The norpurpureine alkaloid from Annona purpurea inhibits human platelet activation in vitro

SummaryThe effect of pyridoxal 5’-phosphate (PALP) and trifluoperazine (TFPZ), the calmodulin antagonist, on in vitro platelet adhesion to collagen and collagen-induced platelet activation was studied using platelet-rich-plasma (PRP) or washed platelets (WPL). Platelet aggregation and [14C]-5HT release induced by “threshold” or low concentrations of collagen (0.6 μg/ ml) in PRP were completely abolished by PALP (24 mM), TFPZ (250 μM) as well as indomethacin (10 μM). At higher concentrations of collagen (10–15 μg/ml) in PRP and WPL, the use of stirred and unstirred platelets treated with collagen enabled a distinction to be made between aggregation and adhesion- mediated release reaction. Platelet aggregation and the aggregation-mediated release reaction induced by these concentrations of collagen in stirred platelets were completely abolished by PALP, TFPZ and indomethacin although neither adhesion to collagen nor the adhesion-mediated release reaction of unstirred platelets was significantly affected by these inhibitors. Interestingly, both adhesion and the adhesion-mediated release reaction were abolished by concentrations of PALP 10–40 fold higher than those required to abolish aggregation. Collagen-induced platelet aggregation, but not platelet adhesion, was inhibited in resuspended platelets pretreated with PALP and NaBH4 indicating a separation in the membrane sites involved in aggregation and adhesion. The results further emphasize the distinction between adhesion and aggregation-mediated events with regards to collagen with the latter being more susceptible to inhibition by antiplatelet agents such as PALP and TFPZ.

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Influence of Mycophenolic Acid and FK–506 on Human Platelet Activation in vitro

Kidney & Blood Pressure Research ◽

10.1159/000025963 ◽

2000 ◽

Vol 23 (2) ◽

pp. 119-124 ◽

Cited By ~ 5

Author(s):

Barbara C. Klein ◽

Dieter Bach ◽

Irene Rehfeld ◽

Elisabeth Kirchhoff ◽

Thomas Hohlfeld ◽

...

Keyword(s):

Platelet Activation ◽

Mycophenolic Acid ◽

Human Platelet ◽

Fk 506

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Reversible Hypothermia-Induced Inhibition of Human Platelet Activation in Whole Blood in Vitro and in Vivo

PsycEXTRA Dataset ◽

10.1037/e464562006-001 ◽

1992 ◽

Author(s):

Alan D. Michelson ◽

Hollace MacGregor ◽

Anita Kestin ◽

Mark R. Barnard ◽

Michael J. Rohrer ◽

...

Keyword(s):

Platelet Activation ◽

Whole Blood ◽

Human Platelet

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Human platelet phospholipase A2 activity is responsive in vitro to pH and Ca2+ variations which parallel those occurring after platelet activation in vivo

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research ◽

10.1016/0167-4889(88)90166-8 ◽

1988 ◽

Vol 971 (1) ◽

pp. 103-111 ◽

Cited By ~ 11

Author(s):

Bruce M. Baron ◽

Lee E. Limbird

Keyword(s):

Phospholipase A2 ◽

Platelet Activation ◽

Human Platelet ◽

Phospholipase A2 Activity

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THE DIHYDROPYRIDINE CALCIUM CHANNEL AGONIST, BAY K 8644, AND THE ANTAGONIST, NIFEDIPINE, INHIBIT U46619-INDUCED HUMAN PLATELET ACTIVATION BY COMPETITIVE BINDING TO THE THROMBOXANE A22/PGH2 RECEPTOR

10.1055/s-0038-1643756 ◽

1987 ◽

Author(s):

G J Johnson ◽

P C Dunlop ◽

M J Rabiet ◽

L A Leis ◽

AH L From

Keyword(s):

Platelet Aggregation ◽

Platelet Activation ◽

Competitive Inhibition ◽

Human Platelet ◽

Ex Vivo ◽

Competitive Binding ◽

Bay K 8644 ◽

Voltage Dependent ◽

Ca2 Channels

The dihydropyridine (DP) Ca2+ channel antagonist, nifedipine (NF), inhibits platelet aggregation .in vitro and ex vivo by an undefined mechanism. Inhibition of Ca2+ influx via Ca2+ channels is a postulated mechanism, but voltage-dependent Ca2+ channels have not been demonstrated in platelets. We previously observed that NF blocked thromboxane A2 (TXA2)-induced platelet aggregation and secretion. In order to further evaluate the mechanism of DP inhibition of platelet activation, we studied the effects of NF and BAY K 8644, (BAY), a DP with opposite (agonist) effects on muscle cells, on human platelet aggregation and secretion induced by the TXA2 mimic, U46619. We also observed the effects of DP on biochemical consequences of platelet activation: cytoplasmic ionized Ca2+ ([Ca2+]i) by fura-2 fluorescence; phosphorylation of 40,000 Dalton protein (40KP) substrate of protein kinase C by SDS-PAGE and [32p] counting; TXA2 formation by RIA of TXB2. 1μM BAY and 10μM NF inhibited the 2nd wave of platelet aggregation and secretion induced by ADP or epinephrine and blocked aggregation and secretion induced by U46619. A Schild plot gave a slope of -1 indicating competitive inhibition of U46619 by BAY (K1[=0.7μM).BAY and NF also blocked U46619-induced phosphorylation of 40KP, rise in [Ca2+]i and TXB2 formation. The (+)-(R) enantiomer of BAY (BAY+) was responsible for BAY inhibition. BAY, BAY(+), and the R enantiomer of another DP, 202-791, all functioned as competitive antagonists of [3H]-U4661 9 binding (K1[ for BAY=2.8 μM-comparable to known receptor antagonists, 13-azaprostanoic acid and BM 13.177; K1 for BAY(+)=0.69μM). Neither BAY nor NF inhibited[3H]-yohimbine binding to α adrenergic receptors.NF, BAY, BAY(+) and BAY(-) in nM concentrations slightly stimulated platelet aggregation,secretion and biochemical events induced by U46619 similar to their effects on muscle. Therefore, DP's do not inhibit platelet activation by blocking voltage-dependent Ca2+ channels. The mechanism of DP inhibition of TXA2-induced platelet activation is stereoselective, competitive binding to the TXA2/PGH2 receptor. DP's may exert similar effects on TXA2-induced vascular smooth muscle contraction.

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Reversible Inhibition of Human Platelet Activation by Hypothermia In Vivo and In Vitro

Thrombosis and Haemostasis ◽

10.1055/s-0038-1642495 ◽

1994 ◽

Vol 71 (05) ◽

pp. 633-640 ◽

Cited By ~ 269

Author(s):

Alan D Michelson ◽

Hollace MacGregor ◽

Marc R Barnard ◽

Anita S Kestin ◽

Michael J Rohrer ◽

...

Keyword(s):

Platelet Aggregation ◽

Platelet Activation ◽

Whole Blood ◽

Peripheral Blood ◽

Bleeding Time ◽

Human Platelet ◽

Platelet Surface ◽

Shed Blood

SummaryA hypothermia-induced hemorrhagic diathesis is associated with cardiopulmonary bypass, major surgery, and multiple trauma, but its pathophysiological basis is not well understood. We examined the hypothesis that hypothermia reversibly inhibits human platelet activation in vitro and in vivo. Platelet activation was studied in normal volunteers by whole blood flow cytometric analysis of modulation of platelet surface GMP-140 and the glycoprotein (GP) Ib-IX complex in: a) shed blood emerging from a standardized in vivo bleeding time wound; b) peripheral blood activated in vitro with either thrombin (in the presence of gly-pro-arg-pro, an inhibitor of fibrin polymerization) or the stable thromboxane (TX) A2 analogue U46619. Platelets in peripheral whole blood were activated at temperatures between 22° C and 37° C. the forearm skin temperature was maintained at temperatures between 22° C and 37° C prior to and during the bleeding time incision. Platelet aggregation was studied in shed blood by flow cytometry and in peripheral blood by aggregometry. Generation of TXB 2 (the stable metabolite of TXA 2) was determined by radioimmunoassay. In vitro, hypothermia inhibited both thrombin- and U46619-induced upregulation of GMP-140, downregulation of the GPIb-IX complex, platelet aggregation, and TXB2 generation. These inhibitory effects of hypothermia were all completely reversed by rewarming the blood to 37° C. In vivo, platelet activation was inhibited by hypothermia as shown by 5 independent assays of shed blood: upregulation of GMP-140, downregulation of the GPIb-IX complex, platelet aggregate formation, TXB 2 ggeneration, and the bleeding time. In summary, by a combination of immunologic, biochemical, and functional assays, we demonstrate that hypothermia inhibits human platelet activation in whole blood in vitro and in vivo. Rewarming hypothermic blood completely reverses the activation defect. These results suggest that maintaining normothermia or rewarming a hypothermic bleeding patient may reduce the need for platelet transfusions.

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Reversible Hypothermia-Induced Inhibition of Human Platelet Activation in Whole Blood in Vitro and in Vivo

10.21236/ada360296 ◽

1992 ◽

Author(s):

A. D. Michelson ◽

H. MacGregor ◽

A. Kestin ◽

M. R. Barnard ◽

M. J. Rohrer

Keyword(s):

Platelet Activation ◽

Whole Blood ◽

Human Platelet

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Human Platelet Aggregation In Response To Multiple Agonists In Plasma Anticoagulated With Heparin

10.1055/s-0038-1652597 ◽

1981 ◽

Author(s):

H A Culliver ◽

N G Ardlie

Keyword(s):

Platelet Aggregation ◽

Platelet Activation ◽

Human Platelet ◽

Platelet Rich Plasma ◽

Adenosine Diphosphate ◽

Human Platelet Aggregation

The lowest concentrations at which epinephrine and vasopressin have been reported to interact positively in causing platelet aggregation in vitro are at least two orders of magnitude greater than the physiological concentrations of these hormones in blood. The aim of this study was to examine the interaction between several agonists of human platelet aggregation. The aggregating agents used were adenosine diphosphate (ADP), epinephrine, norepinephrine, 5-hydroxytryptamine and vasopressin. Platelet-rich plasma (PRP) was prepared from blood anticoagulated with minimal concentrations of heparin in an attempt to more closely reflect the in vivo situation.Aggregation caused by ADP was potentiated by epinephrine at a concentration exceeding the level obtained in circulating blood. When a third agonist (vasopressin) was used in combination with ADP and epinephrine, aggregation was enhanced at concentrations of vasopressin and epinephrine obtained in blood. When used as a fourth agonist norepinephrine and 5-hydroxytryptamine potentiated aggregation at physiological concentrations. The response to multiple agonists was greater in heparinized PRP than citrated PRP. Hirudin decreased the extent of aggregation in heparinized PRP caused by multiple agonists suggesting that thrombin may be involved.Since the concentrations of combined agonists required to induce in vitro platelet aggregation can be obtained in circulating blood these findings may explain why platelet activation occurs in certain pathological states.

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