Factors responsible for ADP-induced release reaction of human platelets

1975 ◽  
Vol 228 (6) ◽  
pp. 1757-1765 ◽  
Author(s):  
JF Mustard ◽  
DW Perry ◽  
RL Kinlough-Rathbone ◽  
MA Packham
Keyword(s):  
Acetylsalicylic Acid ◽  
Sodium Citrate ◽  
Gamma Globulin ◽  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Tyrode Solution ◽  
Human Fibrinogen ◽  
Release Reaction ◽  
Heparin Plasma

Extensive aggregation of human platelets can be induced by ADP without secondaryaggregation or release of granule contents. This occurs with washed platelets in Tyrode solution containing 0.35% albumin, human fibrinogen, and apyrase, and in platelet-rich, heparin- or hirudin-plasma. Conditions that caused release during ADP-inducedaggregation were-citrate as the anticoagulant in platelet-rich plasma; addition of citrate (11-15 mM) to a suspension of washed platelets, or to hirudin-plasma or heparin-plasma; suspension of platelets in a medium containing magnesium but no calcium;and the presence of trace amounts of thrombin or aggregated gamma globulin in the platelet suspensions. Acetylsalicylic acid, phenylbutazone, or sulfinpyrazone inhibited secondary aggregation and release in all these circumstances. Heparin or hirudin inhibited ADP-INDUCED SECONDARY AGGREGATION AND RELEASE PROMOTED BY TRACES OF THROMBIN. Although fibrinogen is required for ADP-induced primary aggregation, it does not support secondary aggregation and release, provided that it has no clot-promoting activity. The main agent responsible for ADP-induced secondary aggregation and release in human, citrated, platelet-rich plasma appears to be sodium citrate. Suspending washed human platelets in a medium without calcium mimics the effect of citrate.

The Effects of Citrate and Extracellular Calcium Ions on the Platelet Release Reaction Induced by Adenosine Diphosphate and Collagen

1979 ◽  
Vol 42 (02) ◽  
pp. 778-793 ◽  
Author(s):  
Stanley Hepinstall ◽  
Patricia M Taylor
Keyword(s):  
Calcium Ions ◽  
Sodium Citrate ◽  
Platelet Rich Plasma ◽  
Adenosine Diphosphate ◽  
Human Platelets ◽  
Extracellular Calcium ◽  
Release Reaction ◽  
Calcium Dependent ◽  
Platelet Release ◽  
Dependent Mechanism

SummaryThe ADP-induced release of 3H-serotonin from human platelets in heparinized platelet rich plasma is markedly stimulated by the addition of sodium citrate. The aggregation and release that is induced by collagen is less affected by citrate. Data is presented that supports the view that the effects of citrate on both ADP-and collagen-induced release are largely via alteration of the concentration of ionized calcium in plasma.Collagen can induce release of 3H-serotonin via extracellular calcium-independent and -dependent mechanisms. The possibility that the calcium-dependent mechanism is aggregation-dependent and that the calcium is required for platelet aggregation rather than directly involved in the release reaction is discussed.

Reactions of Polylysine with Human Platelets in Plasma and in Suspensions of Washed Platelets

1976 ◽  
Vol 36 (02) ◽  
pp. 360-375 ◽  
Author(s):  
M. A Guccione ◽  
M. A Packham ◽  
R. L Kinlough-Rathbone ◽  
D. W Perry ◽  
J. F Mustard
Keyword(s):  
Acetylsalicylic Acid ◽  
Prostaglandin E1 ◽  
Creatine Phosphokinase ◽  
Platelet Rich Plasma ◽  
Creatine Phosphate ◽  
Human Platelets ◽  
Lag Phase ◽  
Tyrode Solution ◽  
Two Phases ◽  
Induced Aggregation

SummaryThe effects of polylysine on human platelets have been examined in citrated platelet-rich plasma (PRP) and in suspensions of washed platelets in various media. In PRP, polylysine caused aggregation after a lag phase. Heparin inhibited this completely. At certain concentrations of polylysine, two phases of aggregation occurred, the second being associated with release of 14C-serotonin from prelabelled platelets; this phase was inhibitable with prostaglandin E1, acetylsalicylic acid, sulphinpyrazone, adenosine, apyrase, or creatine phos- phate/creatine phosphokinase. Polylysine-induced release also occurred in PRP with EDTA or hirudin as anticoagulant. In suspensions of washed platelets in Tyrode solution containing 0.35% or 4% albumin, or 1% gelatin, polylysine caused immediate platelet-to-platelet adherence and very little release of 14C-serotonin or platelet lysis. Heparin inhibited aggregation, but acetylsalicylic acid, prostaglandin E1, adenosine, apyrase, creatine phosphate/creatine phosphokinase or EDTA did not. In a modified Tyrode-albumin medium containing 1 mM magnesium but no calcium, polylysine-induced aggregation was associated with the release of 14C-serotonin which could be inhibited by acetylsalicylic acid or indomethacin; this is similar to the effect of ADP in this medium. In Tyrode solution without albumin or gelatin, polylysine-induced platelet aggregation was associated with release of a large percentage of 14C-serotonin, together with as much as 18% lysis; indomethacin inhibited this release reaction.

Release, Aggregation and Lysis of Human Platelets by Antilymphocyte Globulin and Antiplatelet Serum

1976 ◽  
Vol 36 (02) ◽  
pp. 411-423 ◽  
Author(s):  
Nicholas Lekas ◽  
J. C Rosenberg
Keyword(s):  
Platelet Aggregation ◽  
Gamma Globulin ◽  
Human Platelets ◽  
Plasma Medium ◽  
Release Reaction ◽  
Clinical Events ◽  
Thrombotic Complications ◽  
Platelet Release ◽  
Free Media

SummaryHuman platelets labeled with 51Cr were used to determine the contribution made by platelet lysis to the platelet release reaction and platelet aggregation induced by rabbit antihuman platelet serum (APS) and equine antihuman thymocyte globulin (ATG). Platelets were tested in both plasma (PRP) and non-plasma containing media. Antibodies directed against platelets, either as APS or ATG, induced significant amounts of platelet release and aggregation, as well as some degree of lysis, in the absence of complement. The presence of complement increased platelet lysis and aggregation, but not the release reaction. Non-immune horse gamma globulin produced different responses depending upon whether platelets were investigated in PRP or non-plasma containing media. Aggregation was seen in the latter but not the former. These differences can be explained by the presence of plasma components which prevent non-specific immune complexes from causing platelet aggregation. Since platelets in vivo are always in a plasma medium, one must be wary of utilizing data from platelet studies in synthetic plasma-free media as the basis of explaining clinical events. These observations demonstrate at least two, and possibly three, different mechanisms whereby ATG could activate platelets causing thrombotic complications and thrombocytopenia, i.e., via 1) specific and, 2) non-specific non-lytic pathways and 3) a lytic pathway.

scholarly journals Fibrinogen-independent aggregation and deaggregation of human platelets: studies in two afibrinogenemic patients

Blood ◽  
1987 ◽  
Vol 70 (1) ◽  
pp. 221-226 ◽  
Author(s):  
M Cattaneo ◽  
RL Kinlough-Rathbone ◽  
A Lecchi ◽  
C Bevilacqua ◽  
MA Packham ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Plasma Proteins ◽  
Platelet Rich Plasma ◽  
Platelet Activating Factor ◽  
Adenosine Diphosphate ◽  
Human Platelets ◽  
Human Fibrinogen ◽  
Control Subjects ◽  
Platelet Aggregates ◽  
And Control

Abstract Platelets from two afibrinogenemic patients were used to determine whether fibrinogen is essential for platelet aggregation and to examine whether released fibrinogen contributes to the stabilization of platelet aggregates when platelets have been induced to aggregate and release their granule contents by stimulation with thrombin. The addition of adenosine diphosphate (ADP) to platelet-rich plasma (PRP) or to suspensions of washed platelets from the afibrinogenemic patients caused the formation of small aggregates, which was either not inhibited or only slightly inhibited by the F(ab')2 fragments of an antibody to fibrinogen but was inhibited by an antibody (10E5) to glycoprotein IIb/IIIa. Thus there is a component of ADP-induced platelet aggregation that is not dependent on fibrinogen or other plasma proteins but is dependent on glycoprotein IIb/IIIa. There was little difference in the extent of aggregation and the release of granule contents of normal and afibrinogenemic platelets in response to the release-inducing agents collagen, platelet-activating factor (PAF), sodium arachidonate, or thrombin. With normal or afibrinogenemic platelets, aggregation by thrombin (0.2 U/mL or higher) was not inhibited by the F(ab')2 fragments of an antibody to human fibrinogen. Deaggregation by combinations of inhibitors of platelets aggregated by 1 U/mL thrombin showed no difference between platelets from afibrinogenemic and control subjects, indicating that released fibrinogen does not make a major contribution to the stabilization of platelet aggregates formed by thrombin stimulation.

scholarly journals Thromboxane A2 causes feedback amplification involving extensive thromboxane A2 formation on close contact of human platelets in media with a low concentration of ionized calcium

Blood ◽  
1987 ◽  
Vol 70 (3) ◽  
pp. 647-651 ◽  
Author(s):  
MA Packham ◽  
RL Kinlough-Rathbone ◽  
JF Mustard
Keyword(s):  
Calcium Salt ◽  
Close Contact ◽  
Platelet Rich Plasma ◽  
Thromboxane A2 ◽  
Adenosine Diphosphate ◽  
Human Platelets ◽  
Tyrode Solution ◽  
Low Concentration ◽  
Thromboxane Receptor ◽  
Aggregation Response

Abstract Close platelet-to-platelet contact induced by weak agonists in a medium with a low concentration of Ca2+ leads to thromboxane A2 (TXA2) formation, release of granule contents, and secondary aggregation. These responses do not occur in a medium containing Ca2+ in the physiological range (1 to 2 mmol/L). Experiments were done to determine whether feedback amplification is required to generate amounts of TXA2 that are sufficient to cause secondary aggregation and the reactions associated with it, or whether close platelet-to-platelet contact alone is sufficient to generate enough TXA2 to produce these responses. Platelets were washed and resuspended in a modified Tyrode solution to which no calcium salt was added that contained 0.35% albumin and apyrase. This medium contains 20 mumol/L Ca2+ and 1 mmol/L Mg2+. Platelets were aggregated with adenosine diphosphate (ADP) in the presence of fibrinogen, agglutinated with polylysine, or after pretreatment with chymotrypsin, aggregated with fibrinogen. In the low- Ca2+ medium, all these agonists caused platelets to adhere to each other, followed by secondary aggregation with TXA2 formation and release of granule contents. When Ca2+ (1 to 2 mmol/L), aspirin, or the thromboxane receptor blocker BM 13.177 was present, the secondary responses did not occur; dazoxiben decreased thromboxane formation, but did not prevent secondary aggregation or release. Aspirin-treated platelets were less responsive to ADP, U46619, or TXA2 in the low-Ca2+ medium, which indicated that the secondary responses of untreated platelets were not caused by a generalized increase in sensitivity. The reactions that result from close platelet-to-platelet contact in a low- Ca2+ medium can be caused by a wide variety of weak agonists; the secondary aggregation response and release of granule contents are dependent on TXA2 formation and on feedback amplification by TXA2 or the prostaglandin endoperoxides. The secondary responses caused by weak agonists in citrated platelet-rich plasma (which has a concentration of Ca2+ similar to the low-Ca2+ medium used in the present studies) do not occur at the concentration of Ca2+ in circulating blood and thus may have little biologic relevance.

scholarly journals Platelet release reaction and intracellular cGMP

Blood ◽  
1978 ◽  
Vol 52 (3) ◽  
pp. 524-531 ◽  
Author(s):  
A Weiss ◽  
NL Baenziger ◽  
JP Atkinson
Keyword(s):  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Serotonin Release ◽  
Secretory Process ◽  
Release Reaction ◽  
Intracellular Cgmp ◽  
Exogenous Addition ◽  
Human Thrombin ◽  
Platelet Release

Abstract Enchanced cAMP concentrations inhibit the aggregation and release reaction of isolated human platelets and platelet-rich plasma to all known inducing agents. An opposing role for cGMP in this phenomenon has been proposed by some but not by others, and the function of cGMP in this secretory process is unclear. To further elucidate the role of cGMP in the release reaction, the effect of increased concentrations of this cyclic nucleotide on 14C-serotonin release was evaluated utilizing isolated human platelets and highly purified human thrombin or commercially available bovine thrombin. Several recently described stimulators of guanylate cyclase, including sodium nitroprusside, sodium azide, nitrosoquanidines, and ascorbic acid, were found to markedly augment platelet cGMP levels. Enhanced platelet cGMP concentrations produced by these drugs or by the exogenous addition of cGMP and its analogues neither caused these cells to secrete nor modulated the thrombin-induced serotonin release reaction. The inhibition of serotonin release by increased cAMP concentrations was not counteracted by increased cGMP levels. Platelet cGMP concentrations were unaltered by thrombin. These data indicate that cGMP is not an obligatory signal or a modulator of the thrombin-induced platelet release reaction.

Fibrin-induced release of platelet serotonin

1976 ◽  
Vol 231 (2) ◽  
pp. 344-350 ◽  
Author(s):  
KG Orloff ◽  
D Michaeli
Keyword(s):  
Platelet Aggregation ◽  
Prostaglandin E1 ◽  
Human Platelets ◽  
The Self ◽  
Human Fibrinogen ◽  
Release Reaction ◽  
Activated Platelets ◽  
Platelet Serotonin ◽  
Platelet Release ◽  
Adhesion Of Platelets

Human platelets were reacted with polymerized fibrin formed from human fibrinogen. The platelets adhered to the fibrin particles and this adhesion was followed by the release of serotonin from prelabeled platelets. The adhesion of platelets to fibrin was not inhibited by adenosine or prostaglandin E1. However, the subsequent Ca2+-dependent release of platelet serotonin was completely inhibited by these compounds. After the initial platelet-fibrin interaction, ADP and serotonin released from activated platelets may lead to additional platelet aggregation and release. Therefore, in addition to clot stabilization, fibrin serves as an initiator of the platelet release reaction. This in turn initiates the self-amplifying process of platelet aggregation.

scholarly journals Effect of anti-P1A1 antibody on human platelets. I. The role of complement

Blood ◽  
1979 ◽  
Vol 53 (4) ◽  
pp. 567-577 ◽  
Author(s):  
DB Cines ◽  
AD Schreiber
Keyword(s):  
Platelet Aggregation ◽  
Complement Activation ◽  
Platelet Rich Plasma ◽  
Plasma Concentrations ◽  
Human Platelets ◽  
Serotonin Release ◽  
Buffer System ◽  
Induce Platelet Aggregation ◽  
Platelet Surface ◽  
Release Reaction

Abstract We studied the interaction of complement with human platelets. Complement was activated by IgG anti-P1A1 antibody obtained from 3 patients with the post-transfusion purpura syndrome. We used a heparin- plasma buffer system that permits complement activation and also preserves platelet function. With this system complement activation was efficient, and platelet immune alteration was extensive. Anti-P1A1 antibody was effective only in the presence of complement, in which case both platelet lysis and serotonin release (release reaction) in the absence of lysis were observed. Platelet lysis, as assessed by 51Cr loss, required 10-fold more antibody than was necessary to induce platelet aggregation and release of 14C-serotonin. This platelet release reaction required an intact classic complement sequence through C6. The extent of platelet serotonin release parallelled the depletion of C1 and C4 from platelet-rich plasma. Concentrations of antibody insufficient to induce platelet aggregation and serotonin release could still activate C1 and deposit increased C3 on the platelet surface. These studies demonstrated that complement activation by anti-P1A1 antibody can alter human platelets in a nonlytic system. Several phases of complement-mediated human platelet alteration are possible, depending on the concentration of anti-P1A1 antibody.

scholarly journals Evaluation of Highly Purified Bovine F. VIII and Bovine Plasma as Reagents to Induce Platelet Aggregation

1977 ◽  
Author(s):  
M. A. Lazzari ◽  
C. Simonetti ◽  
G. Casillas ◽  
M. Pavlovsky
Keyword(s):  
Platelet Aggregation ◽  
Optical Density ◽  
Sodium Citrate ◽  
Platelet Rich Plasma ◽  
Human Platelets ◽  
Induce Platelet Aggregation ◽  
Bovine Plasma ◽  
Aggregation Factor ◽  
High Concentrations

Purified bovine F. VIII is a known reagent for human platelets aggregation used for the study of thrombopaties. As bovine plasma (BP)is easier to prepare and standardize it was studied as an alternative reagent, comparing it with purified bovine F. VIII. Platelet aggregation was studied by a turbidimetric technique(Aggregometer Bryston AG 1). The substrates were platelet rich plasma (PRP)and gel filtered platelets using between 150,000 and 200,000 platelets/mm3 to standardize the platelets surface. 1/10 dilution of bovine plasma was found to be the best for PRP and 1/1 for filtered platelets. Incubation of the substrates with bovine plasma at 37°C for 5 min seems to enhance PAF activity of the platelet aggregation factor (PAF). Primary aggregation was obtained in PRP treated with 2% tetrasodic EDTA, 3. 4% sodium citrate and in plasma with aspirin. No secondary aggregation was observed in EDTA or aspirin plasma. Unlike platelets treated with ADP, their shape did not change since the optical density was not modified. No synergism or competition between PAF and ADP or adrenalin were found. High concentrations of NaCl or urea interfere with PAF, and esposure to 56°C inactivates it. In 50 normal PRP the percentage of total aggregation were: BP 1/10 42% ± 12 ; BP 1/1 79% ± 13 ;purified bovine F. VIII 1/10 86% ± 6 and in 10 batches of filtered platelets bp 1/1 82% ± 7. We consider an advantage to use bovine plasma for platelets aggregation studies.

EFFECTS OF HUMAN ATRIAL NATRIURETIC PEPTIDES ON SECRETION REACTION IN HUMAN PLATELETS

1987 ◽  
Author(s):  
A Tanable ◽  
Y Yatomi ◽  
T Ohashi ◽  
H Oka ◽  
T Kariya ◽  
...  
Keyword(s):  
Platelet Rich Plasma ◽  
Atp Release ◽  
Smooth Muscle Contraction ◽  
Inhibitory Effect ◽  
Human Platelets ◽  
Serotonin Release ◽  
Release Reaction ◽  
Induced Aggregation ◽  
Atrial Natriuretic

Human atrial natriuretic peptide (h-ANP) has vasodilating and natriuretic properties, and inhibits smooth muscle contraction, renal renin secretion and adrenal aldosterone release. Although Schiffrin has reported that human platelets have receptors for ANP, its effects in platelets are not established in vivo. We therefore investigated the influence of h-ANP on secretion reaction in human platelets. Eight healthy subjects, males, aged 20 to 24 years, donated blood for the study. Citrated platelet-rich plasma (PRP) was incubated with or without h-ANP at 37 C for 2.5 minutes. The samples of 0.5 ml PRP then used to measure ADP induced aggregation, ATP release reaction and C-serotonin release reaction. H-ANP, at concentration of 1x10 -6M, decreased ADP induced aggregation (after h-ANP: 77.4±9.7 % of control aggregation), and inhibited ATP release reaction (after h-ANP: 31.8±13.1%). Serotonin releasereaction induced by ADP was also inhibited ( control: 15.3±2.2%, after h-ANP: 8.3±0.5 %). The inhibitory effect of h-ANP on aggregation and secretion reaction was maximal by 3 minutes. These data suggest that h-ANP inhibits secretion reaction in human platelets.

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