Aggregation of Rabbit Blood Platelets Produced in Vitro by Saline “Extract” of Tendons

1963 ◽  
Vol 09 (02) ◽  
pp. 248-263 ◽  
Author(s):  
Torstein Hovig
Keyword(s):  
Electron Microscopy ◽  
Blood Coagulation ◽  
Platelet Rich Plasma ◽  
Blood Platelets ◽  
Adenosine Diphosphate ◽  
Blood Platelet ◽  
Saline Extract ◽  
Rabbit Blood ◽  
Platelet Aggregates

SummaryRabbit blood platelet aggregates were produced in vitro by addition of rabbit tendon “extract” to citrated platelet-rich plasma. The aggregating activity was not due to presence of adenosine diphosphate in the “extracts” and was unrelated to blood coagulation. The aggregating principle was destroyed by heating of the “extract” to 60° C for 15 minutes or by incubation with collagenase for 1 hour at 37° C. The aggregating effect was associated with particles which were sedi- mented by centrifugation for 30 minutes at 10,000 G. By means of electron microscopy the particles were identified as cross-striated collagen fibrils.

Release of a Platelet-Aggregating Substance (Adenosine Diphosphate) from Rabbit Blood Platelets Induced by Saline “Extract” of Tendons

1963 ◽  
Vol 09 (02) ◽  
pp. 264-278 ◽  
Author(s):  
Torstein Hovig
Keyword(s):  
Platelet Aggregation ◽  
Blood Platelets ◽  
Adenosine Diphosphate ◽  
Blood Platelet ◽  
Saline Extract ◽  
Rabbit Blood ◽  
Blood Platelet Aggregation

Summary1. Blood platelet aggregation was induced by addition of saline “extract” of tendons to citrated PRP (aggregation time 50—60 seconds). The aggregates were sedimented by centrifugation, and aggregating activity was demonstrated in the supernatant (termed supernat. A) with an aggregation time of about 15 seconds.2. It was demonstrated that the activity of the supernatant was due to ADP released from the platelets.3. The conclusion is drawn that the aggregation reaction initiated by the tendon “extract” can be divided into two steps: 1. release of ADP from the platelets, 2. aggregation caused by released ADP.4. The observations are discussed in relation to the formation of platelet plugs in vivo.

Clumping of Blood Platelets by Heat Aggregated Gamma-Globulin

1969 ◽  
Vol 21 (01) ◽  
pp. 065-075 ◽  
Author(s):  
R. B Davis ◽  
G. C Holtz
Keyword(s):  
Platelet Aggregation ◽  
Arterial Thrombosis ◽  
Gamma Globulin ◽  
Platelet Rich Plasma ◽  
Blood Platelets ◽  
Electron Microscopic ◽  
Rabbit Blood ◽  
Platelet Aggregates

SummaryThe influence of heat-aggregated γ-globulin on the in vitro clumping of rabbit blood platelets was investigated in detail. Clumping was found to be dependent on the amount of γ-globulin added to the system. Clumping began within 60 sec, and was influenced by the amount of heparin in the system. Incubation of platelet-rich plasma with adenosine prior to the addition of heat-aggregated γ-globulin reduced the extent of platelet aggregation but did so only after aggregation was well under way, 150 sec after addition of γ-globulin. The extent of platelet clumping by aggregated γ-globulin was significantly increased by non-aggregating quantities of epinephrine, nor-epinephrine, and 5-hydroxytryptamine in the system; however, bradykinin and histamine were without effect. Electron microscopic observations of platelet aggregates showed that variable destructive changes were present in platelet aggregates, with loss of organelles and platelet hyaloplasm. Marked pseudopod formation was observed in platelet aggregates formed by epinephrine and aggregated γ -globulin. The significance of the findings in relation to the problem of arterial thrombosis is discussed.

The Ultrastructure of Rabbit Blood Platelet Aggregates

1962 ◽  
Vol 08 (03) ◽  
pp. 455-471 ◽  
Author(s):  
Torstein Hovig
Keyword(s):  
Structural Changes ◽  
Blood Platelet ◽  
Saline Extract ◽  
Rabbit Blood ◽  
Different Types ◽  
Dense Substance ◽  
Platelet Aggregates ◽  
Free Spaces

SummaryDifferent types of rabbit blood platelet aggregates formed in vitro were studied by means of electron microscopy.1. Aggregates were produced by addition of small amounts of thrombin to citrate-PRP. The platelets, rich in pseudopods, were lying close together with no detectable fibrin between them. Fibrin was observed only in the periphery of the aggregate. Varying structural changes, with loss of granules and mitochondria, occured in the platelets. The appearance of these aggregates was similar to that of the haemostatic platelet plug formed in vivo. In clots of PRP induced by greater concentrations of thrombin, fibrin was readily detectable as bundles of fibres. Platelets with complete loss of internal structure were scattered in the fibrin framework.2. The platelets were usually loosely packed in thrombin-induced aggregates of washed platelets, with no visible fibrin between them. Considerable structural changes of the platelets were observed. Rods or filaments of electron-dense substance were observed inside the platelets.3. The platelets were lying close together in ADP-induced aggregates, but several free spaces could be observed between them. Small and inconsistent structural changes of the platelets appeared.4. Aggregates induced by saline extract of tendons demonstrated marked structural changes of the platelets, with loss of granules and mitochondria. Platelet membranes were mostly preserved. Several free spaces were observed between the platelets. No electron-dense substance indicative of fibrin was detected.The appearance of the different types of aggregates is compared and discussed in relation to the ultrastructure of the haemostatic platelet plug.

The Effect of Calcium and Magnesium on Rabbit Blood Platelet Aggregation in Vitro

1964 ◽  
Vol 12 (01) ◽  
pp. 179-200 ◽  
Author(s):  
Torstein Hovig
Keyword(s):  
Platelet Aggregation ◽  
Cation Exchange ◽  
Calcium Concentration ◽  
Blood Platelets ◽  
Blood Platelet ◽  
Rabbit Blood ◽  
Calcium And Magnesium ◽  
Induced Aggregation ◽  
Blood Platelet Aggregation

SummaryThe effect of calcium and magnesium on the aggregation of rabbit blood platelets in vitro was studied, with the following results:1. Platelet aggregation induced by ADP or collagen could be prevented by EGTA or EDTA. The aggregating effect was restored by recalcification. The effect was also restored by addition of magnesium in EDTA-PRP, but not in EGTA-PRP unless a surplus of calcium was present.2. Calcium remained in concentrations of the order of 0.15–0.25 mM after dialysis or cation exchange of plasma. Aggregation of washed platelets resuspended in such plasma could not be produced with ADP or collagen, unless the calcium concentration was increased or that magnesium was added.3. The adhesiveness of blood platelets to collagen was reduced in EGTA-PRP and EDTA-PRP. Release of ADP from platelets influenced by collagen could not be demonstrated either in EGTA-PRP (presence of magnesium) or in EDTA-PRP.4. It is concluded that calcium is a necessary factor both for the reaction leading to release of ADP and for the the aggregation produced by ADP.5. Thrombin induced aggregation of washed platelets suspended in tris-buffered saline in the presence of calcium. No effect of magnesium could be observed unless small quantities of calcium were present.

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.

Effects of Lead Acetate on Platelet Aggregation, Ultrastructure and Serotonin Release

1971 ◽  
Vol 26 (03) ◽  
pp. 455-466 ◽  
Author(s):  
R. B Davis ◽  
G. C Holtz
Keyword(s):  
Platelet Aggregation ◽  
Platelet Function ◽  
Lead Acetate ◽  
Platelet Rich Plasma ◽  
Adenosine Diphosphate ◽  
Blood Platelet ◽  
Human Platelets ◽  
Serotonin Release ◽  
Aggregation Of Platelets

SummaryThe effects of lead on blood platelet function and ultrastructure have been investigated. Lead acetate was injected intravenously in 27 rats and was added to rat and human platelet rich plasma in vitro. In vitro studies showed that concentrations of 2.5 × 10-3 M lead acetate reduced or blocked aggregation of rat and human platelets by adenosine diphosphate, collagen, and thrombin. Radioactive serotonin release from human platelets was inhibited by 10-4 M lead acetate. One hour after the injection of lead, platelet aggregation by thrombin was reduced, but platelet aggregation by adenosine diphosphate and collagen showed little change. Three days after lead, aggregation of platelets by collagen and thrombin was blocked and aggregation by adenosine diphosphate reduced. Thrombocytopenia was present 4 days after intravenous lead acetate. Electron micrographs of platelets showed that the mean number of mitochondria per platelet was increased, whereas alpha granules were reduced. Dense bodies were not significantly changed. Lead acetate affects platelet function in concentrations reported in human bone marrow in lead poisoning, and may relate to the binding of free sulfhydryl groups by lead.

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

Abstract 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.

Storage of Human Blood Platelets

1974 ◽  
Vol 32 (02/03) ◽  
pp. 405-416 ◽  
Author(s):  
M. R Hardeman ◽  
Carina J L. Heynens
Keyword(s):  
Storage Temperature ◽  
Platelet Rich Plasma ◽  
Blood Platelets ◽  
Storage Period ◽  
Serotonin Uptake ◽  
Hypotonic Shock ◽  
Glycolytic Intermediates

SummaryStorage experiments were performed at 4°, 25° and 37° C with platelet-rich plasma under sterile conditions. In some experiments also the effect of storing platelets at 4° C in whole blood was investigated.Before, during and after three days of storage, the platelets were tested at 37° C for their serotonin uptake and response to hypotonic shock. In addition some glycolytic intermediates were determined.A fair correlation was noticed between the serotonin uptake and hypotonic shock experiments. Both parameters were best maintained at 25° C. Also platelet counting, performed after the storage period, indicated 25° C as the best storage temperature. Determination of glycolytic intermediates did not justify any conclusion regarding the optimal storage temperature. Of the various anticoagulants studied, ACD and heparin gave the best results as to the serotonin uptake and hypotonic shock response, either with fresh or stored platelets. The use of EDTA resulted in the lowest activity, especially after storage.The results of these storage experiments in vitro, correspond well with those in vivo reported in the literature.

Haemostatic Platelet Plug Formation in the Isolated Rabbit Mesenteric Preparation - An Analysis of Red Blood Cell Participation

1980 ◽  
Vol 44 (01) ◽  
pp. 006-008 ◽  
Author(s):  
D Bergqvist ◽  
K-E Arfors
Keyword(s):  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Adenosine Diphosphate ◽  
In Vitro Test ◽  
Pharmacological Agents ◽  
Vitro Test ◽  
Releasing Effect ◽  
Plug Formation

SummaryIn a model using an isolated rabbit mesenteric preparation microvessels were transected and the time until haemostatic plugs formed was registered. Perfusion of platelet rich plasma gave no haemostasis whereas whole blood did. Addition of chlorpromazine or adenosine to the whole blood significantly prolonged the time for haemostasis, and addition of ADP to the platelet rich plasma significantly shortened it. It is concluded that red cells are necessary for a normal haemostasis in this model, probably by a combination of a haemodynamic and ADP releasing effect.The fundamental role of platelets in haemostatic plug formation is unquestionable but there are still problems concerning the stimulus for this process to start. Three platelet aggregating substances have been discussed – thrombin, adenosine diphosphate (ADP) and collagen. Evidence speaking in favour of thrombin is, however, very minimal, and the discussion has to be focused on collagen and ADP. In an in vitro system using polyethylene tubings we have shown that "haemostasis" can be obtained without the presence of collagen but against these results can be argued that it is only another in vitro test for platelet aggregation (1).To be able to induce haemostasis in this model, however, the presence of red blood cells is necessary. To further study this problem we have developed a model where haemostatic plug formation can be studied in the isolated rabbit mesentery and we have briefly reported on this (2).Thus, it is possible to perfuse the vessels with whole blood as well as with platelet rich plasma (PRP) and different pharmacological agents of importance.

Platelet Aggregation Induced in Vitro by Therapeutic Ultrasound

1977 ◽  
Vol 38 (03) ◽  
pp. 0640-0651 ◽  
Author(s):  
B. V Chater ◽  
A. R Williams
Keyword(s):  
Platelet Aggregation ◽  
Direct Action ◽  
Adenosine Diphosphate ◽  
Ultrasonic Cavitation ◽  
Related Phenomenon ◽  
Release Reaction ◽  
Physical Conditions ◽  
Platelet Aggregates ◽  
Active Substances

SummaryPlatelets were found to aggregate spontaneously when exposed to ultrasound generated by a commercial therapeutic device. At a given frequency, aggregation was found to be a dose-related phenomenon, increasing intensities of ultrasound inducing more extensive and more rapid aggregation. At any single intensity, the extent aggregation was increased as the frequency of the applied ultrasound was decreased (from 3.0 to 0.75 MHz).Ultrasound-induced platelet aggregation was found to be related to overall platelet sensitivity to adenosine diphosphate. More sensitive platelets were found to aggregate spontaneously at lower intensities of sound, and also the maximum extent of aggregation was found to be greater. Examination of ultrasound-induced platelet aggregates by electron microscopy demonstrated that the platelets had undergone the release reaction.The observation that haemoglobin was released from erythrocytes in whole blood irradiated under identical physical conditions suggests that the platelets are being distrupted by ultrasonic cavitation (violent gas/bubble oscillation).It is postulated that overall platelet aggregation is the result of two distinct effects. Firstly, the direct action of ultrasonic cavitation disrupts a small proportion of the platelet population, resulting in the liberation of active substances. These substances produce aggregation, both directly and indirectly by inducing the physiological release reaction in adjacent undamaged platelets.

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