Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P00750 (PLA)
 16,800 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Homozygous plasminogen activator inhibitor-1 (PAI-1)-deficient (PAI-1-/-) mice were generated by homologous recombination in D3 embryonic stem cells. Deletion of the genomic sequences encompassing the transcription initiation site and the entire coding regions of murine PAI-1 was demonstrated by Southern blot analysis. A 3.0-kb PAI-1-specific mRNA was identified by Northern blot analysis in liver from PAI-1 wild type (PAI-1+/+) but not from PAI-1-/- mice. Plasma PAI-1 levels, measured 2-4 h after endotoxin (2.0 mg/kg) injection were 63 +/- 2 ng/ml, 30 +/- 10 ng/ml, and undetectable (< 2 ng/ml) in PAI-1+/+, heterozygous (PAI-1+/-) and PAI-1-/- mice, respectively (mean +/- SEM, n = 4-11). PAI-1-specific immunoreactivity was demonstrable in kidneys of PAI-1+/+ but not of PAI-1-/- mice. SDS-gel electrophoresis of plasma incubated with 125I-labeled recombinant human tissue-type plasminogen activator revealed an approximately 115,000-M(r) component with plasma from endotoxin-stimulated (0.5 mg/kg) PAI-1+/+ but not from PAI-1-/- mice, which could be precipitated with a polyclonal anti-PAI-1 antiserum. PAI-1-/- mice were viable, produced similar sizes of litters as PAI-1+/+ mice, and showed no apparent macroscopic or microscopic histological abnormalities.
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PMID:Plasminogen activator inhibitor-1 gene-deficient mice. I. Generation by homologous recombination and characterization. 825 28

The capacity of macrophages to influence directly and indirectly fibrinolytic processes in atherosclerosis was studied using macrophages isolated from atherosclerotic plaques of patients undergoing surgical repair of distal aortic and femoral arteries. These cells were characterized by their morphology, adherence, esterase positivity, and expression of CD14 antigen. Production of plasminogen activator inhibitor type-1 (PAI-1) by plaque macrophages (6.7 +/- 2.7 ng/10(5) cells/24 hours [mean +/- SEM]) was significantly greater than PAI-1 production by blood monocytes isolated simultaneously from the same patients (1.8 +/- 1.5 ng/10(5) cells/24 hours). Production of tissue type plasminogen activator and urokinase type was not augmented compared to blood monocytes. Conditioned medium from cultured plaque macrophages significantly increased production of PAI-1 by endothelial cells (85 +/- 11% above basal) and vascular smooth muscle cells (25 +/- 10%) in vitro. This response was significantly greater than the response to monocyte-conditioned medium (endothelial cells 38 +/- 11%, vascular smooth muscle cells 2.5 +/- 2.0%). Stimulation of endothelial cell PAI-1 production by macrophage-conditioned medium was partially inhibitable by a monoclonal antibody to transforming growth factor-beta. Tissue type plasminogen activator production by endothelial cells and vascular smooth muscle cells was not affected by plaque macrophage- or monocyte-conditioned medium. Urokinase type plasminogen activator production by endothelial cells and vascular smooth muscle cells was undetectable in control medium and was augmented to similar levels in response to plaque macrophage- and monocyte-conditioned media. These results demonstrate upregulation of PAI-1 production by macrophages in atheromatous plaques and the capacity of soluble products from plaque macrophages to upregulate PAI-1 production by endothelial cells and vascular smooth muscle cells in vitro. These data suggest that macrophages in atherosclerotic plaques may inhibit thrombolysis both directly and indirectly by effects of their soluble products on endothelial cells and vascular smooth muscle cells.
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PMID:Atheromatous plaque macrophages produce plasminogen activator inhibitor type-1 and stimulate its production by endothelial cells and vascular smooth muscle cells. 836 83

S-nitrosothiols may serve as carriers in the mechanism of action of endothelium-derived relaxing factor (EDRF) by stabilizing the labile nitric oxide (NO) radical from inactivation by reactive species in the physiological milieu and by delivering NO to the heme activator site of guanylyl cyclase. Low-molecular-weight thiols, such as cysteine and glutathione, form S-nitrosothiol adducts with vasodilatory and antiplatelet properties, and protein thiols can interact in the presence of NO and/or EDRF to form uniquely stable S-nitroso-proteins. We now show that the S-nitroso-proteins, S-nitroso-albumin, S-nitroso-tissue type plasminogen activator, and S-nitroso-cathepsin B, have potent antiplatelet effects with an IC50 of approximately 1.5 microM. In the dog, S-nitroso-albumin inhibits ex vivo platelet aggregation and significantly prolongs the template bleeding time from 2.15 +/- 0.13 (mean +/- SEM) to 9.70 +/- 1.24 minutes. The antiplatelet action of S-nitroso-proteins is associated with the stimulation of guanylyl cyclase and a significant decrease in fibrinogen binding to platelets. S-Nitroso-proteins undergo thiol-nitrosothiol exchange with low-molecular-weight thiols to form low-molecular-weight S-nitroso-thiols, and they also interact directly with the platelet surface, both of which processes facilitate generation of NO. These data suggest that S-nitroso-proteins are potent antiplatelet agents and may be intermediates in the antiplatelet mechanism of EDRF action.
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PMID:Antiplatelet properties of protein S-nitrosothiols derived from nitric oxide and endothelium-derived relaxing factor. 838 13

Mice with combined homozygous deficiency of tissue-type plasminogen activator (t-PA) and urokinase-type plasminogen activator (u-PA) (T-U-), of t-PA and plasminogen activator inhibitor-1 (PAI-1) (T-P-), of u-PA and PAI-1 (U-P-) or of t-PA, u-PA, and PAI-1 (T-U-P-) were generated by inbreeding of mice with the respective deficiencies. Homologous recombination at the t-PA, u-PA and PAI-1 locus was verified by Southern blot analysis of genomic tail tip DNA, and confirmed by measurement of antigen levels in plasma or urine. T-P- and U-P- mice were apparently healthy and fertile. T-U- mice showed extensive fibrin deposition with calcification in the liver, whereas T-U-P- mice were significantly (p < 0.001) less affected. Spontaneous in vivo clot lysis measured 4 h after injection of a 125I-fibrin-labeled clot prepared from plasma of wild-type (WT) mice into the jugular vein, was (mean +/- SEM of n experiments) 2 +/- 1% (n = 8) for T-P-, 49 +/- 6% (n = 9) for U-P-, 1 +/- 1% (n = 4) for T-U- and 3 +/- 3% (n = 3) for T-U-P- mice, as compared to 32 +/- 4% (n = 10) for WT, 1 +/- 0% (n = 7) for T-, 30 +/- 5% (n = 5) for U- and 58 +/- 10% (n = 6) for P- mice. Plasminogen-dependent lysis of 125I-fibrin-labeled matrix and of 3H-proline-labeled subendothelial matrix (mean +/- SEM; n = 4 to 6) was lower with thioglycollate-stimulated macrophages obtained from U-P- mice (22 +/- 7% and 5 +/- 1%, respectively), as compared to WT mice (57 +/- 14% and 18 +/- 5%, respectively) and T-P- mice (87 +/- 6% and 27 +/- 4%, respectively). A similar decrease was previously observed with U- mice, but not with T- or P- mice. Thus, the phenotype of mice with combined deficiency of t-PA and PAI-1 or of u-PA and PAI-1 is similar to the phenotype observed in mice with single deficiency of the plasminogen activator. Additional deletion of PAI-1 does not affect viability, fertility, macrophage function or thrombolytic potential of the single deficient mice. Additional deletion of PAI in mice with combined deficiency of t-PA and u-PA does not restore the deficient in vivo fibrinolytic capacity, but significantly reduces the thrombotic phenotype, as revealed by fewer, smaller and less calcified fibrin deposits in the liver.
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PMID:Biological effects of combined inactivation of plasminogen activator and plasminogen activator inhibitor-1 gene function in mice. 856 Apr 24

Plasminogen is activated into the fibrinolytic enzyme plasmin via: a tissue type activator and F-XII dependent and F-XII independent systems. The purpose of this study was extract and quantify the tissue-type plasminogen activator present in the salivary glands of rats. The extracted plasminogen activator-EPA- was obtained by homogenizing 1 vol of tissue with 1 vol of 2M KSCN solution. Solution with EPA was applied by triplicated in the standard plasminogen-rich and plasminogen-free fibrin plates. The degree of fibrinolytic activity was observed as areas of liquefaction and measured as the product (mm2) of the two perpendicular diameter of the lysed zones. The submaxillary's EPA produced a mean lytic area of 198 mm2 +/- 18 SEM only in the plasminogen-rich fibrin plate. This activity extrapolated into a standard dilution curve, represented the equivalent to a 50 mg/ml plasmin solution. No lysis was induced by EPA from parotid and sublingual glands. The antifibrinolytic drug E-ACA in a dose dependent inhibitory action, significantly reduced the lytic activity induced by submaxillary's EPA. The observation that EPA produced areas of liquefaction only in plasminogen-rich fibrin plate and that this activity was inhibited by E-ACA is clear indication that the zones of lysis was specific fibrinolysis -activation of plasminogen into plasmin- and not due to non-specific proteolysis.
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PMID:Extraction of plasminogen activator in the rat's submaxillary gland. 858 May 23

Homozygous plasminogen-deficient (Plg-/-) mice had a significantly reduced thrombolytic capacity toward intravenously injected 125I-fibrin labeled plasma clots prepared from Plg-/- murine plasma (9% +/- 3% lysis after 8 hours; (mean +/- SEM, n = 6), as compared with 82% +/- 8% in wild-type mice; P < .0001). Bolus injection of 1 mg purified murine plasminogen in 10- to 17-week-old Plg-/- mice increased the plasminogen antigen and activity levels at 8 hours to normal levels (130 +/- 5 micrograms/mL). Plasminogen administration was associated with significant restoration of thrombolytic potential (64% +/- 7% spontaneous clot lysis; P < .0001 versus lysis without plasminogen injection). Bolus injection of 1 mg plasminogen in homozygous tissue-type plasminogen activator-deficient (t-PA-/-) mice doubled the plasminogen antigen and activity levels after 8 hours and increased 125I-fibrin clot lysis at 8 hours from 13% +/- 3% to 34% +/- 5% (P = .008). Fibrinogen, t-PA antigen and alpha 2-antiplasmin activity levels after 8 hours were not significantly different in the groups with or without plasminogen injection. Injection of plasminogen induced a variable increase (on average 7- to 10-fold) of PAI-1, but no correlation with the extent of spontaneous clot lysis was observed. Histopathologic examination at the end of the experiments revealed that fibrin deposition in the liver of Plg-/- mice was slightly reduced 8 hours after bolus plasminogen injection (P = .007) and markedly reduced after 24 hours (P < .0001). Plasminogen antigen levels in liver extracts were comparable with those found in wild-type mice at 8 hours (130 +/- 20 versus 110 +/- 15 ng/mg protein) and decreased to 25 +/- 3.2 ng/mg protein at 24 hours. Thus, restoration of normal plasminogen levels in Plg-/- mice normalized the thrombolytic potential toward experimentally induced pulmonary emboli, and resulted in removal of endogenous fibrin deposits within 24 hours.
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PMID:Restoration of thrombolytic potential in plasminogen-deficient mice by bolus administration of plasminogen. 870 43

The successful application of thrombolytic therapy to acute myocardial infarction has prompted a reinvestigation of thrombolytic therapy for acute stroke. However, an examination of safety and efficacy of thrombolytic therapy in acute thromboembolic stroke has precluded the entry of patients taking either antiplatelet or anticoagulant therapy. It was therefore of interest, in an established rabbit model of thromboembolic stroke, to examine the use of tissue plasminogen activator therapy in combination with ticlopidine treatment. Following ticlopidine administration (10 mg kg-1, i.v., daily for 5 days), rabbits (n = 7) were embolized by injecting a tin-laden clot into the internal carotid artery with clot placement confirmed by x-ray. Three hours later, t-PA was initiated as a square-wave pulse (6.3 mg kg-1 total dose, given as a 20% bolus, with the remainder administered over 2 h as a continuous infusion). The protocol was continued for a total of 7 h following embolization. Complete clot lysis was demonstrated in 6 of 7 animals. Brain infarct size (triphenyltetrazolium chloride staining) was 36.0 +/- 12.9% hemisphere (mean +/- SEM). Both clot lysis rate and infarct size were very similar to that previously seen following administration of t-PA alone (58% and 31.6 +/- 6.4% hemisphere, respectively) but in marked contrast to previous results seen with intravenous aspirin (no clot lysis). These results suggest that antiplatelet agents used clinically for stroke prophylaxis (aspirin or ticlopidine) may influence the success rate of thrombolysis following initiation of thrombolytic therapy for acute thromboembolic stroke.
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PMID:Combination tissue plasminogen activator and ticlopidine therapy in a rabbit model of acute thromboembolic stroke. 871 36

Staphylokinase, a bacterial plasminogen activator, is a potent, highly fibrin-specific but antigenic thrombolytic agent in humans. In an effort to attenuate the antigenicity of wild-type staphylokinase (SakSTAR variant), 2 of its 3 immunodominant epitopes were altered by substituting clusters of 2 or 3 charged amino acids with alanine, yielding the mutant SakSTAR.M38 (K35A, E38A, K74A, E75A, R77A), which was less antigenic in inbred New Zealand White rabbits. In the present study, groups of 6 baboons (Papio hamadryas) were randomized to SakSTAR (group 1) or SakSTAR.M38 (group 2). The thrombolytic potencies of 50 micrograms/kg compound at baseline, assessed in an extracorporeal thrombosis model, were similar: 77 +/- 2.9% (mean +/- SEM) clot lysis in group 1 and 83 +/- 3.6% in group 2. Groups 1 and 2 were immunized subcutaneously at 2, 3, and 5 weeks with 500 micrograms SakSTAR or SakSTAR.M38, respectively. From 6 weeks, group 1 developed significantly more antibody-related neutralizing activity than group 2 (maximal titer at 8 weeks of 100 +/- 23 micrograms SakSTAR and of 22 +/- 7.1 micrograms SakSTAR.M38 neutralized per milliliter of plasma, respectively). Neutralizing activities subsequently decreased gradually to 10-20% of peak values at 18 weeks. At 6 weeks, both groups were resistant to thrombolysis with 50 micrograms/kg of either compound. Rechallenge at 18 weeks with 250 micrograms/kg of the immunizing compound showed a significantly better recovery of the thrombolytic potency of SakSTAR.M38 (68 +/- 4.5% clot lysis) than of SakSTAR (39 +/- 5.3% clot lysis). Neither agent degraded fibrinogen or depleted alpha 2-antiplasmin. Therefore, SakSTAR.M38 is comparably active and fibrin-specific but less antigenic than wild-type SakSTAR. These findings in outbred primates confirm and extend earlier observations in inbred rabbits and provide a basis for the further development of staphylokinase variants with reduced antigenicity in humans.
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PMID:Comparative antigenicity of recombinant wild-type staphylokinase (SakSTAR) and a selected mutant (SakSTAR.M38) in a baboon thrombolysis model. 876 47

The estimation of the activity of circulating tissue-type plasminogen activator (t-PA) using the Coaset t-PA or Spectrolyse/fibrin t-PA reagent kits involves incubation of plasma with excess plasminogen in the presence of human fibrin fragments, and detection of the plasmin generated by an end-point amidolytic assay. We examined the interference of endogenous inhibitors such as plasminogen activator inhibitor-1 (PAI-1) and alpha 2-antiplasmin, and that of an endogenous activator namely single-chain urokinase-type plasminogen activator (scu-PA) on the Coaset t-PA assay. An extra acidification of the samples already collected into an acidified anticoagulant raised the mean Coaset t-PA activity of 15 samples from 1.39 +/- 0.25 (mean +/- SEM) to 1.71 +/- 0.27 (P < 0.001). In twelve of these samples, the PAI-1 and alpha 2-antiplasmin activities were determined. The increase in the t-PA activity by acidification was found to correlate inversely with PAI-1 (r = -0.66; n = 12; P = 0.019) but not with alpha 2-antiplasmin, demonstrating that the extra acidification step released t-PA from the t-PA-PAI-1 complex, but had no influence on the endogenous alpha 2-antiplasmin in the assay system. Incubation with monoclonal antibody against alpha 2-antiplasmin increased the Coaset t-PA activity in a concentration-dependent manner, linearly up to 2 micrograms/ml (final) of the antibody, irrespective of the extra acidification, the maximal rise being 41.5% and 19.7% in an in-house and commercial plasma pools respectively. In contrast to alpha 2-antiplasmin antibody, monoclonal antibody against scu-PA decreased the Coaset t-PA activity in the in-house and commercial plasma pools again in a concentration-related manner demonstrating that scu-PA in addition to t-PA was being measured by this method. Incubation with an irrelevant antibody (anti-DNA monoclonal antibody) did not affect the Coaset t-PA values. The mean Coaset t-PA activity of 40 subjects decreased from 1.76 +/- 0.10 (mean +/- SEM) to 0.73 +/- 0.07 when incubated with 8 micrograms/ml (final) monoclonal antibody against scu-PA (P < 0.001). The 't-PA' activity in the absence of scu-PA antibody correlated with the u-PA antigen (r = 0.53; n = 40; P < 0.001), and the activity in the presence of scu-PA antibody correlated with the t-PA activity measured by the bio-functional immunosorbent assay (r = 0.86; n = 40; P < 0.001). Hence, unless the Coaset t-PA activity is measured with appropriate amounts of antibodies to alpha 2-antiplasmin and scu-PA, it is best to qualify the activity measured by this method as being that of total plasminogen activators.
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PMID:Interference of an activity assay of tissue-type plasminogen activator in human plasma by endogenous factors. 884 3

The fibrin-specificity and procoagulant effects of recombinant staphylokinase (Sak42D) were compared with those of recombinant tissue-type plasminogen activator (rt-PA) in patients with acute myocardial infarction. Plasma samples were obtained at baseline and at 25 and 90 min, from 24 patients who were randomly assigned to a double bolus (15 mg each, 30 min apart) administration of Sak42D or to accelerated weight-adjusted rt-PA (maximum of 100 mg over 90 min). Baseline levels of fibrinopeptide A (FPA), prothrombin fragment 1 + 2 and thrombin-antithrombin III complex (TAT) were comparable in the Sak42D and rt-PA groups (p > or = 0.6). In patients treated with Sak42D, plasma levels of FPA, prothrombin fragment 1 + 2 and TAT did not markedly increase during treatment (p = 0.06, p = 0.4 and p = 0.03, respectively). In contrast, during administration of rt-PA the levels of FPA, prothrombin fragment 1 + 2 and TAT increased significantly over baseline (p = 0.003, p < 0.0001 and p = 0.001, respectively). As a result, the levels of all three procoagulant parameters were significantly lower during treatment with Sak42D as compared to rt-PA. Thus, FPA levels in the Sak42D group (median values) were 40 ng/ml at 25 min and 11 ng/ml at 90 min, as compared to 88 ng/ml and 50 ng/ml in the rt-PA group (p = 0.0007 and p = 0.009, respectively). Prothrombin fragment 1 + 2 levels in the Sak42D group were 1.3 nM at 25 min and 1.2 nM at 20 min, as compared to 11 nM and 5.3 nM in the rt-PA group (both p < 0.0001). TAT levels were 4.7 ng/ml at 25 min and 6.2 ng/ml at 90 min in the Sak42D group, with corresponding values of 16 ng/ml and 9.6 ng/ml in the rt-PA group (p = 0.02 and p = 0.03, respectively). In the patients treated with Sak42D, no significant systemic fibrinolytic activation was observed, as revealed by unaltered levels of clottable fibrinogen, plasminogen and alpha 2-antiplasmin up to 90 min after the start of therapy. In contrast, the corresponding residual levels at 90 min in patients treated with rt-PA decreased to (mean +/- SEM; n = 12) 62 +/- 6%, 45 +/- 5% and 52 +/- 10%, respectively (all p < or = 0.01 versus the Sak42D group). These data confirm the high degree of fibrin-specificity of Sak42D and demonstrate that this is associated with significantly less generation of procoagulant activity in plasma after intravenous administration in patients with acute myocardial infarction.
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PMID:Procoagulant properties of intravenous staphylokinase versus tissue-type plasminogen activator. 897


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