one-chain as with two-chain tissue plasminogen activator. Transition from first to second phase occurred when the system had been exposed to a certain degree of plasmin digestion. Electrophoretic analysis demonstrated time correlation between the appearance of minimally degraded fibrin (X-fragments) and the transition. No such correlation was found between transition and conversion of 'Glu'-plasminogen to 'Lys'-plasminogen. The effect can result in an acceleration (up to 13-fold) of the fibrinolytic process once a slight degradation of the fibrin has taken place. In vivo, the effect described may constitute a mechanism that protects a fibrin clot from premature lysis.