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1.
Chimeric derivative of fibrolase, a fibrinolytic enzyme from southern copperhead venom, possesses inhibitory activity on platelet aggregation 总被引:3,自引:0,他引:3
Fibrolase, a metalloproteinase isolated from the venom of Agkistrodon contortrix contortrix (southern copperhead snake), is a direct acting fibrinolytic enzyme that has been used to digest occlusive blood clots in animal models. The snake venom enzyme directly degrades fibrin associated with platelet rich blood clots and does not rely on plasminogen activation. Rethrombosis is a serious complication that is experienced in a significant percentage of patients treated with thrombolytic agents to remove occlusive vascular thrombi. The involvement of platelets in the initiation of rethrombosis is well known. Arg-Gly-Asp-(RGD)-containing agents have been shown to inhibit rethrombosis following thrombus dissolution by plasminogen activators. In an effort to create a more effective fibrinolytic enzyme and to target the enzyme to platelet-rich thrombi, thereby decreasing the potential for rethrombosis, a chimeric derivative of fibrolase has been produced. This report describes the construction and biochemical characterization of the chimeric enzyme and an evaluation of its in vitro activities. The chimera was formed by covalently incorporating an RGD-like peptide into fibrolase. The site of peptide attachment was determined to be a single lysine residue remote from the enzymes active site. Covalent modification of fibrolase with the RGD-like peptide did not inhibit either fibrinolytic activity of the enzyme nor platelet aggregation inhibitory activity of the peptide. The chimera not only retained the same level of enzymatic activity as native fibrolase, but also acquired the ability to inhibit platelet aggregation by binding to the fibrinogen receptor (integrin alphaIIbbeta3) on platelets. 相似文献
2.
Erythrocyte aggregation (examined microscopically in diluted blood), lipid and protein plasma profiles, and fibrinolytic activity were studied in endurance athletes. Division of the athletes into two subgroups by cluster analysis showed that a higher level of fitness was associated with a lower plasminogen activity; enhanced fibrinolysis; increased blood fluidity; lower fibrinogen, cholesterol, and triglyceride levels; a relatively low erythrocyte aggregation index; and high suspension stability of the blood. Fibrinogen was the key plasma factor determining erythrocyte aggregation. Its level was closely correlated with plasminogen activity. Discriminant analysis showed that most differences between groups of athletes were connected with plasminogen activity, the von Willebrand factor, and fibrinolytic activity. 相似文献
3.
Gramicidin S (GS) is a cyclic decapeptide antibiotic active against both Gram‐positive and Gram‐negative bacteria as well as against several pathogenic fungi. However, clinical application of GS is limited because of GS hemolytic activity. The large number of GS analogues with potentially attenuated hemolytic activity has been developed over the last two decades. For all new GS derivatives, the antimicrobial test is accompanied with the hemolytic activity assay. At the same time, neither GS nor its analogues were tested against other blood cells. In the present work, the effects of GS on platelets and platelet aggregates have been studied. GS interaction with platelets is concentration dependent and leads either to platelet swelling or platelet shape change. Effect of GS on platelets is independent of platelet aggregation mechanism. GS induces disaggregation of platelet aggregates formed in the presence of aggregation agonists. The rate of the GS interaction with platelet membranes depends on membrane lipid mobility and significantly increases with temperature. The interaction of GS with the platelet membranes depends strongly on the state of the membrane lipids. Factors affecting the membrane lipids (temperature, lipid peroxidation and ionising irradiation) modify GS interaction with platelets. Our results show that GS is active not only against erythrocytes but also against other blood cells (platelets). The estimated numbers of GS molecules per 1 µm2 of a blood cell required to induce erythrocyte hemolysis and disaggregation of platelet aggregates are comparable. This must be considered when developing new antimicrobial GS analogues with improved hemolytic properties. Copyright © 2012 European Peptide Society and John Wiley & Sons, Ltd. 相似文献
4.
N A Fedorov N A Gorbunova T A Balakina V N Kolpikov 《Biulleten' eksperimental'no? biologii i meditsiny》1978,86(7):116-120
Vascular-thrombocytic and plasma hemostasis was studied in dogs after blood loss (40--50 ml/kg weight) and subsequent hypervolemic (60--65 ml/kg weight) transfusion of homologous (from 3 donors) platelet and leukocyte rich in plasma. After a short phase of hypercoagulation all the tested dogs displayed hypocoagulation accompanied by a decrease in the count of platelets and a fall of their aggregation function, prolongation of bleeding time, diminution of capillary wall resistance, a decrease of plasma fibrinogen concentration and factor XIII activity, and by a rise of blood fibrinolytic activity. The data obtained show the development of the acute disseminated intravascular coagulation syndrome (DIC). DIC model is porposed on the basis of the results obtained. 相似文献
5.
We have used a technique for the simultaneous measurement of platelet activation and aggregation in whole blood using two-color immunofluorescence and flow cytometry to study the relationship between the release reaction and aggregation. A monoclonal antibody specific for the alpha granule membrane protein GMP-140 was used to measure the release reaction, and a monoclonal antibody specific for platelet membrane glycoprotein Ib (GPIb) was used to identify platelets and platelet aggregates. Aggregates were identified as particles expressing both levels of GPIb and size larger than that of resting single platelets. Anticoagulated whole blood was incubated with platelet agonists. At various times samples of the blood were removed and immediately fixed with paraformaldehyde. Blood that had been anticoagulated with ethylenediamine tetraacetic acid showed progressive release of platelets but little or no aggregation. However, blood anticoagulated with citrate or heparin showed correlated release and aggregation. The degree of aggregation was greater in heparin than in citrate. The expression of GPIb and GMP-140 increased in direct proportion to the size of the aggregates. Aggregates were observed varying in apparent diameter up to approximately 20 microns. During prolonged incubation there was progressive disaggregation of adenosine diphosphate (ADP)-induced aggregates. After disaggregation the proportion of GMP-140 negative single platelets increased, indicating that both released and nonreleased platelets participated in the aggregation. There was little or no disaggregation of phorbol myristate acetate (PMA)-induced aggregates. The relatively small size and reversibility of platelet aggregates that we have observed in whole blood may be relevant to phenomena occurring in vivo and in extracorporeal circulation.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
6.
Iu Iu Chirkov N N Belushkina I A Tyshchuk I S Severina 《Biulleten' eksperimental'no? biologii i meditsiny》1991,111(2):152-154
Activity of guanylate cyclase (GC) and its capacity for sodium nitroprusside (SNP) activation were determined in platelets with different state of aggregation. The development of ADP-induced reversible aggregation was accompanied by a decrease in the basal GC activity and by an increase in the SNP activation of GC. It was shown that elevation of GC sensitivity to SNP during the aggregation might be due to the decrease in the state of enzyme blood deficiency. Preincubation of platelets with SNP before ADP adding markedly diminished or even prevented aggregation, depending on SNP concentration. GC parameters in platelets with prevented aggregation were just the same as in control. It is suggested that the regulatory role of cGMP system in platelet aggregation may be seen in the increase in GC sensitivity to endogenous activator, presumably to NO. 相似文献
7.
N A Gorbunova E P Romanova T A Balakina E I Agafonova 《Biulleten' eksperimental'no? biologii i meditsiny》1985,99(1):3-4
It was established in dog experiments that aggregation activity of platelets and the rate of aggregation were higher in arterial than in venous blood, with the platelet content being identical. Antiaggregation activity of vascular walls of the arterial system. The data obtained indicate the presence of the arteriovenous difference of the function of the vascular-platelet hemostasis, which is of a role in the processes of regulating the state of blood aggregation in the body. 相似文献
8.
9.
Cho MJ Pestina TI Steward SA Jackson CW Kent Gartner T 《Biochemical and biophysical research communications》2002,292(4):916-921
The work presented here demonstrates that platelets from mice lacking LAT (linker for the activation of T cells) show reversible aggregation in response to concentrations of collagen that cause TxA2/ADP-dependent irreversible aggregation of control platelets. The aggregation defect of the LAT-deficient platelets was shown to be the result of almost no TxA2 production and significantly diminished ADP secretion. In contrast, the LAT deficiency does not affect aggregation induced by high concentrations of collagen because that aggregation is not dependent on TxA2 and/or ADP. Even though ADP and TxA2 provide amplification signals for platelet activation in response to low concentrations of collagen, LAT-deficient platelets hyperaggregate to low levels of U46619, a TxA2 analog, or ADP. Though the mechanism(s) of costimulatory signals by collagen, ADP, and TxA2 remains unidentified, it is clear that LAT plays a positive role in collagen-induced, TxA2/ADP-dependent aggregation, and a negative role in TxA2 or ADP-induced platelet aggregation. 相似文献
10.
目的:明确京尼平苷和京尼平的抗血栓和纤溶的作用效果。方法:用旁路循环血栓形成模型和颈总动脉血栓模型,测定CT、BT、PRT、PT,全血溶栓,计算血栓形成抑制率。结果:试验表明,京尼平苷及其苷元能显著延长凝血、出血时间,减少两个血栓模型的血栓重量,京尼平苷高剂量组与阳性药组相比,溶栓作用具有显著性差异(P<0.05),优于阳性药;京尼平、京尼平苷抗凝血及抗血栓作用与阿司匹林组相比无显著差异(P>0.05)。结论:实验表明中药栀子能延长凝血和出血时间,可能有一定的抗血栓和溶栓作用。 相似文献
11.
Platelet activation as a marker for in vivo prothrombotic activity: detection by flow cytometry 总被引:7,自引:0,他引:7
Tomer A 《Journal of biological regulators and homeostatic agents》2004,18(2):172-177
Circulating platelets play a pivotal role in hemostasis. The platelet hemostatic function involves the direct interaction with damaged vessel walls, and circulating coagulation factors, primarily thrombin resulting in platelet activation, aggregation and formation of hemostatic plug. Flow cytometry is a useful technique for the study of platelet activation in circulating blood. Platelet activation markers for ex vivo analysis may include a) activation-dependent epitopes of the membrane glycoprotein (GP) IIb/IIIa (CD41a) receptor, as demonstrated by the binding of activation-specific monoclonal antibodies (MoAbs) PAC1, anti-LIBS1 and anti-RIBS); b) the expression of P-selectin (CD62p), the alpha-granule GP translocated to the platelet surface following release reaction; and c) platelet procoagulant activity, as demonstrated by the binding of i) annexin V protein to the prothrombinase-complex (prothrombin, activated factor X (Xa) and V (Va)) binding sites on the surface of activated platelets, and of ii) MoAbs against activated coagulation factors V and X bound to the surface of activated platelets. Using this method, platelet activation as a marker for in vivo prothrombotic activity can be demonstrated in various clinical conditions including coronary angioplasty, orthostatic challenge in primary depression, sickle cell disease in clinical remission and during pain episode, and in pregnancy-related hypertension with marked increase during preeclampsia. The finding of platelet procoagulant activity is corroborated by increased levels of plasma markers for thrombin generation and fibrinolytic activity. 相似文献
12.
Srihirun S Sriwantana T Unchern S Kittikool D Noulsri E Pattanapanyasat K Fucharoen S Piknova B Schechter AN Sibmooh N 《PloS one》2012,7(1):e30380
BACKGROUND: Nitrite is a nitric oxide (NO) metabolite in tissues and blood, which can be converted to NO under hypoxia to facilitate tissue perfusion. Although nitrite is known to cause vasodilation following its reduction to NO, the effect of nitrite on platelet activity remains unclear. In this study, the effect of nitrite and nitrite+erythrocytes, with and without deoxygenation, on platelet activity was investigated. METHODOLOGY/FINDING: Platelet aggregation was studied in platelet-rich plasma (PRP) and PRP+erythrocytes by turbidimetric and impedance aggregometry, respectively. In PRP, DEANONOate inhibited platelet aggregation induced by ADP while nitrite had no effect on platelets. In PRP+erythrocytes, the inhibitory effect of DEANONOate on platelets decreased whereas nitrite at physiologic concentration (0.1 μM) inhibited platelet aggregation and ATP release. The effect of nitrite+erythrocytes on platelets was abrogated by C-PTIO (a membrane-impermeable NO scavenger), suggesting an NO-mediated action. Furthermore, deoxygenation enhanced the effect of nitrite as observed from a decrease of P-selectin expression and increase of the cGMP levels in platelets. The ADP-induced platelet aggregation in whole blood showed inverse correlations with the nitrite levels in whole blood and erythrocytes. CONCLUSION: Nitrite alone at physiological levels has no effect on platelets in plasma. Nitrite in the presence of erythrocytes inhibits platelets through its reduction to NO, which is promoted by deoxygenation. Nitrite may have role in modulating platelet activity in the circulation, especially during hypoxia. 相似文献
13.
By means of CM-Sephadex C-50 column chromatography and gel filtration on Sephadex G-75 and G-50 columns, a potent platelet aggregation inhibitor was purified and characterized. It was a glycoprotein with a molecular weight of 31,000. It was devoid of phospholipase A, ADPase, esterase and fibrino(geno)lytic activities. It inhibited dose-dependently the aggregation of washed platelets induced by collagen, thrombin, sodium arachidonate, platelet activating factor and ionophore A23187 with a similar IC50 (5-10 micrograms/ml). It was also active in platelet-rich plasma, with an IC50 of 10-15 micrograms/ml. The venom inhibitor reduced the elasticity of whole blood clot and inhibited the thrombin-induced clot retraction of platelet-rich plasma. These activities were related to its inhibitory activity on platelet aggregation rather than blood coagulation. The venom inhibitor had various effects on [14C]serotonin release stimulated by aggregation agonists. It had no effect on thromboxane B2 formation of platelets stimulated by sodium arachidonate, collagen and ionophore A23187. The presence of this venom inhibitor prior to the initiation of aggregation was a prerequisite for the maintenance of its maximal activity. It showed a similar inhibitory effect on collagen or thrombin-induced aggregation even when it was added after the platelets had undergone the shape change. High fibrinogen levels partially antagonized its activity. The venom inhibitor completely inhibited the fibrinogen-induced aggregation of alpha-chymotrypsin-treated platelets. It is concluded that this venom inhibitor interferes with the interaction of fibrinogen with fibrinogen receptors, leading to inhibition of aggregation. 相似文献
14.
Y P C?té J G Filep B Battistini J Gauvreau P Sirois A R Beaudoin 《Biochimica et biophysica acta》1992,1139(1-2):133-142
The inner layer of the aorta contains the enzyme ATP diphosphohydrolase (ATPDase: EC 3.6.1.5) which catalyzes the sequential phosphorolysis of ATP----ADP----AMP. Two zones of the inner layer, the intima and media, were separated and both were shown to contain ATPDase activity of similar specific activity (0.08 and 0.10 U/mg protein, respectively). However, the media exhibited about 100-times more enzyme activity than the intima. Both preparations were virtually identical with respect to pH optima (7.5), migration patterns after electrophoresis under non-denaturing conditions, relative rates of ATP and ADP hydrolysis and potency to inhibit ADP-induced platelet aggregation in both human platelet-rich plasma and whole blood. The IC50 values for ADP (2 microM)-induced aggregation were 6.8 and 12.9 mU/ml in platelet-rich plasma and whole blood, respectively. Addition of ATPDase to platelets pre-aggregated with ADP resulted in a dose-dependent disaggregation in platelet-rich plasma (IC50 4.9 mU/ml), but not in whole blood. When both ATPDase (5.6-58.7 mU/ml) and ATP (0.5-10 microM) were added to platelet-rich plasma, there was an immediate dose-dependent aggregation of platelets followed by a slowly developing disaggregation. These data show that ATPDase is present in both the intima and media layers of bovine aorta and suggest a dual role for this enzyme in platelet activation. By converting ATP released from damaged cells into ADP, the enzyme could facilitate platelet aggregation at the site of vascular injury, whereas the subsequent conversion of ADP to AMP could inhibit or reverse platelet aggregation. The consequence of these activities would be to control the growth of a platelet thrombus. 相似文献
15.
A E Rozenberg I L Bordiukova Kh M Markov 《Biulleten' eksperimental'no? biologii i meditsiny》1984,98(7):121-122
Three different methods of blood collection from rats are studied and compared, namely decapitation, catheterization of the carotid artery and puncture of the heart left ventricle. The latter method is preferable in studies of platelet aggregation. The method for isolation of washed platelets from rat blood is described in detail. The platelets stored for several hours at 20 degrees C did not lose the ability for aggregating under exposure to low concentrations of ADP. The curves of aggregation of washed and plasma platelets are provided. To study the rate of aggregation, a model is offered based on the approximation of the aggregation curves by the exponential A = A0exp (-alpha t). Such an approach made it possible to treat the data objectively and to reveal the features of platelet functional activity, particularly in arterial hypertension. 相似文献
16.
Pankaj Ganguly Harvey R. Bradford 《Biochimica et Biophysica Acta (BBA)/General Subjects》1982,714(2):192-199
The aggregation of human blood platelets by thrombin, adenosine diphosphate, wheat germ agglutinin or ristocetin was inhibited by primary amines. In general, thrombin-induced platelet aggregation was strongly affected by the amines while the effect was weak on cell aggregation by ristocetin. Usually, the diamines were stronger inhibitors of aggregation than the monoamines with cadaverine as the strongest and ethylamine as the weakest inhibitor. At concentration where platelet aggregation was inhibited, the amines neither displaced serotonin from serotinin-loaded platelets nor caused lysis of human red cells. The lectin activity of wheat germ agglutinin on human red cells was not affected by the amines indicating that the amines probably acted on platelets and not on the agglutinin. The clotting activity of thrombin on fibrinogen was partially inhibited by the amines while its esterolytic activity remained unaltered. The inhibitory action of the amines on platelet aggregation could be overcome with small amounts of calcium while other divalent cations tested had little effect. It is suggested that the amines affect platelet aggregation by interfering with the actions of membrane-associated calcium. 相似文献
17.
苗芳 《中国应用生理学杂志》1985,(4)
本文讨论了实验性缺氧对大鼠凝血机能产生的影响。实验结果表明,大鼠在模拟8000米高度两小时后,血小板凝聚性、血小板因子3活性明显增加,纤溶活性下降,同时,纤维蛋白原含量和因子X也明显下降。大鼠在模拟7000米经36天间歇性慢性缺氧,血小板计数、血小板凝聚性,血小板因子3活性、纤维蛋白原含量、因子X活性均显著增加,部分凝血活酶时间缩短、纤溶活性下降,明显地出现凝血增强的趋势。本文还讨论了抗缺氧药物复方党参、异叶青兰对人鼠急性缺氧时凝血机能的影响。 相似文献
18.
Construction and characterization of novel staphylokinase variants with antiplatelet aggregation activity and reduced immunogenecity 总被引:3,自引:0,他引:3
Su HB Zhang YG He JT Mo W Zhang YL Tao XM Song HY 《Acta biochimica et biophysica Sinica》2004,36(5):336-342
Thrombotic complication of cardiovascular diseases isa main cause of death and disability and consequently,thrombolysis favorably influences the outcome of such life-threatening diseases as myocardial infarction. Throm-bolytic agents are plasminogen activators that convertplasminogen, the inactive proenzyme in the blood, to pro-teolytic enzyme plasmin, which dissolves the fibrin ofblood clot [1]. Currently, thrombolytic agents approved orunder clinical investigation on patients with acute myo-… 相似文献
19.
K D Lomazova A M Poliakova O S Astrina D B Tsukerman 《Biulleten' eksperimental'no? biologii i meditsiny》1989,107(5):538-540
The effect of PAF in aggregation of platelets induced by endotoxin was studied in experiments in vitro. It is indicated that in high concentration (1.10(-7)-1.10(-6) M) PAF did not affect the degree of aggregation of platelets induced by lipopolysaccharides (LPS) S. typhimurium and N. meningitidis. Successive addition to PRP LPS and PAF or joint addition of PAF and LPS did not change the degree of aggregation of each inductor or their sum. A lower concentration of PAF (1.10(-11)-1.10(-9) M) and endotoxin caused a more expressive aggregation of platelets than their successive addition. Stimulating activity of PAF on endotoxin-induced aggregation, perhaps, is caused by involvement of metabolism of arachidonic acid during blood platelets activation. 相似文献
20.
A Dmoszyńska-Giannopoulou J Roliński B Soko?owska A Nowakowski 《Polski tygodnik lekarski (Warsaw, Poland : 1960)》1989,44(21-22):496-498
An effect of vitamin E on blood platelets functioning was studied in 39 patients with diabetes mellitus type 1. Control group included 20 healthy blood donors. Vitamin E in a daily dose of 1000 mg produced statistically significant decrease in platelets aggregation, number of circulating platelet aggregates and release of the platelet factory 4 in diabetics after 7 days of treatment. No adverse reactions were seen in any patient treated with vitamin E. The obtained results indicate that vitamin E inhibits increased platelets activity in the patients with diabetes mellitus type 1 and does not exert toxic reactions during the treatment. 相似文献