Regulation and secretion of plasminogen activators and their inhibitors in a human leukemic cell line (K562)

The secretion of tissue plasminogen activator (t-PA), urokinase (u-PA) and their inhibitors by the human leukemia cell line K562 was examined. K562 cells normally secrete both t-PA and u-PA in a ratio of 3:1. After addition of 10 or 1 ng/mL phorbol myristate acetate (PMA) to K562 cells, a marked decrease in enzymatic activity is observed in the medium. However, when t-PA antigen rather than activity is measured, an increased amount is found in the medium under these conditions. PMA also induces secretion of the two inhibitors of plasminogen activator: plasminogen activator inhibitor 1 (PAI-1) and plasminogen activator inhibitor 2 (PAI-2). This accounts for the decrease in total enzymatic activity under conditions when production of t-PA antigen is increased. A study of the time course of induction revealed that the synthesis of plasminogen activator occurred before that of its inhibitors. Low concentrations of PMA (0.1 ng/mL) induce t-PA antigen primarily and not the inhibitors. This results in an increase in total enzymatic activity, with 94% of the secreted activity being t-PA. Thus, the secretion of plasminogen activators and their inhibitors can be manipulated in certain leukemic cells by inducers such as PMA.     

Plasminogen activator inhibitor 2 and urokinase-type plasminogen activator in plasma and leucocytes in patients with severe sepsis

Proteins influencing plasminogen activation to plasmin, namely plasminogen activators tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA) and their principal inhibitors, plasminogen activator inhibitor 1 (PAI-1) and PAI-2, were measured in the plasma, the polymorph and mononuclear cell fractions taken from patients with major sepsis who were entering a general intensive care unit. The purpose of this study was to elucidate the factors favouring the persistence of fibrin in the microvasculature and thus contributing to multiple organ failure. Levels of u-PA antigen in plasma rose in sepsis and u-PA activity, not detectable in normal plasma, appeared. Levels of u-PA antigen in the cell fractions fell concomitantly. t-PA antigen in plasma and in the mononuclear cell fraction rose in sepsis, but t-PA activity was not detectable. Plasma PAI-1 antigen levels were strikingly raised in sepsis, presumably accounting for the complete neutralization of t-PA activity. PAI-2 antigen, not normally detected in plasma, appeared in the plasma of some patients, whereas it disappeared from the cellular fractions. Appearance of PAI-2 in plasma was associated with non-survival of the patient. The observations indicate that all the agents involved in plasminogen activation are released into the plasma in major sepsis. The levels of PAI-1 reached were quantitatively sufficient to suppress all activity of the released t-PA, but the inhibitors did not prevent expression of u-PA activity in the circulation. Circulating active u-PA and PAI-2 in the plasma of patients with severe sepsis may represent material originating from leucocytes. Leucocyte release of these agents within fibrin deposits may influence the persistence of fibrin and thus the development of multiple organ failure.     

The release of plasminogen activators (t-PA and u-PA) and plasminogen activator inhibitor (PAI-1) after venous stasis.

The aim of the present study was to compare plasma levels of urokinase-type plasminogen activator (u-PA), before and after 20 min of venous stasis, with those of tissue-type plasminogen activator (t-PA), type 1 plasminogen activator inhibitor (PAI-1) and t-PA/PAI-1 complexes, to determine whether both plasminogen activators and their inhibitor respond similarly to the same stimulus. We studied 36 patients with recurrent venous thrombosis in whom no coagulation defects predisposing them to thrombosis had been detected (mean age 38.2 years, range 15-70 years). Twenty healthy individuals (mean age 34.3 years, range 20-60 years) served as a control group. t-PA, PAI-1 and u-PA activity and antigen, as well as the t-PA/PAI-1 complex antigen, were measured before and after venous stasis. Post-stasis fibrinolytic parameters were corrected for the haemoconcentration which occurred during the venous occlusion test. Pathologically high PAI-1 levels (antigen and activity) were found in four out of 36 patients who were excluded from study. Functional and antigenic u-PA increased significantly after venous stasis when analysed as the absolute differences between paired samples (P less than 0.01). This increase in u-PA did not correlate with changes in t-PA or PAI-1 (r = 0.28 and r = 0.36 respectively). This leads us to suggest that different mechanisms relating to clearance and/or release from diverse sources might be involved in elevations of u-PA in response to a local endothelial stimulus. We conclude that venous stasis might not be the elective choice when evaluating 'bad responders' predisposed to thrombosis.     

Plasminogen activator in acute myeloid leukaemic marrows: u-PA in contrast to t-PA in normal marrow

Leukaemic and normal bone marrow samples were compared in terms of their content of the fibrinolytic agents, tissue plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA) and their inhibitors, plasminogen activator inhibitors 1 and 2 (PAI-1 and PAI-2). Normal marrow contained t-PA as the principal plasminogen activator, whereas in leukaemic marrow samples u-PA was the predominant activator. Both normal and leukaemic marrows contained PAI-1 in similar amounts, but whereas normal marrow contained significant amounts of PAI-2 the leukaemic marrows contained very little. Plasminogen activators were present in uncomplexed, active forms and plasmin–α₂-antiplasmin complexes were generated locally more prominently in leukaemic marrows. u-PA associated with blast cells may contribute to the severe forms of haemorrhage sometimes occurring in myeloid types of leukaemia.     

Interleukin-4 Modulation of Platelet-Derived Growth Factor–Induced Smooth Muscle Cell Urokinase Plasminogen Activator

Platelet-derived growth factor (PDGF) stimulates smooth muscle cell (SMC) migration owing to stimulation of SMC tissue plasminogen activator (t-PA) production. In this study we examined the effects of the T-cell lymphokine interleukin-4 (IL-4) on PDGF induction of human aortic SMC antigen levels of urokinase-type plasminogen activator (u-PA) and those of plasminogen activator inhibitor-1 (PAI-1), the endogenous inhibitor of t-PA and u-PA, measured by enzyme-linked immunosorbent assays (ELISAs). u-PA antigen levels from human aortic SMC incubated with PDGF 100 ng/mL and IL-4 500 U/mL were significantly greater than those incubated with PDGF 100 ng/mL alone. Coincubation of PDGF with IL-4 did not significantly increase SMC u-PA antigen levels in cellular lysates. Coincubation with PDGF 100 ng/mL and IL-4 500 U/mL did not significantly affect SMC PAI-1 antigen levels in conditioned media or cellular lysates. Therefore, interleukin-4 modulates vascular SMC u-PA production induced by PDGF.     

DIFFERENCE IN EXPRESSION OF THE PLASMINOGEN ACTIVATION SYSTEM IN SYNOVIAL TISSUE OF PATIENTS WITH RHEUMATOID ARTHRITIS AND OSTEOARTHRITIS

Protcolytic joint destruction in inflammatory and non-inflammatoryarthropathy is believed to be mediated, at least in part, bythe plasminogen activation (PA) system. To further investigatepossible involvement of the PA system, we quantified immunoreactiveurokinase-type plasminogen activator (u-PA), tissue-type plasminogenactivator (t-PA), both plasminogen activator inhibitors (PAI-1and PAI-2) and u-PA-receptor (u-PAR) in synovial tissue extractsof 14 patients with rheumatoid arthritis (RA) and 12 with osteoarthritis(OA). u-PA, PAI-1, PAI-2 and u-PAR concentrations were significantlyhigher in RA than in OA patients. t-PA antigen levels were significantlylower in RA than in OA synovial tissue extracts. Immunohistochemistrywas performed to compare the distribution and staining intensityof these components in samples of RA and OA synovial tissue.Intense immunostaining of u-PA, u-PAR, PAI-1 and, to a lesserdegree, PAI-2 was observed predominantly in the synovial liningof RA patients. In OA patients, u-PA, PAI-1, PAI-2 and u-PARwere barely detectable. t-PA immunostaining was restricted tothe endothelial side of vascular walls in both groups. We concludethat the observed increase of u-PA, u-PAR and PAI expression,distributed mainly in the synovial lining area of proliferativeand invasively growing synovial tissue in RA patients, supportsa pathogenic role for the PA system in destructive arthritis.Depressed t-PA-mediated plasminogen activation might contributeto delayed intra-articular fibrin removal. KEY WORDS: Urokinase, Plasminogen activation, Immunohistochemistry, Rheumatoid arthritis, Osteoarthritis     

Tumor necrosis factor induces the production of urokinase-type plasminogen activator by human endothelial cells

Endothelial cells play an important role in the regulation of fibrinolysis by the production of several key regulatory proteins. The cytokines tumor necrosis factor (TNF), lymphotoxin, and interleukin-1 (IL-1), but not interleukin-6, increase the production of plasminogen activator inhibitor-1 (PAI-1) by endothelial cells, whereas they have no stimulatory effect on the production of tissue-type plasminogen activator (t-PA). Primary cultures of human endothelial cells release very little urokinase-type plasminogen activator (u-PA). We report here that TNF and lymphotoxin induce, in a concentration-dependent way, the production of both cellular and secreted u-PA antigen in primary and subcultured human endothelial cells. The TNF-induced increase was accompanied by a more than 10-fold increase in u-PA mRNA. Upon stimulation of early passage umbilical vein endothelial cells by TNF, u-PA was predominantly secreted at the basolateral side, whereas PAI activity and t-PA were found in more equal amounts at the apical and basolateral sides of the cell monolayers. TNF-stimulated u-PA secretion by subcultured human aorta endothelial cells showed only a marginal polarity. The u-PA antigen was present in a plasmin-activatable form (single chain u-PA) and in a nonactivatable form (probably u-PA: PAI-1 complex). During the induction of u-PA by TNF, the ratio between plasmin-activatable u-PA and total u-PA decreased markedly. This may indicate that TNF also increases the degree of u-PA activation. The parallel induction of the synthesis and secretion of both u-PA and PAI-1 by endothelial cells adds a new aspect to the alterations of the fibrinolytic system caused by inflammatory mediators. This aspect may be significant for the regulation of cell-associated and interstitial plasminogen activator activity.     

Gastric mucosal plasminogen activators inHelicobacter pylori infection

Long-termH. pylori associated gastritis is recognized as a pathogenic factor in gastric carcinogenesis. In gastric carcinomas the amount and activity of the tissue-type plasminogen activator (t-PA) have been reported to be decreased, whereas those of the urokinase-type plasminogen activator (u-PA) were increased, contributing to the neoplastic and invasive process. The present study was performed to determine t-PA and u-PA levels and activity in gastric mucosa from 102 patients and to investigate whether these levels are influenced byH. pylori infection. The antigen concentration and activity of t-PA and u-PA in corpus mucosa were low (P<0.001) compared with those in antral mucosa, although for the u-PA activity this did not reach statistical significance. InH. pylori-associated antral gastritis the mucosal t-PA antigen concentration and activity were found to be decreased (P<0.001) compared with normal mucosa, whereas inH. pylori-associated pangastritis the corpus t-PA levels were not affected. The antigen concentration and activity of u-PA were found to be significantly (P<0.005) increased, both inH. pylori-associated gastritis of antrum and corpus mucosa. Levels of u-PA in histologically normal corpus mucosa of patients with anH. pylori-associated antral gastritis were also found to be increased (P<0.05). In conclusion, the alterations in the plasminogen activator profile found inH. pylori-associated gastritis, ie, a decrease in t-PA and an increase in u-PA, show a similar tendency as the previously found alterations in gastric carcinomas, which provides additional support for the possible involvement ofH. pylori-associated gastritis in the pathogenesis of gastric carcinoma.     

Fibrinogenolysis in thrombotic thrombocytopenic purpura

Coagulo-fibrinolytic factors were studied in five patients suffering from thrombotic thrombocytopenic purpura (TTP). The change in coagulation factors in the acute stage was mild compared with that found in disseminated intravascular coagulation (DIC). We observed a slight increase of fibrin-fibrinogen degradation products (FDP) in the plasma of four patients during the acute stage of TTP, but the level of the D-dimer remained within normal variation and was extremely low compared with that in 27 samples from patients with DIC showing the same level of FDP. At the same time, both antigen levels of tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type 1 (PAI-1) were elevated in three of the four patients tested. Although a similar change was recognized in DIC patients' plasma, the elevation of PAI-1 in the acute stage of TTP was far higher than in overt DIC. The antigen levels of t-PA and PAI-1 were normal in remission, and a mild elevation of PAI-1 was detected in one of the three patients during the early stage of TTP relapse. Enzymography revealed the appearance of free t-PA and an increase of a substance with a 110 kD molecule, assumed to be a t-PA and PAI-1 complex, in TTP plasma in the acute stage, but the findings were normal for plasma from cases in remission and the early stage of relapse. Enzymography also showed a decrease of urokinase-type plasminogen activator (u-PA) only in the acute stage of TTP. These changes in the coagulo-fibrinolytic factors in the acute stage of TTP suggest that fibrinogenolysis might be induced by t-PA, released through vascular reaction at an uninvolved area of vascular lesions caused by platelet agglutinates, which would then release large amounts of PAI-1 inhibiting t-PA and u-PA activities at the occlusive lesion.     

Antithrombotic regulation in human endothelial cells by fibrinolytic factors

Vascular endothelial cells (ECs) modulate the blood fibrinolytic system by secreting tissue-type plasminogen activator (t-PA), urokinase-type plasminogen activator (u-PA), and their inhibitor, type-1 plasminogen activator inhibitor (PAI-1). ECs also express t-PA receptors (t-PAR) and u-PA receptors (u-PAR) on their cell surfaces, assembling both enzymes to regulate the cellular fibrinolytic activity. In addition, ECs modulate these factors in response to several stimuli. Fibrin clots on ECs induce the up- and downregulation of t-PA and PAI-1 production, respectively, thus causing an effective lysis of the fibrin clot. Heat shock (43 degrees C) increases the expression of u-PA, t-PA, PAI-1, and u-PAR by which ECs become more fibrinolytic around the cells. Furthermore, because ECs possess t-PAR and u-PAR on their cell surfaces, the binding of t-PA and u-PA is a critical event, which affords ECs the localized and condensed fibrinolytic potential. Therefore, ECs play a central role in antithrombotic activity by regulating the levels of these fibrinolytic factors.     

Residual plasminogen activator inhibitor activity after venous stasis as a criterion for hypofibrinolysis: a study in 83 patients with confirmed deep vein thrombosis

In eighty-three patients with confirmed deep vein thrombosis, the fibrinolytic system was studied before and after a 10-minute venous occlusion. Blood was collected at least 3 months after the last acute episode, and PAI-1 antigen and activity, as well as tissue-type plasminogen activator (t-PA) antigen, urokinase-type plasminogen activator (u-PA) antigen, and fibrinolytic activity were measured in these samples. During venous stasis, plasminogen activator inhibitor (PAI) activity decreased in almost all patients (81 of 83), from a median value of 8.2 to 2.9 U/mL (P less than .001, Wilcoxon signed-rank test). Because PAI-1 antigen augmented from a median value of 16 to 19.2 ng/mL (P less than .001), the decline in PAI activity was attributed to an increase in t-PA antigen from a median value of 10 to 21.7 ng/mL (P less than .001). Neutralization of PAI activity thus reflects the patient's capacity to overcome basal inhibitory potential through t-PA release. Based on residual PAI activity after 10-minute stasis, patients were classified as good or bad responders (PAI activity below detection limit, ie, less than or equal to 1.0 and greater than 1.0 U/ml, respectively). Good responders had a significantly higher fibrinolytic response after stasis than bad responders (median euglobulin clot lysis time 60 v 180 minutes; dilute whole blood clot lysis time 60 v 120 minutes; fibrinolytic activity on fibrin plates 7.7 v 0 U/mL). Furthermore, good responders, as compared with bad responders, had higher t-PA release (median 16.5 v 11.5 ng/mL), lower basal PAI activity (median 4.8 v 11.2 U/mL), and lower basal PAI-1 (median 11 v 21 ng/mL) and u-PA antigen (median 7.9 v 9.0 ng/mL, P less than .02). Hypofibrinolysis, as defined by the inability of released t-PA to overcome PAI-1 basal inhibitory potential, was observed in 45 of 83 patients (54%) and resulted either from an insufficient release of t-PA or from an increased basal PAI activity.     

Changes in the fibrinolytic components of cultured human umbilical vein endothelial cells induced by endotoxin, tumor necrosis factor-alpha and interleukin-1alpha

BACKGROUND AND OBJECTIVE: Vascular fibrinolysis, a major natural defense mechanism against thrombosis, is a highly regulated process. The aim of this study was to evaluate the effect of endotoxin, tumor necrosis factor-alpha (TNFalpha) and interleukin-1alpha (IL-1alpha), on the fibrinolytic potential of cultured human umbilical vein endothelial cells (HUVEC). DESIGN AND METHODS: Samples of stimulated conditioned media were collected over a period of 24 hours to determine: plasminogen activator (PA) and plasminogen activator inhibitor (PAI) activity, PAI-1 mRNA, tissue-type plasminogen activator (t-PA) antigen and urokinase-type plasminogen activator (u-PA) antigen. RESULTS: Similar changes were observed after endotoxin and cytokine stimulation: there was a significant increase of PAI activity (p<0.01), starting at 6 hours, which remained 24 hours after stimulation. PAI-1 mRNA also showed an important rise with these agents, although cytokines induced an earlier and more intense inhibitor response (up to 6-fold increase). PA activity increased significantly at 6 hours (p<0.01) to drop at 24 hours and was mainly related to the presence of u-PA. INTERPRETATION AND CONCLUSIONS: We conclude that endotoxin,+TNFalpha and IL-1alpha induce profound alterations in the fibrinolytic potential of HUVEC, characterized by an initial rise of activators (u-PA) followed by a strong increase of PAI-1. These changes may be of pathophysiologic significance for thrombosis and inflammatory reactions.     

Increased tissue-type plasminogen activator: a facade in the fibrinolytic system in type 2 diabetes

To determine whether the increased concentration in blood of total tissue-type plasminogen activator (t-PA), accompanying increased plasminogen activator inhibitor type-1 (PAI-1) seen with diabetes reflects increased free t-PA and hence activity or simply increased t-PA complexed with PAI-1, we measured each in 39 people with type 2 diabetes. The increased total t-PA occurred in the absence of an increase in t-PA activity detectable in venous blood samples despite the increase in the concentration of total t-PA protein. BACKGROUND: Type 2 diabetes is known to be associated with increased concentrations in blood of total tissue-type plasminogen activator (t-PA) [free t-PA plus t-PA complexed with plasminogen activator inhibitor type-1 (PAI-1)] and PAI-1. This study was performed to determine whether the increased total t-PA is indicative of increased t-PA activity attributable to free t-PA or whether it is simply a manifestation of compensatory processes maintaining t-PA activity detectable in venous blood samples at normal or subnormal levels. METHODS: Total t-PA, free t-PA, and PAI-1 antigen and activity were measured by enzyme-linked immunosorbent assay and immunofunctional chromogenic substrate kinetic assays in peripheral venous blood samples obtained under fasting conditions from 39 people with type 2 diabetes. RESULTS: The results indicate that both PAI-1 and total t-PA antigen concentrations were increased in association with diabetes but the concentration of free t-PA and hence the t-PA activity was not. Thus, the increased total t-PA is a facade potentially masking the impaired fibrinolytic system activity associated with type 2 diabetes. CONCLUSIONS: The results indicate that the increase in the concentration of total t-PA protein in blood of people with diabetes occurs in the absence of an increase in free t-PA. Hence t-PA activity detectable in peripheral venous blood samples does not increase and may even decline modestly.     

Coagulation and fibrinolysis in preeclampsia and neonates.

Coagulation and fibrinolysis were determined in 67 Indonesian women admitted to the University Hospitals for delivery in Medan. They were diagnosed to be at term gestation (mean 39.3 +/- 1.1 weeks) with moderate and severe preeclampsia (n=32) and in labor, and 8 had preterm labor (gestation mean 33.5 +/- 2.6 weeks). Twenty-seven normal pregnant women in labor (gestation mean 39.7 +/- 1.0 weeks) served as controls. Cord blood from 23 neonates from normal pregnancy and 31 neonates from preeclampsia was also evaluated. Preeclamptic women in labor showed further enhanced coagulation activation (F(1+2)) with raised urokinase-like plasminogen activator (u-PA) activity and reduced plasminogen activator inhibitor-2 (PAI-2) levels. In preterm preeclampsia, significantly reduced antithrombin III (ATIII) and PAI-2 levels with further elevated tissue-type PA (t-PA) antigen and plasminogen activator inhibitor-1 (PAI-1) antigen were seen compared to normal pregnancy. These would suggest a state of enhanced thrombin generation with elevated fibrinolytic/inhibitor proteins in preterm preeclampsia. The reduced PAI-2 levels seen in preeclampsia have been suggested to be associated with reduced placental function. Neonates born to mothers of either normal pregnancy or preeclampsia at term showed similar hemostatic changes with reduced fibrinogen, ATIII, t-PA, u-PA antigen, PAI-1 levels, and coagulation activation compared to their respective maternal plasma levels. No significant differences in hemostatic parameters studied between the neonates of both cohorts were seen, and this would suggest that the neonates were protected from the adverse effects of preeclampsia and their hemostatic system was physiologically balanced.     

Plasminogen activators in experimental colorectal neoplasia: a role in the adenoma-carcinoma sequence?

An important step in the transition from adenomatous polyp to invasive carcinoma is the degradation of the epithelial basement membrane. By the generation of plasmin, plasminogen activators may play an important role in regulating the extracellular protease activity required for this event to occur. The production of biofunctional urokinase and of tissue plasminogen activator was therefore investigated in the dimethylhydrazine induced rat model of colorectal neoplasia. Both adenomatous polyps (p values less than 0.001) and colorectal carcinomas (p values less than 0.001) were demonstrated to produce a significant excess of both urokinase and tissue plasminogen activator when compared with macroscopically normal colon. There was, however, no increased production of either enzyme by macroscopically normal preneoplastic colon when compared with control colon. This enhanced capacity of colorectal tumours to produce plasminogen activators and generate plasmin is thus a feature of both the premalignant as well as the malignant phenotype. These enzymes may contribute to the malignant potential of adenomatous polyps and to the invasive capacity of established carcinomas.     

Functional characteristics of receptor-bound urokinase on human monocytes: catalytic efficiency and susceptibility to inactivation by plasminogen activator inhibitors

We compared urokinase-type plasminogen activator (u-PA) in fluid phase and u-PA bound with its receptor on human blood monocytes with respect to proteolytic activity and susceptibility to inactivation by the plasminogen activator inhibitors PAI-1 and PAI-2. Receptor-bound u-PA is catalytically twice as efficient as fluid-phase u-PA. Fluid-phase u-PA is susceptible to rapid inhibition by PAI-1 and PAI-2 at an estimated PAI:u-PA molar ratio of 2:1. In contrast, u-PA bound to endogenously occupied receptors is inhibited by PAI-2 only at PAI:u-PA molar ratios of 20:1, but is not inhibited by PAI-1, u-PA/PAI-1 and u-PA/PAI-2 complexes bind to the receptor with a tenfold lower affinity than u-PA itself. Thus, competition of u-PA/PAI complexes with fluid-phase u-PA for binding to the receptor is unlikely to affect the overall plasminogen activator activity of the monocyte. These findings demonstrate that the activity of receptor-bound u-PA can be modulated by PAI-2, but not by PAI-1, to adjust the cell's proteolytic activity to different local situations.     

Tissue-type plasminogen activator activity in morphologically normal tissues adjacent to gastrointestinal carcinomas is associated with the degree of tumor progression

Purpose: To investigate whether the level of plasminogen activator (PA) activity assayed in gastrointestinal carcinomas and the “morphologically normal tissues” adjacent to them is associated with the degree of tumor progression. Methods: Tumor and “normal tissues” were obtained from gastrointestinal surgical samples to assess urokinase-type (u-PA) and tissue-type plasminogen activator (t-PA) activities by radial caseinolytic assay and the expression of PA inhibitor-1 (PAI-1) by ELISA. We compared the PA system between the tumor and “normal tissues” and we investigated the existence of correlations between: (a) PA production in the tumor and “normal tissues”, (b) different components of the PA system, and (c) PA system and the degree of tumor progression. Results: (1) Total PA activity, u-PA activity and PAI-1 expression are significantly higher in tumor than in “normal tissues”, whereas t-PA activity does not differ between them. (2) Total PA activity mainly correlates with u-PA activity in tumor tissues and similarly with u-PA and t-PA activities in “normal tissues”. (3) There is a significant association between t-PA activity in tumor and “normal tissues” and the degree of tumor progression. Conclusions: “Morphologically normal tissues” adjacent to carcinomas present abnormal t-PA activity that is associated with the degree of tumor progression. Assaying of this activity could be useful as a predictive parameter.     

Acute inflammatory intestinal vascular lesions and in situ abnormalities of the plasminogen activation system in Crohn's disease.

OBJECTIVES: The distribution of the intestinal vascular lesions and their relation with the fibrinolysis process are poorly known in Crohn's disease (CD). The mediators of the plasminogen activator system, namely urokinase-type plasminogen activator (u-PA), tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type-1 (PAI-1), are a key complex involved in fibrinolysis. The aims of this study were: (1) to further define vascular lesions and their distribution in the intestine; and (2) to study concomitantly the qualitative in situ expression and the levels of u-PA, t-PA and PAI-1 in the ileum of patients with CD. PATIENTS AND METHODS: Histological, immunohistochemical and ultrastructural studies of vascular lesions in the resected ileum of 27 patients with CD were performed and compared with 36 control patients. Levels of u-PA, t-PA and PAI-1 measured by ELISA methods were compared in healthy and inflamed ileal tissues of 17 patients with CD. RESULTS: Acute vascular lesions involving mainly serosal venules and capillaries were present in 63% of patients with CD vs 3/36 controls and were associated with PAI-1 expression. They were prominent on the mesenteric border beneath macroscopically normal mucosa. In contrast, chronic vascular lesions were present in all layers beneath mucosal ulcerations, where a significant increase of PAI-1 levels was found. CONCLUSIONS: These results suggest that vascular involvement associated with abnormalities of PAI-1 expression is an early and widespread event in CD. Their prominence on the mesenteric border might explain the characteristic location of CD ulceration along the mesenteric margin.     

Effect of a Change in the Sleep/Wake Cycle on the Diurnal Variation of Fibrinolytic Parameters

The mechanism underlying the circadian rhythm of fibrinolysis is not well understood. To evaluate the influences of wakefulness and of the intrinsic circadian rhythm on fibrinolytic activity, we examined diurnal changes (8:00 am vs. 8:00 pm) in plasminogen activator inhibitor-1 (PAI-1) activity, tissue plasminogen activator (t-PA) antigen levels, and t-PA activity, as well as in plasma serum cortisol levels, in 10 healthy males (21 ± 2 years) for two consecutive days. On the first day, subjects remained awake all day and night. They slept during the daytime (8:30 am to 5:30 pm) on the following day. PAI-1 activity and cortisol levels were significantly decreased, and t-PA activity tended to increase during the daytime on the first day. On the morning following overnight wakefulness, PAI-1 activity and cortisol levels did not return to the levels of the previous morning. On the second day, the afternoon decrease in PAI-1 activity, but not cortisol levels, was still observed, although its magnitude was substantially attenuated. No significant diurnal changes were observed in the levels of t-PA antigen throughout the study period. These findings suggest that the diurnal variation of fibrinolytic activity may be governed by an intrinsic circadian rhythm of PAI-1, which can be modified by a change in the time of wakefulness.