Binding of Anti-Actin Auto-Antibodies to Platelets

It was recently shown (Gabbiani & Coll., Am. J. Path. 72, 473-488, 1973) that a natural antibody occurring in patients with chronic hepatitis is specifically directed against smooth and striated muscle actin or, in the case of platelets, Thrombosthenin A. Thus it is proposed to change its name from Smooth Muscle Antibody (SMA) to Anti Actin Antibody (AAA). The possibility that AAA would contain a contaminant directed against cell membranes was ruled out because it does not bind (nor its titer in supernatant decrease) to intact platelets, but only to damaged ones (by freeze-thawing). However AAA binds to a platelet-rich clot, and bright fluorescent staining is obtained both for such clots and bone-marrow megacaryocytes. AAA does not induce platelet aggregation in vitro when added to platelet-rich plasma; it does not inhibit or potentiate ADP, Adrenalin or Collagen induced aggregation, and does not inhibit platelet-rich plasma clot retraction.With aggregation controlled in an aggregometer, the following observations were made : AAA adsorption (and consequent fluorescent staining) begins immediately after addition of the trigger, and becomes maximal when a plateau is reached (so-called irreversible aggregation). However, when liminary amounts of ADP are added, so as to obtain reversible aggregation, a large number of disaggregated platelets do bind with AAA, suggesting that actin-related antigens have become available to antibody. This implies that a major change in membrane permeability to large molecules has occurred. The same holds true when ADP-induced aggregation is prevented by EDTA.

Human Platelet Acetylcholinesterase: The Effects of Anticholinesterases on Platelet Function

10.1055/s-0038-1657065 ◽

1979 ◽

Vol 42 (05) ◽

pp. 1615-1619 ◽

Cited By ~ 1

Author(s):

Martin J Smith ◽

Boyd Braem ◽

Kent D Davis

Keyword(s):

Platelet Function ◽

Ache Activity ◽

Room Temperature ◽

Platelet Rich Plasma ◽

Complete Inhibition ◽

Clot Retraction ◽

Plasma Clot ◽

Normal Platelet ◽

Aggregation Factor

SummaryPlatelet acetylcholinesterase (AChE) activity was measured in gel-filtered platelet preparations. Three different anticholinesteratic agents (eserine, neostigmine, and diiso- propylphosphorofluoridate) at final concentrations of 10 μM caused complete inhibition of AChE activity after 30 min incubation at room temperature with either platelet-rich plasma or gel-filtered platelets. Complete inhibition of platelet AChE had no effect on platelet aggregation, factor-3 availability, and plasma clot retraction. We conclude that platelet membrane AChE activity is not required for normal platelet function as measured by these in vitro parameters.

Dipyridamole Inhibits Platelet Aggregation in Whole Blood

10.1055/s-0038-1665327 ◽

1983 ◽

Vol 50 (04) ◽

pp. 852-856 ◽

Cited By ~ 104

Author(s):

P Gresele ◽

C Zoja ◽

H Deckmyn ◽

J Arnout ◽

J Vermylen ◽

...

Keyword(s):

Platelet Aggregation ◽

Whole Blood ◽

Ex Vivo ◽

Platelet Rich Plasma ◽

Significant Negative Correlation ◽

Significant Inhibition ◽

Oral Drug ◽

Human Blood Samples ◽

SummaryDipyridamole possesses antithrombotic properties in the animal and in man but it does not inhibit platelet aggregation in plasma. We evaluated the effect of dipyridamole ex vivo and in vitro on platelet aggregation induced by collagen and adenosine- 5’-diphosphate (ADP) in human whole blood with an impedance aggregometer. Two hundred mg dipyridamole induced a significant inhibition of both ADP- and collagen-induced aggregation in human blood samples taken 2 hr after oral drug intake. Administration of the drug for four days, 400 mg/day, further increased the antiplatelet effect. A significant negative correlation was found between collagen-induced platelet aggregation in whole blood and dipyridamole levels in plasma (p <0.001). A statistically significant inhibition of both collagen (p <0.0025) and ADP-induced (p <0.005) platelet aggregation was also obtained by incubating whole blood in vitro for 2 min at 37° C with dipyridamole (3.9 μM). No such effects were seen in platelet-rich plasma, even after enrichment with leukocytes. Low-dose adenosine enhanced in vitro inhibition in whole blood.Our results demonstrate that dipyridamole impedes platelet aggregation in whole blood by an interaction with red blood cells, probably involving adenosine.

Decreased Platelet Vitamin C in Diabetes Mellitus; Possible Role in Peraggregatoon

10.1055/s-0039-1687640 ◽

1979 ◽

Author(s):

K.E. Sarji ◽

J. Gonzalez ◽

H. Hempling ◽

J.A. Colwell

Keyword(s):

Ascorbic Acid ◽

Arachidonic Acid ◽

Platelet Aggregation ◽

Vitamin C ◽

Platelet Rich Plasma ◽

Solution Ph ◽

Dose Dependent Fashion ◽

Insulin Dependent Diabetic ◽

To determine whether Vitamin C might relate to the increased platelet sensitivity in the diabetic, we have measured levels of platelet Vitamin C and studied the effects of Vitamin C on platelet aggregation. Ascorbic acid levels in washed platelets from diabetics were significantly lower than from normals (4s.2±3 μg/1010 platelets vs. 2s.s±2 μg/1010 platelets, p<.001). The effects of ascorbic acid on platelet aggregation in vitro were studied by adding ascorbic acid in buffered solution (pH 7.35) prior to-aggregating agents. Ascorbic acid in platelet-rich plasma consistently inhibited platelet aggregation with threshold concentrations of ADP, epinephrine, and collagen. With washed platelets, ascorbic acid inhibited arachidonic, acid-induced aggregation. When platelets were incubated at 37°C for 10 minutes with varying concentrations of ascorbic acid, rewashed, and aggregation with arachidonic acid tested, aggregation was inhibited in a linear dose-dependent fashion. Oral ingestion of ascorbic acid (2 gm/day) for seven days by normal non-smoking males produced a marked inhibition of aggregation. In a similar study, platelets from an insulin-dependent diabetic showed no change in aggregation. These results suggest that platelet levels of ascorbic acid may relate to the hyperaggregat ion of platelets from diabetics.

Effects of Ethanol on Pathways of Platelet Aggregation In Vitro

10.1055/s-0038-1647500 ◽

1988 ◽

Vol 59 (03) ◽

pp. 383-387 ◽

Cited By ~ 42

Author(s):

Margaret L Rand ◽

Marian A Packham ◽

Raelene L Kinlough-Rathbone ◽

J Fraser Mustard

Keyword(s):

Arachidonic Acid ◽

Platelet Aggregation ◽

Platelet Rich Plasma ◽

Secondary Phase ◽

Primary Phase ◽

Rabbit Platelet ◽

Membrane Phospholipids ◽

Rabbit Platelets ◽

SummaryEthanol, at physiologically tolerable concentrations, did not affect the primary phase of ADP-induced aggregation of human or rabbit platelets, which is not associated with the secretion of granule contents. Potentiation by epinephrine of the primary phase of ADP-induced aggregation of rabbit platelets was also not inhibited by ethanol. However, ethanol did inhibit the secondary phase of ADP-induced aggregation which occurs with human platelets in citrated platelet-rich plasma and is dependent on the formation of thromboxane A2. Inhibition by ethanol of thromboxane production by stimulated platelets is likely due to inhibition of the mobilization of arachidonic acid from membrane phospholipids, as ethanol had little or no effect on aggregation and secretion induced by arachidonic acid or the thromboxane mimetic U46619. Rabbit platelet aggregation and secretion in response to low concentrations of collagen, thrombin, or PAF were inhibited by ethanol. Inhibition of the effects of thrombin and PAF was also observed with aspirin-treated platelets. Thus, in addition to inhibiting the mobilization of arachidonate for thromboxane formation that occurs with most agonists, ethanol can also inhibit aggregation and secretion through other effects on platelet responses.

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

10.1055/s-0039-1689380 ◽

1975 ◽

Cited By ~ 1

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 ◽

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.

Effects of gas bubbling and other forms of convection on platelets in vitro

Journal of Applied Physiology ◽

10.1152/jappl.1989.67.3.1250 ◽

1989 ◽

Vol 67 (3) ◽

pp. 1250-1255 ◽

Cited By ~ 3

Author(s):

D. M. Pickles ◽

D. Ogston ◽

A. G. MacDonald

Keyword(s):

Shear Stress ◽

Platelet Rich Plasma ◽

Human Platelets ◽

Air Plasma ◽

Physiological Level ◽

Gentle Agitation ◽

Rocking Motion ◽

Experimental Treatments ◽

Citrated platelet-rich human plasma was subjected to one of three experimental treatments at 37 degrees C for 15 min: stirring, bubbling (with stirring), and gentle agitation achieved by a rocking motion. The last two were “equiconvective” as judged by equilibration rates with CO2 and O2 but presumably differed in the shear stress they imposed on the cells. Stirring platelets in normal air or 5% CO2-air caused no significant aggregation. Bubbling air through platelet-rich plasma increased its pH and marked aggregation occurred. Bubbling CO2-air caused the platelet-rich plasma pH to attain its physiological level of 7.4 with less aggregation. In both cases, subsequent ADP-induced aggregation was diminished. Rocking (without stirring) in the presence of CO2-air caused negligible aggregation in platelets and an enhanced response to ADP. Because of the marked difference between the two equiconvective treatments, bubbling and rocking, the main factor in activating the human platelets is suggested to be shear stress (potentiated by high pH), with perhaps a lesser contribution from the air-plasma interface.

PDK1 selectively phosphorylates Thr(308) on Akt and contributes to human platelet functional responses

10.1160/th13-06-0484 ◽

2014 ◽

Vol 111 (03) ◽

pp. 508-517 ◽

Cited By ~ 30

Author(s):

Carol Dangelmaier ◽

Bhanu Kanth Manne ◽

Elizabetta Liverani ◽

Jianguo Jin ◽

Paul Bray ◽

...

Keyword(s):

Protein Kinase ◽

Protein A ◽

Human Platelets ◽

Clot Retraction ◽

Functional Responses ◽

Atp Secretion ◽

Dependent Protein ◽

Summary3-phosphoinositide-dependent protein kinase 1 (PDK1), a member of the protein A,G and C (AGC) family of proteins, is a Ser/Thr protein kinase that can phosphorylate and activate other protein kinases from the AGC family, including Akt at Thr308, all of which play important roles in mediating cellular responses. The functional role of PDK1 or the importance of phosphorylation of Akt on Thr308 for its activity has not been investigated in human platelets. In this study, we tested two pharmacological inhibitors of PDK1, BX795 and BX912, to assess the role of Thr308 phosphorylation on Akt. PAR4-induced phosphorylation of Akt on Thr308 was inhibited by BX795 without affecting phosphorylation of Akt on Ser473. The lack of Thr308 phosphorylation on Akt also led to the inhibition of PAR4-induced phosphorylation of two downstream substrates of Akt, viz. GSK3β and PRAS40. In vitro kinase activity of Akt was completely abolished if Thr308 on Akt was not phosphorylated. BX795 caused inhibition of 2-MeSADP-induced or collagen-induced aggregation, ATP secretion and thromboxane generation. Primary aggregation induced by 2-MeSADP was also inhibited in the presence of BX795. PDK1 inhibition also resulted in reduced clot retraction indicating its role in outside-in signalling. These results demonstrate that PDK1 selectively phosphorylates Thr308 on Akt thereby regulating its activity and plays a positive regulatory role in platelet physiological responses.

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

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y89-155 ◽

1989 ◽

Vol 67 (9) ◽

pp. 989-993 ◽

Cited By ~ 30

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 ◽

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.

The Role of Prostaglandins in the ADP-Induced Aggregation of Rabbit Platelets Shown by the Use of 15-Hydroxyprostaglandin Dehydrogenase

10.1055/s-0038-1646749 ◽

1978 ◽

Vol 39 (03) ◽

pp. 725-732 ◽

Cited By ~ 2

Author(s):

Robert B Wallis

Keyword(s):

Platelet Rich Plasma ◽

Shape Change ◽

Rabbit Platelet ◽

Direct Role ◽

Initial Shape ◽

Rabbit Platelets ◽

Induced Aggregation ◽

Aggregation Of Platelets

SummaryThe initial shape change and subsequent aggregation of platelets in citrated rabbit platelet-rich plasma caused by ADP in vitro was inhibited by 15-hydroxyprostaglandin dehydrogenase. This inhibition was NAD-dependent and was also seen when shape change and aggregation were initiated by sodium arachidonate or by collagen. The aggregation of gel-filtered rabbit platelets by thrombin was not, however, affected by removal of 15-hydroxyprostaglandins.Indomethacin was found to inhibit ADP-induced aggregation but at a concentration (250 μM) much higher than that required to inhibit collagen-induced aggregation. Moreover the platelet release reaction had not taken place 3 min after ADP stimulation. The direct role 15-hydroxyprostaglandin production in ADP-induced aggregation of rabbit platelets is proposed. The involvement of 15-hydroxyprostaglandins in platelet aggregation caused by other inducers is also discussed.

Effect of Lidoflazine (R 7904) on Human Platelet Function in Vitro

10.1055/s-0038-1649059 ◽

1972 ◽

Vol 28 (02) ◽

pp. 228-236 ◽

Cited By ~ 3

Author(s):

F De Clerck

Keyword(s):

Platelet Function ◽

Mode Of Action ◽

Human Platelet ◽

Platelet Rich Plasma ◽

Serotonin Release ◽

Clot Retraction ◽

Plasma Coagulation ◽

Release Reaction ◽

Platelet Release

SummaryThe effect of lidoflazine and of cinnarizine on human platelet function in vitro was compared to that of dipyridamole.Pre-incubation for 30 min at 37° C of platelet rich plasma with lidoflazine or with dipyridamole 5 ×10–4 M resulted in an appreciable inhibition of collagen aggregation in particular and to a lesser extent of ADP aggregation; cinnarizine was marginally active only.Clot retraction was inhibited by lidoflazine and by dipyridamole. Experiments on biphasic ADP aggregation and C14-serotonin release during aggregation show that lidoflazine reduces the platelet release reaction.The possible mode of action of the compound is discussed.Plasma coagulation and PF – 3 availability were not affected.