High plasminogen activator inhibitor and tissue plasminogen activator levels in plasma precede a first acute myocardial infarction in both men and women: evidence for the fibrinolytic system as an independent primary risk factor

BACKGROUND: In patients with established ischemic heart disease, prospective cohort studies have indicated that plasminogen activator inhibitor (PAI-1), the inhibitor of the fibrinolytic system, may predict cardiovascular events. So far, there have been no primary prospective studies of PAI-1. METHODS AND RESULTS: The aim of the present study was to test whether plasma levels of PAI-1, tissue-type plasminogen activator (tPA), von Willebrand factor (vWF), and thrombomodulin (TM) could predict the occurrence of a first acute myocardial infarction (AMI) in a population with high prevalence of coronary heart disease by use of a prospective nested case-control design. Mass concentrations of PAI-1 and tPA were significantly higher for the 78 subjects who developed a first AMI compared with the 156 references matched for age, sex, and sampling time; for tPA, this increase was independent of smoking habits, body mass index, hypertension, diabetes, cholesterol, and apolipoprotein A-I. The ratio of quartile 4 to 1 for tPA was 5.9 for a patient to develop a first AMI. The association between tPA and AMI was seen in both men and women. Increased levels of vWF were associated with AMI in a univariate analysis. High levels of TM were associated with AMI in women but not in men. CONCLUSIONS: The plasma levels of PAI-1, tPA, and vWF are associated with subsequent development of a first AMI; for PAI-1 and tPA, this relation was found in both men and women. For tPA but not for PAI-1 and vWF, this association is independent of established risk factors.     

Renal tissue angiotensins during converting enzyme inhibition of angiotensin I in spontaneously hypertensive rat

To compare the effects of an angiotensin-converting enzyme inhibitor on circulating and tissue renin-angiotensin system, we measured different renin-angiotensin system parameters during the first day of treatment (Day 1) as well as after two weeks of treatment (Day 14). Ramipril was given orally once daily to adult male spontaneously hypertensive rats. Renin activity, angiotensin-converting enzyme activity and levels of angiotensin I and angiotensin II in the plasma, renal cortex and renal medullar were assessed at Day 1 and Day 14 of the treatment. In the plasma, both renin activity and angiotensin I increased 10 to 15 fold one to four hours after acute as well as at Day 14 of ramipril treatment and then returned to basal values within 24 hours. Plasma angiotensin II levels were not significantly decreased at Day 1 or Day 14. The decrease in the angiotensin II/angiotensin I ratio suggested a sustained inhibition of plasma angiotensin-converting enzyme at Day 14. In the renal cortex and medulla, a clearly different pattern was observed: in ramipril treated rats, renin activity in the renal cortex and medulla did not change at Day 1 but at Day 14 we observed a slight and sustained increase in renin activity. Despite very high basal levels of renin activity, angiotensin I levels in the renal cortex were comparable to those in the plasma. The angiotensin I level increased only one-fold one hour after ramipril intake at Day 1 and Day 14. This suggests that angiotensinogen may have a limiting role in the synthesis of angiotensin I in the kidney. Angiotensin II levels were slightly higher in the renal cortex and medulla than in the plasma suggesting local synthesis of the peptide. In the kidney, angiotensin II levels decreased one and four hours after the acute or prolonged ramipril treatment and the angiotensin II/angiotensin I ratio was reduced at the same time. Our results show that the responses of the plasma and kidney components of the renin-angiotensin system to angiotensin-converting enzyme inhibition are different in the plasma and the kidney suggesting that the circulating and tissue renin-angiotensin system are at least in part independent.     

Dimethyl sulfoxide-induced apoptosis in human leukemic U937 cells

The effects of two oral contraceptives, containing gestodene and either 20 micrograms or 30 micrograms ethinylestradiol, on hemostatic parameters was investigated in a six-month randomized study involving a total of 40 healthy women between the ages of 18 and 30 years. A large number of hemostatic parameters were measured, which were categorized as either pro-coagulatory, anti-coagulatory, profibrinolytic, anti-fibrinolytic or indicative of fibrin turnover. Additionally, tissue plasminogen activator (t-PA) and plasminogen activator inhibitor (PAI-1) were measured before and after venous occlusion and delta and ratio values calculated. Pro-coagulatory factors as well as reaction products reflecting in vivo coagulatory activity (thrombin-antithrombin III complex, prothrombin fragment 1 + 2) were found to increase. Among the anti-coagulatory parameters, only protein S concentration and protein S activity decreased, most notably in the 30 micrograms EE group. There was a corresponding increase in fibrinolytic activity reflected by reaction products of in vivo fibrinolysis (plasmin-antiplasmin 2-complex, fibrin-degradation products). Measurement of t-PA and PAI-1, before and after venous occlusion, revealed that the fibrinolytic response was more pronounced in the 20 micrograms EE group. There was also an increase in the threshold of fibrinolytic inhibition (ratio PAI-1) in both groups, which was less pronounced in the 20 micrograms EE group. Apart from isolated measurements, all parameters remained within their normal ranges and values returned to baseline in the follow-up cycle. It is concluded that both preparations had a balanced effect on the hemostatic system stimulating both pro-coagulant and fibrinolytic activity. No statistically significant differences were observed between the two groups; however, there was a trend towards greater fibrinolytic capacity in the 20 micrograms EE group.     

Fibrinolytic factors and the risk of myocardial infarction or sudden death in patients with angina pectoris. ECAT Study Group. European Concerted Action on Thrombosis and Disabilities

BACKGROUND: Disturbances of the fibrinolytic system that lead to decreased removal of fibrin deposits may be important risk factors for coronary thrombosis. There is as yet no consensus on the prognostic value of fibrinolytic parameters, which may be attributed in part to the choice of confounding variables controlled for. METHODS AND RESULTS: The ECAT study is a prospective multicenter study of 3043 patients with angina pectoris followed for 2 years. Baseline measurements included 10 fibrinolytic variables. The results were analyzed in relation to the subsequent incidence of myocardial infarction or sudden coronary death. They are presented before and after adjustment for clusters of confounding variables that are markers of different mechanisms: insulin resistance (body mass index, triglyceride, and HDL cholesterol), inflammation (fibrinogen and C-reactive protein), and endothelial cell damage (von Willebrand factor). An increased incidence of events was associated with higher baseline concentrations of tissue plasminogen activator (TPA) antigen (P = .0002), plasminogen activator inhibitor-1 (PAI-1) activity (P = .02), and PAI-1 antigen (P = .001). The associations of PAI-1 activity and PAI-1 antigen with risk of events disappeared after adjustment for parameters reflecting insulin resistance but were not affected by other adjustments. TPA antigen was affected to a similar extent by adjustment for parameters reflecting insulin resistance. Inflammation, or endothelial cell damage, but the risk association disappeared only after combined adjustments. CONCLUSIONS: The prognostic role of PAI-1 in predicting coronary events is related principally to insulin resistance, whereas that of TPA antigen could be explained only by its relationship with different mechanisms, including insulin resistance, inflammation and endothelial cell damage.     

Initiation of a fibrinolytic system in hepatic resection: the roles of tissue-type plasminogen activator and plasminogen activator inhibitor-1

The factors related to the initiation of fibrinolysis, especially with regard to the tissue-type plasminogen activator (tPA) and the plasminogen activator inhibitor-1 (PAI-1), were investigated in 15 patients who underwent hepatic resection, and the findings were compared between those with normal livers and those with diseased livers. It was found that tPA increased before hepatic division, whereas PAI-1 increased after hepatic division and reached a peak immediately following the operation. Plasminogen decreased during hepatectomy, reaching its lowest point on postoperative day 1, and increasing later. Decreased levels of both plasminogen and the alpha 2-plasmin inhibitor were considered to be partly due to plasmin formation in the blood. Patients with a diseased liver tended to have higher intraoperative values of euglobulin lysis activity and higher postoperative values of plasminogen activator, but significantly lower postoperative values of alpha 2-plasmin inhibitor than those with a normal liver. The results of this study suggest that activation of the fibrinolytic system occurs both during hepatectomy and in the early postoperative period, and that patients with a diseased liver are prone to develop hyperfibrinolysis during hepatectomy. Moreover, the increased levels of both tPA and PAI-1 can serve as one of the most sensitive markers for the vital reaction against surgical stress.     

Tissue markers as predictors of postoperative adhesions

BACKGROUND: Postoperative adhesion formation has been associated with a decreased capacity to degrade intra-abdominally deposited fibrin. Adhesions, once lysed, have a high propensity for reformation. This study tested the hypothesis that patients with a high propensity for adhesion formation as well as adhesion tissue had a reduced fibrinolytic capacity. METHODS: Peritoneal biopsies were taken during abdominal surgery from 21 patients who had previously undergone operation; previously formed adhesion tissue was sampled from ten of these patients. Adhesion formation was scored. The fibrinolytic capacity of peritoneum was determined in tissue extracts. RESULTS: At the time of opening of the abdominal cavity, levels of plasminogen activator inhibitor (PAI) type 1 (P = 0.009) and tissue-type plasminogen activator (tPA)/PAI complex (P = 0.008) were increased in peritoneal samples from patients with severe adhesions compared with those in samples from patients with less severe adhesions. Adhesion tissue similarly had reduced fibrinolytic capacity as judged by a decrease in tPA activity (P = 0.005) and an increase in PAI-1 level (P = 0.01), reflected in an increased level of tPA/PAI complex (P = 0.008) compared with unaffected peritoneum. CONCLUSION: These observations demonstrate reduced fibrinolytic capacity in peritoneal tissue in patients with a greater propensity for development of adhesions and likewise in adhesion tissue. This suggests that components of the fibrinolytic system may be used as markers of an increased risk of adhesion development.     

Concentration of endogenous tPA antigen in coronary artery disease: relation to thrombotic events, aspirin treatment, hyperlipidemia, and multivessel disease

Tissue plasminogen activator (tPA) is the major plasminogen activator responsible for dissolving blood clots found in blood vessels. However, elevated concentrations of tPA antigen were found to be related to adverse events in patients with coronary artery disease (CAD). Considerable controversy about the significance of these results exists. The goal of this cross-sectional study was to identify independent determinants for tPA antigen concentrations in patients with CAD, to possibly clarify the above paradoxical relationship. The baseline tPA antigen concentrations of 366 patients with angiographic evidence of coronary sclerosis were determined. Univariate analysis showed that age (P=0.013), angiographic extent of disease (P<0.001), presence of angina at rest (P<0.001), diabetes mellitus (P=0.004), hypercholesterolemia (P=0. 045), hypertriglyceridemia (P=0.015), and chronic intake of nitrates (P<0.001) were significantly and positively related to tPA antigen concentration, while the chronic intake of aspirin was inversely related to tPA antigen (P<0.001). In addition, plasminogen activator inhibitor type 1 (PAI-1) activity was found to be significantly and positively associated with tPA antigen concentration (P<0.001). A multivariate analysis identified chronic low-dose aspirin therapy (P<0.001), PAI-1 activity (P<0.001), hypertriglyceridemia (P=0.005), the type of angina (P=0.026), multivessel disease (P=0.041), and hypercholesterolemia (P=0.043) as significant and independent determinants of tPA antigen. While hypertriglyceridemia and hypercholesterolemia both are related to the underlying disease, the type of angina and the number of involved vessels are linked to the severity and extent of disease, and all of them are indicators of a prothrombotic state found during the progression of CAD. In contrary, low-dose aspirin rather would decrease the likelihood of thrombotic events. The relation of tPA antigen to PAI-1 activity furthermore underlines the relation between tPA antigen concentration and a prothrombotic state. Therefore, the positive or-in case of aspirin therapy-negative correlation of these parameters with tPA antigen concentration would indicate that thrombus formation and simultaneous endothelial cell activation might be major determinants for tPA antigen concentration in CAD.     

The fibrinolytic effects of intermittent pneumatic compression: mechanism of enhanced fibrinolysis

BACKGROUND AND OBJECTIVES: Intermittent pneumatic compression (IPC) is an effective form of deep vein thrombosis prophylaxis for general surgery patients. The antithrombotic effect of IPC is thought to be the result of increased venous velocity and stimulation of endogenous fibrinolysis. However, the mechanism of enhanced fibrinolytic activity and the relative effects on normal and postthrombotic veins have not been defined. The purposes of this study are 1) to quantify changes in fibrinolytic activity with IPC; 2) to study the mechanism of fibrinolytic enhancement with IPC; and 3) to evaluate whether postthrombotic patients have the same capacity for fibrinolytic enhancement with IPC as do normal subjects. METHODS: Twelve volunteers (6 normal and 6 postthrombotic) had 5 IPC devices applied for 120 minutes in random fashion, 1 per week x 5 weeks. The devices included single-chamber, sequential, foot, calf, and long-leg compression. Subjects had an indwelling antecubital venous cannula placed for blood drawn at baseline, 60, 120, and 180 minutes after IPC devices were applied. Global fibrinolytic activity (euglobulin fraction, fibrin plate assay), tissue plasminogen activator (tPA) antigen (Ag) and activity (Act), plasminogen activator inhibitor-1 (PAI-1) Ag and Act, alpha-2-antiplasmin-plasmin complexes, and von Willebrand factor (vWF) antigen were assayed. RESULTS: A striking elevation in fibrinolytic activity was noted at 180 minutes with all devices in normal subjects and postthrombotic patients (p = 0.01-0.0001); however, baseline and stimulated fibrinolytic activity was attenuated in postthrombotic patients (<0.03). The tPA-Act increased only in normal subjects (3.8 +/- 1.9%) (p = 0.057), despite a decrease in plasma tPA-Ag, which was observed in both normal subjects (-12.4 +/- 3.8%) (p = 0.009) and patients (-17.2 +/- 3.1%) (p = 0.001). PAI-1-Ag decreased in both normal subjects (-13.4 +/- 3.8%) (p = 0.007) and patients (-12.0 +/- 3.1%) (p = 0.013) with a marked reduction in PAI-1-Act in both normal subjects (p = 0.003) and patients (p = 0.004). There were no changes in vWF, and alpha-2-antiplasmin-plasmin complexes increased only in postthrombotic patients (p = 0.021). CONCLUSIONS: Stimulation of endogenous fibrinolytic activity occurs after IPC, both in normal subjects and postthrombotic patients; however, baseline and overall fibrinolytic response in postthrombotic patients is reduced. The mechanism of increased fibrinolytic activity is likely because of a reduction in PAI-1, with a resulting increase of tPA activity.     

Small GTP-binding proteins in the brain-corpus cardiacum-corpus allatum complex of the silkworm, Bombyx mori: involvement in the secretion of prothoracicotropic hormone

BACKGROUND: Proteolysis, modulated in part by intramural fibrinolytic system proteins and their inhibitors, appears to influence vascular smooth muscle cell (SMC) migration and proliferation and remodeling of extracellular matrix (ECM). Alterations of fibrinolysis in circulating blood and of proteolysis within vessel walls in experimental animals and patients with diabetes have been associated with accelerated vascular disease. Hyaluronan, a prominent component of ECM in normal vessels, is increased in the tunica media of macroscopically normal arterial vessels from patients with type 2 diabetes. OBJECTIVE: To determine whether hyaluronan alters the expression of the fibrinolytic system protein, plasminogen activator inhibitor type-1 (PAI-1), in human vascular SMCs, thereby potentially accelerating vascular disease in patients with type 2 diabetes. METHODS: Urokinase-type and tissue-type plasminogen activators (uPA and tPA) and PAI-1 were assayed in vascular SMC conditioned media and in cell lysates, using enzyme-linked immunosorbent assay and western blotting. RESULTS: Hyaluronan increased the 24-h release of PAI-1 into conditioned media in a concentration-dependent and time-dependent manner (1.8-fold compared with control with 1 mg/ml hyaluronan; n = 9, P < 0.01). Although the accumulation of uPA in conditioned media tended to increase also, uPA content was reduced in cell lysates (64% of control with 0.1 mg/ml hyaluronan at 24 h; n = 9, P < 0.01) without any change in PAI-1. Concentrations of tPA in conditioned media and cell lysates were unchanged. Digestion of hyaluronan with hyaluronidase (50 turbidity reducing units (TRU)/ml) or exposure of the smooth muscle cells to antihuman CD44 antibody (1 microgram/ml) that binds to the hyaluronan cell surface receptor obviated the effects of hyaluronan. CONCLUSION: Our results indicate that increases in hyaluronan increase vascular SMC expression of PAI-1, a phenomenon that may alter the balance between proteolysis and its inhibition in vessels of patients with type 2 diabetes, thereby contributing to the acceleration of macroangiopathy.     

Evaluation of nutritional status in persons with spinal cord injury: a prerequisite for successful rehabilitation

A minority of patients with acute pulmonary embolism (PE) show failure of resolution when assessed by serial ventilation/perfusion (V/Q) radionuclide lung imaging. The fibrinolytic systems were studied in six such patients (group I), and in 11 patients in whom PE had resolved (group II), together with 17 healthy control subjects. Assays of the fibrinolytic system included euglobulin clot lysis times (ECLT), tissue plasminogen activator (tPA), and plasminogen activator inhibitor-1 (PAI-1). Euglobulin clot lysis times were not prolonged in the unresolved PE group, but were significantly longer in patients in group II when compared to control subjects (P < 0.03). This could not be explained either on the basis of tPA levels, which were higher in group II when compared to group I (P < 0.05) and control subjects (P < 0.02), or on the basis of PAI-1 levels which did not differ significantly between the three groups. Our inability to demonstrate derangements of fibrinolysis in the patients with unresolved PE makes defective fibrinolysis an unlikely aetiological factor in the persistence of thrombosis in these patients.     

Effects of experimental desmotomy on material properties and histomorphologic and ultrasonographic features of the accessory ligament of the deep digital flexor tendon in clinically normal horses

OBJECTIVE: Acute physical and psychological stressors affect blood coagulation and fibrinolysis, but little is known about hemostatic factors associated with chronic psychological stress. Prolonged psychological stress may end in a state of vital exhaustion, which has been shown to be a risk factor for first myocardial infarction and recurrent events after coronary angioplasty. The present study tested the hypothesis that vital exhaustion resulting from chronic psychological stress is associated with impaired fibrinolytic capacity and increased coagulation factors. METHODS: On the basis of a validated questionnaire and subsequent structured interview, a well-defined group of otherwise healthy exhausted men was recruited (N = 15) and compared with age-matched not-exhausted controls (N = 15). Fibrinolytic measures included tissue plasminogen activator (TPA) antigen and plasminogen activator inhibitor (PAI-1) activity, and as coagulation factors we examined factors VIIc, factor VIIIc, and fibrinogen. Control variables were: blood pressure, smoking status, triglycerides, cholesterol, and standard hematological measures. Samples were collected twice to correct for intraindividual fluctuations. Statistical analyses were performed using 2 x 2 mixed model analysis of variance with subsequent univariate testing. RESULTS: Vital exhaustion was associated with significantly elevated levels of PAI-1 activity (p = .023). The higher PAI-1 activity in exhausted subjects (median = 13.0 U/ml vs. 6.0 U/ml) was not accounted for by smoking status or serum lipids. No significant differences were observed in TPA antigen, factor VIIc, factor VIIIc, and fibrinogen. The groups did not differ in blood pressure, smoking status, triglycerides, cholesterol, or standard hematological measures. CONCLUSION: These data suggest a reduced fibrinolytic capacity in exhausted individuals. Therefore, the relationship between vital exhaustion and risk of myocardial infarction may be mediated in part by an imbalance between blood coagulation and fibrinolysis.     

Secretion of cardiac plasminogen activator during hypoxia-induced right ventricular hypertrophy

In the circulation, fibrinolytic activity is determined to a large degree by the relative levels of tissue plasminogen activator (tPA) and its major inhibitor (PAI-1). Vascular beds in different organs secrete tPA and PAI-1 into the circulation, and the total secretory rate of each protein is balanced by its half-life in the bloodstream. We are testing the hypothesis that in the heart, ventricular hypertrophy will alter the rates of formation of tPA and/or PAI-1 and the rates of their release into the cardiac vasculature. In this study, we have examined the effects of continuous hypoxia on PA activity in extracts of rat heart ventricles, on the activity secreted into the cardiac vasculature of perfused hearts, and on the levels of mRNAs for tPA and PAI-1. Rats were subjected to hypobaric hypoxia at 0.5 atm for 1-21 days. The treatment caused polycythemia within 1-3 days, and right ventricular hypertrophy by 3 days. PA activity in extracts of both right and left ventricles was significantly elevated after 3 days of hypoxia, continued to increase for 4 additional days, and remained elevated for 3 weeks. The actions of inhibitors of urokinase and tPA indicated that the PA activity in heart extracts was exclusively tPA. Fibrin zymography confirmed that result. The mRNAs for tPA and for PAI-1 were elevated after 1 day of hypoxia and then returned to near control levels on days 2 and 3. After 7 days, hearts from hypoxic rats secreted more tPA activity into perfusates than did hearts from controls. The difference in secretory rates was proportional to the differences in the levels of tPA in the corresponding heart extracts.     

Atherogenic, hemostatic, and other potential risk markers in subjects with previous isolated myocardial infarction compared with long-standing uncomplicated stable angina

BACKGROUND: Several atherogenic, hemostatic, inflammatory, and genetic parameters and markers have been implicated as risk factors in coronary artery disease, although whether they are risk factors for acute as opposed to chronic coronary disease is unclear. METHODS AND RESULTS: Fifty subjects with an isolated myocardial infarction >3 months previously were compared with 50 subjects with a minimum 3-year history of stable angina, documented coronary artery disease, normal electrocardiogram and normal ventricular wall motion, and no episode suggesting infarction or unstable angina. Biologic variables analyzed included apolipoprotein B (apo B), lipoprotein (a), C-reactive protein (CRP), fibrinogen, factor VII, tissue plasminogen activator (TPA) and inhibitor (PAI-1), thrombin-antithrombin (TAT), fragment 1+2 (F1+2), von Willebrand factor (vWF), activated protein C resistance, homocyst(e)ine, anticardiolipin antibodies, blood group, and the angiotensin-converting enzyme insertion/deletion (I/D) and angiotensin II receptor gene polymorphisms. There were no significant differences between the 2 groups for any of the variables studied, although fibrinogen and F 1+2 tended to be slightly higher in the angina group (P = .09 for each). These significant correlations were present: age with fibrinogen, homocyst(e)ine, and vWF; factor VII with apo B, homocyst(e)ine, and TPA; apo B with TPA and CRP; CRP with fibrinogen, TPA, PAI-1, and factor VII; fibrinogen with vWF. CONCLUSIONS: Examination of atherogenic, hemostatic, inflammation, and genetic variables in the clinically quiescent state permitted no distinction between subjects with a previous isolated myocardial infarction in contrast to those with long-standing uncomplicated stable angina, favoring the notion that acute coronary events occur at random on a varying background of atherosclerosis. The multiple correlations found among these variables also underscore their complex interaction in the atherosclerotic process.     

Relationship between serum angiotensin-converting enzyme activity and plasma plasminogen activator inhibitor activity in patients with recent myocardial infarction

BACKGROUND: An elevated level of angiotensin-converting enzyme (ACE) has been demonstrated to increase the risk of myocardial infarction. Plasminogen activator inhibitor (PAI) is the most important physiological inhibitor of tissue plasminogen activator in plasma. An elevated level of PAI has been reported to be associated with decreased fibrinolytic capacity and to constitute a marker of the risk for recurrent coronary thrombosis. METHODS: We measured the serum ACE activity and plasma PAI activity in 34 patients with recent myocardial infarction, and evaluated the correlation between these two values by linear regression analysis. We also administered captopril (37.5 mg/day) to 17 of these patients and placebo to the other 17 patients at random, and compared the changes in PAI activity and ACE activity in these two groups over a 1-month period. RESULTS: There was a significant correlation between the serum ACE activity and the plasma PAI activity at baseline in the patients (r = 0.498, P < 0.01). The captopril-treated patients showed significantly reduced PAI activity (P < 0.01), and a concomitant decrease in ACE activity. CONCLUSION: These results suggest that elevated ACE activity is associated with impaired fibrinolysis and that treatment with an ACE inhibitor improves the fibrinolytic function in patients with recent myocardial infarction. The results also suggest that the renin-angiotensin system plays a role in the increased risk of ischemic cardiovascular events when it is activated, and in the reduction of risk of recurrent myocardial infarction by ACE inhibition.     

Expression of fibrinolytic antigens in redistributed cardiac mast cells in auricular thrombosis

Recent data suggest that auricular thrombosis is associated with an increase and accumulation of mast cells (MC) in the subendothelial region of the upper endocardium. However, the molecular basis and the functional role of MC in this process are not known. In the current study, expression of fibrinolytic and antifibrinolytic antigens in human cardiac MC was analyzed by immunohistochemistry. MC were found to react with antibodies against tissue-type plasminogen activator (tPA) and urokinase receptor (uPAR/CD87), but not with antibodies against urokinase (uPA) or plasminogen activator inhibitors (PAI-1, PAI-2). Significant changes were observed when the phenotype of accumulated MC in the upper endocardium in patients with auricular thrombosis was compared with the phenotype of myocardial MC in the same patients or with MC in normal hearts. These redistributed MC stained less intensely with antibodies against tPA and chymase but retained their staining for tryptase and uPAR. Together, these data indicate that cardiac MC are a source of fibrinolytic antigens and that accumulation of MC in auricular thrombosis is associated with phenotypic changes of MC and loss of cellular tPA. It is hypothesized that MC and their products may play a role in endogenous fibrinolysis in auricular thrombosis.     

Coordinated expression of tissue-type plasminogen activator and plasminogen activator inhibitor type 1 during corpus luteum formation and luteolysis in the adult pseudopregnant rat

Proteolytic activity generated by the plasminogen activator (PA) system is associated with many biological processes. Using an adult pseudopregnant rat model, we have studied how two components of the PA system, tissue-type plasminogen activator (tPA) and plasminogen activator inhibitor type 1 (PAI-1), are expressed temporally and spatially during different developmental stages of the corpus luteum (CL). Northern blot analysis, in situ hybridization, in situ zymography, and fibrin overlay were used to analyze the expression and distribution of tPA and PAI-1 messenger RNA (mRNA) as well as PA activity in CL of different ages. We demonstrated that during the luteinization period (approximately days 1-2), tPA mRNA was highly and evenly expressed in newly formed CL, whereas PAI-1 mRNA was mainly detected in the central part of the same CL. In accordance with these findings, proteolytic activity generated by tPA was detected in the outer region of newly formed CL by in situ zymography. During the luteotropic period (approximately days 3-10), tPA mRNA expression was very low. PAI-1 mRNA expression was also low, but increased on day 10. As expected, proteolytic activity was very low during this period. During functional luteolysis (days 13-14) and subsequent structural luteolysis, tPA mRNA was elevated. PAI-1 mRNA was also expressed during this period. Moreover, the net PA activity, as determined by fibrin overlay, was relatively high during this period. Our studies indicate that tPA and PAI-1 are coordinately expressed in the CL, resulting in increased proteolytic activities during the luteinization and luteolytic periods. PA-mediated proteolysis may, therefore, play a role in both CL formation and luteolysis in rats.     

Distinction of endothelial cell growth and fibrinolytic activity between WKY/Izm and SHRSP/Izm in vitro

1. In the present study, endothelial cells (EC) growth and fibrinolytic activity of WKY/Izm and SHRSP/Izm were investigated in vitro. 2. EC were isolated from the thoracic aortas of WKY and SHRSP at the age of 8-9 weeks. Proliferative activities of EC were analysed with doubling time and DNA synthesis. Fibrinolytic activity was determined by tissue type plasminogen activator (tPA) and plasminogen activator inhibitor type 1 (PAI-1) activities in cultured medium. 3. SHRSP-EC growth rate was significantly greater than WKY-EC growth rate in doubling time. In the assay for DNA synthesis, 5-bromo-2'-deoxy uridine incorporation rate in SHRSP-EC was significantly increased compared with that in WKY-EC. 4. The tPA activity in cultured medium of WKY-EC was 2-fold greater than that of SHRSP-EC, while PAI-1 activities were nearly equal in them. 5. These physiological distinctions of EC of SHRSP/Izm from that of WKY/Izm with close genetic background could be contributory genetic factors to hypertension-related vascular diseases in SHRSP.     

Insulin sensitivity, insulin action, and fibrinolysis activity in nondiabetic and diabetic obese subjects

Because inconsistencies occur with regard to the relative contribution of insulin to the hypofibrinolysis characteristic of obesity and diabetes, we explored the relationship between insulin and fibrinolysis, assessing both insulin sensitivity and insulin action. Seventeen markedly obese subjects (body mass index [BMI], 34.0+/-1.6 kg/m2; 12 nondiabetic and five diabetic) were studied using the three-step euglycemic-hyperinsulinemic clamp technique. Since the circadian rhythm of the fibrinolytic system may obscure a true effect of insulin, variations in fibrinolysis parameters observed during the glucose clamp were compared with those occurring spontaneously because of the circadian rhythm. Compared with six normal-weight subjects (BMI, 21.0+/-0.9 kg/m2), all obese subjects exhibited basal hyperinsulinism (fasting plasma insulin, 16.0+/-1.4 v 9.8+/-1.3 microU/microL, P < .001; fasting plasma C-peptide, 1.4+/-0.2 v 0.5+/-0.2 ng/mL, P < .001), hypofibrinolysis (euglobulin lysis time [ELT], 378+/-29 v 222+/-31 minutes, P=.01; tissue plasminogen activator [tPA] antigen, 7.8+/-0.9 v 4.2+/-0.5 ng/mL, P=.04; plasminogen activator inhibitor type 1 [PAI-1] activity, 22.2+/-2.5 v3.9+/-0.6 AU/mL, P=.004), and marked insulin resistance (M value, ie, the maximal glucose disposal rate, 9.1+/-0.6 v 18.6+/-0.8 mg/(kg x min), P < .001). The M value correlated inversely with tPA antigen (r=-.46, P=.05). During insulin infusion, values for fibrinolysis parameters decreased, but were not different compared with variations due to the circadian rhythm. In conclusion, our findings together with previously reported data reinforce the idea that chronic hyperinsulinism is linked to hypofibrinolysis, but insulin does not seem to acutely regulate the fibrinolysis system.     

The mast cell as site of tissue-type plasminogen activator expression and fibrinolysis

Recent data suggest that mast cells (MC) and their products (heparin, proteases) are involved in the regulation of coagulation and fibrino(geno)lysis. The key enzyme of fibrinolysis, plasmin, derives from its inactive progenitor, plasminogen, through catalytic action of plasminogen activators (PAs). In most cell systems, however, PAs are neutralized by plasminogen activator inhibitors (PAIs). We report that human tissue MC as well as the MC line HMC-1 constitutively produce, express, and release tissue-type plasminogen activator (tPA) without producing inhibitory PAIs. As assessed by Northern blotting, highly enriched lung MC (>98% pure) as well as HMC-1 expressed tPA mRNA, but did not express mRNA for PAI-1, PAI-2, or PAI-3. The tPA protein was detectable in MC-conditioned medium by Western blotting and immunoassay, and the MC agonist stem cell factor (c-Kit ligand) was found to promote the release of tPA from MC. In addition, MC-conditioned medium induced fibrin-independent plasmin generation as well as clot lysis in vitro. These observations raise the possibility that MC play an important role in endogenous fibrinolysis.     

Lovastatin modulates in vivo and in vitro the plasminogen activator/plasmin system of rat proximal tubular cells: role of geranylgeranylation and Rho proteins

Interstitial fibrosis is one of the most deleterious events during the progression of renal deterioration after renal mass reduction. In vivo, hydroxymethylglutaryl CoA reductase inhibitors (HRI) were shown to reduce progression of glomerulosclerosis, but the mechanisms are still unclear. The present study investigates, in vivo, whether lovastatin, a potent HRI, was able to modulate the plasminogen-plasmin pathway, one of the most efficient systems involved in extracellular matrix remodeling, and characterizes in vitro the cellular mechanisms of these effects. Proximal tubules freshly isolated from rats treated for 2 d with lovastatin (4 mg/kg per d) showed increased tissue-type plasminogen activator (tPA) and urokinase (uPA) activities and antigens. Incubation with lovastatin (5 microM) of proximal tubules isolated from untreated rats induced an increase in tPA and uPA and a decrease in plasminogen activator inhibitor-1 (PAI-1) activities. In vitro, supernatants, cytosols, and membranes of renal proximal tubular cells in primary cultures had no detectable uPA activity, and lovastatin (0.1 to 10 microM) induced an increase in tPA and a decrease in PAI-1 activities and antigens. These effects were reversed by mevalonate and geranylgeranyl-pyrophosphate (GGPP) but not by farnesyl-pyrophosphate or LDL cholesterol. C3 exoenzyme, an inhibitor of the geranylgeranylated-activated Rho protein, reproduced the effect of lovastatin on tPA and PAI- activity and blocked its reversion by GGPP. The effect of lovastatin was associated with a disruption of cellular actin stress fibers, which was reversed by GGPP and reproduced by C3 exoenzyme. In conclusion, HRI can modify the fibrinolytic potential of proximal tubules, most likely via inhibition of geranylgeranylated Rho protein and disruption of the cytoskeleton. The resulting increase of proteolytic activity of tubular cells may serve to prevent extracellular matrix deposition and renal interstitial fibrosis.