Hydrogen Peroxide (H2O2) an Inhibitor of the Platelet Release Reaction

1975 ◽  
Author(s):  
M. J. Stuart ◽  
H. Holmsen ◽  
F. A. Oski
Keyword(s):  
Steady State ◽  
Platelet Rich Plasma ◽  
Adenosine Diphosphate ◽  
Minute Period ◽  
Bacterial Interaction ◽  
Steady State Levels ◽  
In Vivo Effects ◽  
Induced Aggregation

The effect of H2O2 on platelet metabolism, aggregation and release was studied by the in vitro exposure of 3H-adenine labelled platelet rich plasma (PRP) to H2O2. On incubation of PRP with H2O2 (100 to 500 μM final cone.) over a 30 minute period, there was a mean drop of 38% in the baseline steady state levels of radioactive metabolic ATP, the fall occurring in the first 3 minutes of incubation, with a corresponding increase in the levels of radioactive inosinemonophosphate and hypoxanthine. This was not a nonspecific lytic effect on the platelet since no extracellular leakage of platelet nucleotides occurred during the incubation. Further, mean decreases of 8 to 38% in steady state levels of platelet metabolic ATP were observed during incubations of PRP with 5 to 500 μM H2O2 respectively. Finally, the action of H2O2 on adenosine diphosphate (ADP) induced biphasic aggregation and release was studied in PRP preincubated for 3 minutes with H2O2 (100 to 500 μM). Partial inhibition of the primary wave, and complete inhibition of the second wave of ADP induced aggregation was observed in the H2O2 pretreated platelets, con-commitant with inhibition of release of platelet non-metabolic ATP and ADP, when compared to the control saline preincubated platelets. Since H2O2 is generated in vivo by bacteria and leucocytes during phagocytosis, the in vivo effects of the described inhibition of H2O2 and its possible role in platelet-leucocyte-bacterial interaction requires further elucidation.

RAP1 is required for BAS1/BAS2- and GCN4-dependent transcription of the yeast HIS4 gene

1991 ◽  
Vol 11 (7) ◽  
pp. 3642-3651 ◽  
Author(s):  
C Devlin ◽  
K Tice-Baldwin ◽  
D Shore ◽  
K T Arndt
Keyword(s):  
Steady State ◽  
Binding Site ◽  
Binding Sites ◽  
Binding Activity ◽  
Micrococcal Nuclease ◽  
Mrna Levels ◽  
High Affinity ◽  
Steady State Levels

The major in vitro binding activity to the Saccharomyces cerevisiae HIS4 promoter is due to the RAP1 protein. In the absence of GCN4, BAS1, and BAS2, the RAP1 protein binds to the HIS4 promoter in vivo but cannot efficiently stimulate HIS4 transcription. RAP1, which binds adjacently to BAS2 on the HIS4 promoter, is required for BAS1/BAS2-dependent activation of HIS4 basal-level transcription. In addition, the RAP1-binding site overlaps with the single high-affinity HIS4 GCN4-binding site. Even though RAP1 and GCN4 bind competitively in vitro, RAP1 is required in vivo for (i) the normal steady-state levels of GCN4-dependent HIS4 transcription under nonstarvation conditions and (ii) the rapid increase in GCN4-dependent steady-state HIS4 mRNA levels following amino acid starvation. The presence of the RAP1-binding site in the HIS4 promoter causes a dramatic increase in the micrococcal nuclease sensitivity of two adjacent regions within HIS4 chromatin: one region contains the high-affinity GCN4-binding site, and the other region contains the BAS1- and BAS2-binding sites. These results suggest that RAP1 functions at HIS4 by increasing the accessibility of GCN4, BAS1, and BAS2 to their respective binding sites when these sites are present within chromatin.

Aspirin-Dependent Effects on Purinergic P2Y1 Receptor Expression

2019 ◽  
Vol 119 (05) ◽  
pp. 726-734 ◽  
Author(s):  
Isabella Massimi ◽  
Laura Alemanno ◽  
Maria Guarino ◽  
Raffaella Guerriero ◽  
Massimo Mancone ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Thromboxane A2 ◽  
Adenosine Diphosphate ◽  
Receptor Expression ◽  
P2y1 Receptor ◽  
Biochemical Pathway ◽  
Thromboxane A2 Receptor ◽  
Induced Aggregation

AbstractChronic treatment with aspirin in healthy volunteers (HVs) is associated with recovery of adenosine diphosphate (ADP)-induced platelet activation. The purinergic P2Y1 receptor exerts its effects via a Gq-protein, which is the same biochemical pathway activated by thromboxane-A2 receptor. We hypothesized that recovery of ADP-induced platelet activation could be attributed to increased P2Y1 expression induced by chronic aspirin exposure. We performed a multi-phase investigation which embraced both in vitro and in vivo experiments conducted in (1) human megakaryoblastic DAMI cells, (2) human megakaryocytic progenitor cell cultures, (3) platelets obtained from HVs treated with aspirin and (4) platelets obtained from aspirin-treated patients. DAMI cells treated with aspirin or WY14643 (PPARα agonist) had a significant up-regulation of P2Y1 mRNA, which was shown to be a PPARα-dependent process. In human megakaryocytic progenitors, in the presence of aspirin or WY14643, P2Y1 mRNA expression was higher than in mock culture. P2Y1 expression increased in platelets obtained from HVs treated with aspirin for 8 weeks. Platelets obtained from patients who were on aspirin for more than 2 months had increased P2Y1 expression and ADP-induced aggregation compared with patients on aspirin treatment for less than a month. Overall, our results suggest that aspirin induces genomic changes in megakaryocytes leading to P2Y1 up-regulation and that PPARα is the nuclear receptor involved in this regulation. Since P2Y1 is coupled to the same Gq-protein of thromboxane-A2 receptor, platelet adaptation in response to pharmacological inhibition seems not to be receptor specific, but may involve other receptors with the same biochemical pathway.

Inhibition of Human Platelet Aggregation by the Proteolytic Effect of Streptokinase (SK). Role of the Factor VIII Related Antigen

1975 ◽  
Author(s):  
R. Castillo ◽  
S. Maragall ◽  
J. A. Guisasola ◽  
F. Casals ◽  
C. Ruiz ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Factor Viii ◽  
Platelet Rich Plasma ◽  
Gel Filtration ◽  
Inhibitory Effect ◽  
Factor Viii Related Antigen ◽  
Human Factor Viii ◽  
Induced Aggregation

Defective ADP-induced platelet aggregation has been observed in patients treated with streptokinase. This same effect appears “in vitro” when adding SK to platelet rich plasma (PRP). Classic hemophilia and normal platelet poor plasmas (PPP) treated with SK inhibit the aggregation of washed platelets; plasmin-treated normal human serum also shows an inhibitory effect on platelet aggregation. However, von Willebrand SK-treated plasmas do not inhibit the aggregation of washed platelets. The same results appear when plasmas are previously treated with a rabbit antibody to human factor VIII.This confirms that the antiaggregating effect is mainly linked to the digested factor VIII related antigen.The inhibition of ADP-induced platelet aggregation has been proved in gel filtration-isolated and washed platelets from SK-treated PRP.Defective ristocetin-induced platelet aggregation has also been observed- This action does not appear in washed platelets from SK-treated PRP in presence of normal PPP, but it does in presence of SK-treated PPP, which suggests that the inhibition of the ristocetin-induced aggregation is due to the lack of factor VIII and not to the factor VIII-related products.Heparin, either “in vivo” or “in vitro”, has corrected the antiaggregating effect of SK.

The Effects Of Prostacyclin (PGI2) On Intravascular Platelet Aggregation

1981 ◽  
Author(s):  
G G Duncan ◽  
G Mallarkey ◽  
G M Smith
Keyword(s):  
Platelet Count ◽  
Guinea Pigs ◽  
Adenosine Diphosphate ◽  
Before And After ◽  
Dependent Inhibition ◽  
Induced Aggregation ◽  
Dose Dependent ◽  
Anaesthetised Rats

Intravascular aggregation can be measured by counting the number of circulating platelets before and after the injection of aggregation agents. The Technicon Autocounter was modified to count platelets continuously and connected via a double cannula in a carotid artery to an anaesthetised animal.Adenosine diphosphate (ADP) and collagen gave dose- dependent falls in the circulating platelet count when injected into rats, guinea pigs and rabbits. This enabled aggregation to be accurately quantitated in vivo.The infusion of PGI2 (0.25-1 ug/kg/min) in anaesthetised rats and rabbits produced a dose-dependent inhibition of the fall in platelet count produced by ADP and collagen. The formation of PGI2 can be inhibited in vitro by 15- hydroperoxyarachidonic acid (15HPAA). When 20 ug/kg/min of 15HPAA was infused into rats, aggregation produced by collagen was significantly increased suggesting that PGI2 is continuously formed by the rat vascular endothelium. This observation was confirmed by infusing 6-keto PGF1α antiserum. This antibody also prevented the inhibitory activity of PGI2 on collagen-induced aggregation. The study of continuous platelet counting in guinea pigs has been hampered by the occurrence of thrombocytopenia in certain animals. When 2 ug/kg/min of PGI2 was infused for 10 mins, a rise in the circulating platelet count to a steady plateau 4-5 × 105 platelets occurredThese experiments have shown that PGI2 will prevent aggregation by ADP and collagen and will reverse spontaneous thrombocytopenia and that PGI2 is continuously released from the vessels of anaesthetised rats.

scholarly journals Zinc-induced platelet aggregation is mediated by the fibrinogen receptor and is not accompanied by release or by thromboxane synthesis

Blood ◽  
1985 ◽  
Vol 66 (1) ◽  
pp. 213-219 ◽  
Author(s):  
P Heyns A du ◽  
A Eldor ◽  
R Yarom ◽  
G Marx
Keyword(s):  
Platelet Aggregation ◽  
Dense Body ◽  
Platelet Rich Plasma ◽  
Adenosine Monophosphate ◽  
Adenosine Diphosphate ◽  
Creatine Phosphate ◽  
Calcium Flux ◽  
Fibrinogen Receptor ◽  
Induced Aggregation

Abstract We demonstrate that zinc (0.1 to 0.3 mmol/L) induces aggregation of washed platelet suspensions. Higher concentrations (1 to 3 mmol/L) of zinc were needed to aggregate platelets in platelet-rich plasma obtained from blood anticoagulated with low-molecular-weight heparin, probably due to the binding of zinc to the plasma proteins. Zinc- induced aggregation of normal washed platelets required added fibrinogen and no aggregation occurred with thrombasthenic platelets or with normal platelets pretreated with a monoclonal antibody (10E5) that blocks the platelet fibrinogen receptor. These data indicate that the platelet membrane fibrinogen receptor-glycoproteins IIb and IIIa mediate the effect of zinc. Zinc-induced aggregation was blocked by the agent TMB-8, which interferes with the internal calcium flux, and by prostacyclin, which elevates platelet cyclic adenosine monophosphate levels. Zinc-induced aggregation was not accompanied by thromboxane synthesis or by the secretion of dense-body serotonin and was not affected by preexposure of platelets to acetylsalicylic acid. Experiments with creatine phosphate/creatine phosphokinase showed that the zinc effect on platelets was independent of extracellular adenosine diphosphate (ADP). Zinc had an additive effect when platelet aggregation was stimulated with subthreshhold concentrations of collagen or ADP. Together with the known effects of nutritional zinc on in vivo bleeding, on platelet aggregation, and on lipid metabolism, the results suggest that zinc may have an important bearing on normal hemostasis, thrombosis, and atherosclerosis.

The pharmacology of L-670,596, a potent and selective thromboxane/prostaglandin endoperoxide receptor antagonist

1989 ◽  
Vol 67 (9) ◽  
pp. 989-993 ◽  
Author(s):  
A. W. Ford-Hutchinson ◽  
Y. Girard ◽  
A. Lord ◽  
T. R. Jones ◽  
M. Cirino ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Guinea Pig ◽  
Receptor Antagonist ◽  
Competitive Inhibition ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Prostaglandin Endoperoxide ◽  
Induced Aggregation

L-670,596 ((−)6,8-difluoro-9-p-methylsulfonyl benzyl-1,2,3,4-tetrahydrocarbazol-1-yl-acetic acid) has been shown to be a potent receptor antagonist as evidenced by the inhibition of the binding of 125I-labeled PTA-OH to human platelets (IC50, 5.5 × 10−9 M), inhibition of U-44069 induced aggregation of human platelet rich plasma (IC50, 1.1 × 10−7 M), and competitive inhibition of contractions of the guinea pig tracheal chain induced by U-44069 (pA2,9.0). The compound was also active in vivo as shown by inhibition of arachidonic acid and U-44069 induced bronchoconstriction in the guinea pig (ED50 values, 0.04 and 0.03 mg/kg i.v., respectively), U-44069 induced renal vasoconstriction in the pig (ED50, 0.02 mg/kg i.v.), and inhibition of ex vivo aggregation of rhesus monkey platelets to U-44069 (active 1–5 mg/kg p.o.). The selectivity of the compound was indicated by the failure to inhibit, first, ADP-induced human or primate platelet aggregation and, second, bronchoconstriction in the guinea pig in vivo and contraction of the guinea pig tracheal chain in vitro to a variety of agonists. It is concluded that L-670,596 is a potent, selective, orally active thromboxane A2/prostaglandin endoperoxide receptor antagonist.Key words: thromboxane A2, thromboxane antagonist, prostaglandin endoperoxides, platelet aggregation.

scholarly journals Do patients with thromboembolic disease have circulating platelet aggregates?

Blood ◽  
1984 ◽  
Vol 64 (1) ◽  
pp. 205-209 ◽  
Author(s):  
FH Kohanna ◽  
MH Smith ◽  
EW Salzman
Keyword(s):  
Platelet Rich Plasma ◽  
Venous Blood ◽  
Adenosine Diphosphate ◽  
Normal Subjects ◽  
Thromboembolic Disease ◽  
Circulating Platelet Aggregates ◽  
Platelet Aggregates ◽  
Lower Ratio

Reports of circulating platelet aggregates (ie, microemboli) in thromboembolism and other vascular disorders are based on a method (Wu and Hoak , 1974) in which venous blood is collected via scalp vein needle and tubing into either formaldehyde, which fixes aggregates, or EDTA, which disperses them. The ratio of platelet counts in platelet- rich plasma (PRP) from the two blood samples after centrifugation is interpreted as a measure of platelet aggregates in the circulation in vivo. We compared this standard Wu and Hoak technique with a modified one, in which blood was drawn directly into a syringe, and with a third method that avoided centrifugation by counting single platelets in whole blood. Both modified techniques could detect aggregates generated in vitro with adenosine diphosphate (ADP). In 12 normal subjects, the three methods were equivalent, but in 37 patients with thromboembolic disorders, the standard Wu and Hoak method gave a lower ratio than the other methods. Similar results were found in a subset of eight patients with myocardial infarction. Heparin treatment of patients did not influence the results. The data suggest that formation of platelet aggregates occurred during venipuncture. Platelets may be hyperactive in patients with thromboembolic disease and may form aggregates in vitro during collection, but the concept of chronic microembolism in such patients should be reassessed.

In Vivo Effects of a Low Molecular Weight Heparin Fragment on Platelet Aggregation and Platelet Dependent Hemostasis in Dogs

1988 ◽  
Vol 60 (02) ◽  
pp. 232-235 ◽  
Author(s):  
B Ljungberg ◽  
H Johnsson
Keyword(s):  
Molecular Weight ◽  
Platelet Aggregation ◽  
Low Molecular Weight Heparin ◽  
Platelet Rich Plasma ◽  
Skin Flap ◽  
Low Molecular Weight ◽  
Dog Plasma ◽  
In Vivo Effects ◽  
Induced Aggregation

SummaryPlasma defibrinogenated dogs were used to study the influence of conventional heparin and a low molecular weight heparin fragment (Fragmin®, mean MW 5,000 d) on platelet dependent hemostasis. The heparins were given intravenously in gravimetri- cafly equal doses. The bleeding from standardized skin flap wounds and platelet aggregation (ADP and thrombin) was studied. In comparison, higher doses of the fragment than of heparin were required to increase the bleeding. ADP-induced aggregation in defibrinogenated platelet rich plasma (after addition of normal dog plasma) was potentiated by both heparins. After injection of heparin or the fragment, ADP induced platelet aggregation without prior addition of normal plasma to the testtube.In conclusion the heparin fragment affected bleeding to a less extent than conventional heparin. One explanation might be a weaker inhibition of thrombin-induced platelet aggregation.

In Vitro and In Vivo Effects of Adrenaline and Phentolamine on Platelet Function

1978 ◽  
Vol 40 (02) ◽  
pp. 428-438 ◽  
Author(s):  
Oreste Ponari ◽  
Emilio Civardi ◽  
Alessandro Megha ◽  
Mario Pini ◽  
Raffaele Poti’ ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
Threshold Concentration ◽  
Two Phase ◽  
Platelet Factor ◽  
Induce Platelet Aggregation ◽  
In Vivo Effects ◽  
In Vivo Administration

Summary In vitro and in vivo effects of adrenaline (ADR) on platelet aggregation, on platelet factor 3 (PF3) availability and on platelet factor 4 (PF4) release were studied in man. Inhibitory action of an alpha-blocker, phentolamine (PHEN) was investigated in the same conditions.The threshold concentration (TC) of ADR inducing the typical two-phase response in aggregation tests when added to platelet-rich plasma (PRP) varied in different pools of plasma, but always induced an evident PF4 release and increased PF3 availability. A further increase in both parameters was obtained with higher concentrations but without any significant dose/response correlation.Adding PHEN alone to PRP did not induce platelet aggregation or modify PF4 release induced by stirring, but it reduced PF3 availability. On the other hand, PHEN prevented the effects of ADR in different platelet tests, at appropriate concentrations.Intravenous infusion of ADR lowered the TC, and increased PF3 availability and PF4 release. In vivo administration of PHEN, in contrast, increased TC and reduced PF3 availability, while PF4 remained unchanged.

Lu 23051, A New Potent Inhibitor Of Platelet Aggregation

1981 ◽  
Author(s):  
H D Lehmann ◽  
J Gries ◽  
D Lenke
Keyword(s):  
Platelet Aggregation ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Coronary Vessels ◽  
Inhibitory Effect ◽  
Spontaneously Hypertensive ◽  
Dependent Inhibition ◽  
Induced Aggregation

6- [p-(2-(Chiorpropionylamino)phenyl] -4.5-dihydro-5-methyl-3(2H)-pyridazinone, LU 23051, is primarily characterized by its strong inhibition of platelet aggregation under in vitro and in vivo conditions. In vitro there is a concentration-dependent inhibition of ADP and collagen induced aggregation in platelet rich plasma of man, rat and dog. The inhibitory concentration EC 33 % is 0.0010-0.030 mg/1 (man: ADP-0.030, col 1.-0.013 mg/l) depending on species and type of aggregation. When administered orally in ex vivo experiments on rats and dogs the substance is found to have a dose-dependent antiaggregatory effect in the range from 0.1-3.16 mg/kg. The ED 33 % is 0.27-0.63 mg/kg.-In addition after oral administration the substance has a good inhibitory effect in models being based on intravascular platelet aggregation. Thus, a dose of 1 mg/kg inhibits laser-induced aggregation in mesenteric venules of rats. Mortality after i.v. injection of collagen in mice is reduced by 50 % after a dose of 0.02 mg/kg. A dose of 0.039 mg/kg prolongs the bleeding time of rats by 50 %. The aggregation-inhibiting action is of long duration (0.1 mg/kg p.o.∼24 h). The substance does not interfere with clotting.Besides its effect on platelet aggregation LU 23051 acts as vasodilatator as well. Dilatation of coronary vessels by 100 % is seen in isolated guinea-pig hearts at a concentration of 0.1 mg/l. In spontaneously hypertensive rats the substance has an anti hypertensive effect. The ED 20 % is 0.36 mg/kg p.o.The combination of antiaggregatory and vasodilatatory effects opens up interesting aspects with respect to the pharmacotherapeutic use of the new substance

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