Enhancement of Serrapeptase Hyper Producing Mutant by Combined Chemical and UV Mutagenesis and its Potential for Fibrinolytic Activity

SummaryWe analyzed fibrinolytic parameters in 20 healthy men and 20 healthy women, aged from 25 to 59, before and after 10 and 20 min venous occlusion. The 10 min post-occlusion fibrinolytic activity measured directly in diluted unfractionated plasma by a highly sensitive 125I-fibrin plate assay correlated well with the activity of euglobulins determined by the classical fibrin plate assay (r = 0.729), but pre-stasis activities determined with these two methods did not correlate (r = 0.084). The enhancement of fibrinolytic activity after venous occlusion was mainly due to an increase of t-PA in the occluded vessels (4-fold increase t-PA antigen after 10 min and 8-fold after 20 min venous occlusion). Plasminogen activator inhibitor (PAI) activity and plasminogen activator inhibitor 1 (PAI-1)1 antigen levels at rest showed considerable dispersion ranging from 1.9 to 12.4 U/ml, respectively 6.9 to 77 ng/ml. A significant increase of PAI-1 antigen levels was observed after 10 and 20 min venous occlusion. At rest no correlation was found between PAI activity or PAI-1 antigen levels and the fibrinolytic activity measured by 125I-FPA. However, a high level of PAI-1 at rest was associated with a high prestasis antigen level of t-PA and a low fibrinolytic response after 10 min of venous stasis. Since the fibrinolytic response inversely correlated with PAI activity at rest, we conclude that its degree depends mainly on the presence of free PAI.

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Coagulation and Fibrinolytic Activity in Behçet's Disease

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646409 ◽

1991 ◽

Vol 66 (03) ◽

pp. 292-294 ◽

Cited By ~ 57

Author(s):

K K Hampton ◽

M A Chamberlain ◽

D K Menon ◽

J A Davies

Keyword(s):

Plasminogen Activator ◽

Tissue Plasminogen Activator ◽

Behçet’S Disease ◽

Fibrinolytic Activity ◽

Behcet’S Disease ◽

Behçet's Disease ◽

Plasminogen Activator Inhibitor ◽

Behcet's Disease ◽

Tissue Plasminogen ◽

Von Willebrand

SummaryCoagulation and fibrinolytic activities were studied in 18 subjects with Behçet's disease and compared with results from 14 matched control patients suffering from sero-negative arthritis. Significantly higher plasma concentrations (median and range) were found in Behçet's patients for the following variables: fibrinogen 3.7 (1.7-6.9) vs 3.0 (2.0-5.1) g/1, p <0.05; von Willebrand factor antigen, 115 (72-344) vs 74 (60-119)%, p <0.002; plasminogen activator activity (106/ECLT2) 219 (94-329) vs 137 (78-197) units, p <0.002; tissue plasminogen activator inhibitor (t-PA-I) activity, 9.1 (5.5-19.3) vs 5.1 (1.8-12.0) IU/ml, p <0.002; and PAI-1 antigen, 13.9 (4.5-20.9) vs 6.4 (2.4-11.1) ng/ml, p <0.002. Protein C antigen was significantly lower: 97 (70-183) vs 126 (96-220)%, p <0.02. No differences were observed in antithrombin III activity or antigen, factor VIII coagulant activity, fibrinopeptides A and Bβ15-42, plasminogen, α-2-antiplasmin, functional and immunological tissue-plasminogen activator, thrombin-antithrombin complexes and D-dimer. Levels of tissue plasminogen activator inhibitor (activity and antigen) correlated with disease activity while fibrinogen and von Willebrand factor concentrations did not. Seven of the 18 subjects with Behçet's disease had suffered thrombotic events but it was not possible to distinguish these from the 11 patients without thrombosis using the assays performed. The results suggest the abnormal fibrinolytic activity in Behçet's disease is due to increased inhibition of tissue plasminogen activator. No abnormality of coagulation or fibrinolytic activity specific to Behçet's disease was detected.

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Dose-response Curves in the Fibrin Plate Assay Fibrinolytic Activity of Proteases

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647705 ◽

1974 ◽

Vol 32 (02/03) ◽

pp. 356-365 ◽

Cited By ~ 12

Author(s):

F Haverkate ◽

D. W Traas

Keyword(s):

Dose Response ◽

Fibrinolytic Activity ◽

Enzyme Concentration ◽

Response Curves ◽

Agar Gel ◽

Porcine Tissue ◽

Plate Assay ◽

Fibrin Plate ◽

Different Types ◽

Dose Response Curves

SummaryIn the fibrin plate assay different types of relationships between the dose of applied proteolytic enzyme and the response have been previously reported. This study was undertaken to determine whether a generally valid relationship might exist.Trypsin, chymotrypsin, papain, the plasminogen activator urokinase and all of the microbial proteases investigated, including brinase gave a linear relationship between the logarithm of the enzyme concentration and the diameter of the circular lysed zone. A similar linearity of dose-response curves has frequently been found by investigators who used enzyme plate assays with substrates different from fibrin incorporated in an agar gel. Consequently, it seems that this linearity of dose-response curves is generally valid for the fibrin plate assay as well as for other enzyme plate bioassays.Both human plasmin and porcine tissue activator of plasminogen showed deviations from linearity of semi-logarithmic dose-response curves in the fibrin plate assay.

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Influence of Moderate and Strenuous Daily Physical Activity on Fibrinolytic Activity of Blood: Possibility of Plasminogen Activator Stores Depletion

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646834 ◽

1979 ◽

Vol 41 (04) ◽

pp. 745-755 ◽

Cited By ~ 22

Author(s):

Dušan Keber ◽

Mojca Stegnar ◽

Irena Keber ◽

Bojan Accetto

Keyword(s):

Physical Activity ◽

Plasminogen Activator ◽

Fibrinolytic Activity ◽

Regular Physical Activity ◽

Clot Lysis ◽

Moderate Activity ◽

Lysis Time ◽

Strenuous Activity ◽

Activity Measurements ◽

Clot Lysis Time

SummaryFibrinolysis was studied in 10 alpinists during regular physical activity of different intensity. Blood was sampled at rest and after exposure to submaximal workload on the treadmill on three occasions: before and after 6 months physical conditioning (moderate physical activity), and after 6 weeks of an alpinistic expedition (strenuous physical activity). Measurements included submaximal working capacity, fibrinogen, euglobulin clot lysis time (ELT), whole plasma clot lysis time, and estimations derived from ELT - percent increase in fibrinolytic activity after exercise (RFS), and absolute increase in fibrinolytic activity after exercise (PAR).Regular moderate activity increased the resting level of ELT, but strenuous activity decreased is. After each treadmill testing, a marked increase in fibrinolytic activity was observed. RFS was unaltered at all three testings. PAR increased after moderate activity, but decreased after strenuous activity.The results indicate that regular physical activity can lead from enhanced to decreased resting activity of plasminogen activator in blood. It is presumed that increased release of activator during prolonged stress causes partial depletion of endothelial stores with the consequence of decreased activator activity in the blood.

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Impaired Fibrinolysis in Obstructive Jaundice - Evidence from Clinical and Experimental Studies

Thrombosis and Haemostasis ◽

10.1055/s-0038-1647628 ◽

1988 ◽

Vol 60 (01) ◽

pp. 025-029 ◽

Cited By ~ 8

Author(s):

M Colucci ◽

D F Altomare ◽

G Chetta ◽

R Triggiani ◽

L G Cavallo ◽

...

Keyword(s):

Renal Failure ◽

Bile Duct ◽

Obstructive Jaundice ◽

Plasminogen Activator ◽

Fibrinolytic Activity ◽

Experimental Studies ◽

Plasminogen Activator Inhibitor ◽

Venous Stasis ◽

Microvascular Thrombosis ◽

Duct Ligation

SummaryMicrovascular thrombosis is considered an important pathogenetic factor in renal failure associated with obstructive jaundice but the mechanisms leading to fibrin deposition are still unknown. The plasma levels of plasminogen activator inhibitor (PAI) in 29 patients with obstructive jaundice were found significantly increased as compared to 20 nonjaundiced patients. Fibrin autography of plasma supplemented with tissue plasminogen activator (t-PA) revealed that in icteric samples most of the added activator migrated with an apparent Mr of 100 kDa, corresponding to t-PA-PAI complex, whereas in control samples virtually all t-PA migrated as free enzyme. PAI activity detected in icteric samples is similar to the endothelial type PAI since it is neutralized by a monoclonal antibody against PAI-1.Venous stasis in jaundiced patients was neither associated with an increase in blood fibrinolytic activity nor with a decrease in PAI activity. Immunologic assay showed that t-PA release was impaired in 3 out of 4 patients. In controls, venous occlusion induced an increase in both fibrinolytic activity and t-PA antigen and a reduction in PAI activity. Bile duct recanalization in jaundiced patients subjected to surgery was accompanied by a decrease in plasma PAI activity which paralleled the decrease in serum bilirubin levels. In nonjaundiced patients, surgical treatment did not cause significant changes in either parameter. Rabbits made icteric by bile duct ligation showed an early and progressive increase in plasma PAI activity indicating that obstructive jaundice itself causes the elevation of circulating PAI. It is concluded that obstructive jaundice is associated with a severe impairment of fibrinolysis which might contribute to microvascular thrombosis and renal failure.

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Fibrinolysis in Decidual Spiral Arteries in Late Pregnancy

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646752 ◽

1978 ◽

Vol 39 (03) ◽

pp. 751-758 ◽

Cited By ~ 12

Author(s):

B L Sheppard ◽

J Bonnar

Keyword(s):

Electron Microscopy ◽

Endothelial Cells ◽

Fibrinolytic Activity ◽

Physiological Mechanism ◽

Late Pregnancy ◽

Full Term ◽

Fibrin Deposition ◽

Placental Villi ◽

The Media ◽

Slide Technique

SummaryThe fibrinolytic activity of the intimal cells of decidual spiral arteries and the syncytium of placental villi was studied by electron microscopy in ten normal full-term human pregnancies using a modification of the fibrin slide technique. Endothelial cells lining the intima of the decidual spiral arteries showed a considerably greater fibrinolytic activity than intimal cytotrophoblast and the syncytiotrophoblast showed no activity.The replacement of endothelial cells by an intimal lining of cytotrophoblast, and the presence of cytotrophoblast in the media, appears to play an important role in the reduction of the fibrinolytic activity of the vessel. This inhibition of fibrinolytic activity in the utero-placental arteries may be the physiological mechanism which controls fibrin deposition in these vessels and on the placental villi.

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Binding of Tissue Plasminogen Activator to Vascular Grafts

Thrombosis and Haemostasis ◽

10.1055/s-0038-1646541 ◽

1989 ◽

Vol 61 (01) ◽

pp. 131-136 ◽

Cited By ~ 8

Author(s):

Richard A Harvey ◽

Hugh C Kim ◽

Jonathan Pincus ◽

Stanley Z Trooskin ◽

Josiah N Wilcox ◽

...

Keyword(s):

Plasminogen Activator ◽

Tissue Plasminogen Activator ◽

Fibrinolytic Activity ◽

Polymer Surface ◽

Enzymic Activity ◽

Vascular Prostheses ◽

Chemically Modified ◽

Insoluble Surfactant ◽

Tissue Plasminogen

SummaryTissue plasminogen activator labeled with radioactive iodine (125I-tPA) was immobilized on vascular prostheses chemically modified with a thin coating of water-insoluble surfactant, tridodecylmethylammonium chloride (TDM AC). Surfactant- treated Dacron, polytetrafluoroethylene (PTFE), silastic, polyethylene and polyurethane bound appreciable amounts of 125I- tPA (5-30 μg 125I-tPA/cm2). Upon exposure to human plasma, the amount of 125I-tPA bound to the surface shows an initial drop during the first hour of incubation, followed by a slower, roughly exponential release with a t½ of appoximately 75 hours. Prostheses containing bound tPA show fibrinolytic activity as measured both by lysis of clots formed in vitro, and by hydrolysis of a synthetic polypeptide substrate. Prior to incubation in plasma, tPA bound to a polymer surface has an enzymic activity similar, if not identical to that of the native enzyme in buffered solution. However, exposure to plasma causes a decrease in the fibrinolytic activity of both bound tPA and enzyme released from the surface of the polymer. These data demonstrate that surfactant-treated prostheses can bind tPA, and that these chemically modified devices can act as a slow-release drug delivery system with the potential for reducing prosthesis-induced thromboembolism.

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Gastric Fibrinolysis

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651400 ◽

1975 ◽

Vol 34 (02) ◽

pp. 409-418 ◽

Cited By ~ 9

Author(s):

I. M Nilsson ◽

S.-E Bergentz ◽

U Hedner ◽

K Kullenberg

Keyword(s):

Gastric Ulcer ◽

Gastric Juice ◽

High Activity ◽

Fibrinolytic Activity ◽

Duodenal Juice ◽

Bleeding Tendency ◽

Local Fibrinolysis ◽

Heated Plates

SummaryGastric juice from 15 normals, 20 patients with gastric ulcer and 4 patients with erosive haemorrhagic gastroduodenitis was investigated in respect of its activity on unheated and heated fibrin plates and its content of FDP and plasminogen or plasmin with immunochemical methods. Gastric juice from normals showed no activity on unheated and heated fibrin plates, and no FDP or plasminogen could be demonstrated. In the patients with gastric ulcer the gastric juice showed little or no fibrinolytic activity on fibrin plates except in 2, who had regurgitation of duodenal juice and neutral pH of the juice. These patients had equally high activity on heated as on unheated plates and no plasmin could be demonstrated. It was shown that this activity was not due to fibrinolysis, but to non-specific proteolytic activity (probably trypsin). The patients with erosive haemorrhagic gastroduodenitis exhibited quite a different picture. The gastric juice from these patients showed extremely high activity on fibrin plates, the activity was higher on unheated than on heated plates. The activity was inhibited in vitro by addition of EACA and in vivo after administration of AMCA. The occurrence of plasmin could be demonstrated directly immunologically in the gastric juice. By comparison of plasmin and trypsin in various assays it could further be proved that the gastric juice in these cases contained plasminogen activator and plasmin. The patients with erosive haemorrhagic gastroduodenitis showed no increase in fibrinolysis in the blood, but low values for plasminogen and α2M, and the serum contained FDP. These findings in the blood and gastric juice were interpreted as signs of local fibrinolysis in the stomach and duodenum. There is reason to assume that this gastric fibrinolysis contributes substantially to the bleeding tendency. The effect of administration of AMCA on fibrinolytic activity and the haemorrhage lends support to the assumption of such a mechanism.

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Plasma Fibrinolytic Activity Following Oral Anabolic Steroid Therapy

Thrombosis and Haemostasis ◽

10.1055/s-0038-1651371 ◽

1975 ◽

Vol 34 (01) ◽

pp. 236-245 ◽

Cited By ~ 3

Author(s):

I. D Walker ◽

J. F Davidson ◽

P Young ◽

J. A Conkie

Keyword(s):

Heart Disease ◽

Ischaemic Heart Disease ◽

Anabolic Steroid ◽

Fibrinolytic Activity ◽

Anabolic Steroids ◽

Long Term Effects ◽

Lysis Time ◽

Detailed Evaluation ◽

Euglobulin Lysis Time

SummarySix anabolic steroids were assessed for their ability to enhance plasma fibrinolytic activity in males with ischaemic heart disease. Five 17α-alkylated steroids (Ethyloestrenol, Norethandrolone, Methandienone, Methylandrostenediol and Oxymetholone) were examined and all produced a significant increase in plasma plasminogen activator as measured by the euglobulin lysis time. The only non-17α-alkylated steroid studied (Methenolone acetate) failed to enhance fibrinolysis. The 17α-alkylated steroids studied all deserve more detailed evaluation of their long term effects on plasma fibrinolytic activity.

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