Decrease Of Platelet Aggregation In Rats Under Gastric Ulcer-Causing Stress

1981 ◽  
Author(s):  
T Terano ◽  
T Hamazaki ◽  
A Hirai ◽  
Y Tamura ◽  
A Kumagai
Keyword(s):  
Gastric Ulcer ◽  
Platelet Aggregation ◽  
Room Temperature ◽  
Platelet Rich Plasma ◽  
Water Immersion ◽  
Ether Anesthesia ◽  
Normal Rat ◽  
Antiaggregatory Activity ◽  
Stomach Ulceration ◽  
Rat Platelets

Decreased platelet aggregability may play an important role in gastric ulcer continuation. We attempted to identify an antiaggregatory substance involved in stress induced gastric ulceration in rats. METHODS: 1) Male SD rats (180-220g) were stressed by water immersion, one group for 30 min and another for 120 min. Platelet rich plasma (PRP) and platelet poor plasma (PPP) were obtained from stressed rats and normal rats under ether anesthesia. Platelet aggregation was carried out by addition of 1/10 vol of 50μM ADP and was measured using a Sienco aggregometer. 2) Platelet-plasma mixing experiments. PRP from normal rats and from rats stressed for 30 min was centrifuged and the supernatant was discarded. Packed platelets from normal rats were resuspended in PPP from stressed rats and platelets from stressed rats were resuspended in PPP from normal rats. Also, platelets from each rat were resuspended in PPP of the same rat. Aggregation was then observed. 3) Stability tests of the activity in PPP from stressed rats which depressed platelet aggregation. PPP from normal and stressed rats was stored for 4. hr either in ice or at room temperature. Platelets from a normal rat were then mixed with each PPP. 4) 6 keto PGF1α was measured by RIA. RESULTS: 1) The aggregability of rat platelets was decreased to 1/4-1/5 and to 1/10 of the normal value, after 30 min and 120 min stress, respectively. 2) Normal platelets suspended in PPP from stressed rats did not aggregate. Platelets from stressed rats suspended in normal PPP showed significant aggregability. 3) The antiaggregatory activity of PPP from stressed rats was lost significantly after storage at room temperature for 4 hr, while storage in ice did not greatly affect this activity. DISCUSSION: During water immersion stress which causes stomach ulceration of rats there appear platelet antiaggregatory substance(s) in plasma. Like PGI2, the substance observed in this experiment is more stable at 0°C than at room temperature. However, plasma 6 keto PGF1αconcentrations in stressed rats did not increase enough to explain the decreased platelet aggregation. Identification of this antiaggregatory substance is now under way.

Platelet Retention in Glass Bead Columns: Further Evidence for the Importance of ADP

Blood ◽  
1974 ◽  
Vol 44 (3) ◽  
pp. 411-425 ◽  
Author(s):  
V. J. McPherson ◽  
M. B. Zucker ◽  
N. M. Friedberg ◽  
P. L. Rifkin
Keyword(s):  
Platelet Aggregation ◽  
Acetylsalicylic Acid ◽  
Glass Bead ◽  
Room Temperature ◽  
Creatine Phosphokinase ◽  
Platelet Rich Plasma ◽  
Creatine Phosphate ◽  
Inhibitory Effect ◽  
Normal Platelet ◽  
Normal Individuals

Abstract Plasma of normal heparinized blood contained 0.284 µM ± SD 0.097 (ADP + ATP) with an ATP:ADP ratio of 2.5:1. Plasma from thrombocytopenic blood contained only 0.106 µM ± 0.073 (ADP + ATP). Blood with normal platelet retention released 0.234 µM ± 0.187 (ADP + ATP) during passage through a glass bead column, with an ATP:ADP ratio of 1.6:1. Significantly less was released in blood with low retention, i.e., samples from patients with von Willebrand’s disease, thrombasthenia, or thrombocytopenia, and some samples from normal individuals. Thus, nucleotides in the plasma of pre- and postcolumn blood appear to be derived from platelets; their release within glass bead columns is closely associated with normal platelet retention. Since release occurred at room temperature and was not prevented by acetylsalicylic acid or accompanied by measurable release of 14C-serotonin, the classic release reaction may not have been responsible. The low retention in platelet-rich plasma was variably increased by adding 0.5 µM ADP, an increase at least partly due to trapping of preformed aggregates. Retention in undisturbed blood was markedly inhibited by creatine phosphokinase with creatine phosphate (CPK-CP) and moderately inhibited by apyrase I (ATPase:ADPase 0.8:1) at an ADP-removing activity between 1 and 5 U/ml, indicating that ADP is essential for retention. At less than 1 U/ml, both apyrase I and II (ATPase: ADPase 2.8:1) enhanced retention in undisturbed blood, but CPK-CP was still inhibitory. These results suggest that enhancement is due to conversion of released ATP to ADP, as shown to occur in studies of platelet aggregation with ATP and ADP. At less than 1 U/ml, all three enzymes protected against the inhibitory effect of disturbance; this protection was marked with apyrase II, moderate with apyrase I and slight with CPK-CP. These observations provide additional evidence that ADP is responsible for the low retention caused by disturbance of the blood.

scholarly journals The Effect of Cold on Platelets. II. Platelet Function After Short-term Storage at Cold Temperatures

Blood ◽  
1972 ◽  
Vol 40 (5) ◽  
pp. 688-696 ◽  
Author(s):  
Herman E. Kattlove ◽  
Benjamin Alexander ◽  
Frances White
Keyword(s):  
Platelet Aggregation ◽  
Room Temperature ◽  
Platelet Rich Plasma ◽  
Adenosine Diphosphate ◽  
Serotonin Release ◽  
Prolonged Storage ◽  
Cold Temperatures ◽  
Aggregation Of Platelets ◽  
Term Storage

Abstract Citrated platelet-rich plasma (PRP) was kept at cold temperatures or room temperature. After 4 hr or more at these temperatures, the PRPs were warmed 1 hr at 37°C. This prevents the spontaneous aggregation seen in chilled PRP that is stirred immediately after warming. Platelet aggregation in response to connective tissue (CT), epinephrine, and adenosine diphosphate (ADP) was considerably greater in the PRPs originally kept at cold temperatures. In addition, chilling would restore the aggregation of platelets whose function had deteriorated due to prolonged storage at warm temperatures. Neither ADP-induced refractoriness, serotonin uptake, or CT-induced serotonin release was affected by cold. Retention in glass bead columns was greater in platelets that had been chilled than in platelets kept at room temperature or 37°C. Thus, the storage of platelets at cold temperatures leads to changes that improve platelet aggregation but may also increase platelet adhesion, which would account for the decreased in vivo survival of platelets preserved for transfusion at cold temperatures.

EFFECTS OF INHIBITORS ON COLLAGEN INDUCED PLATELET AGGREGATION IN SIX DIFFERENT SPECIES.

1987 ◽  
Author(s):  
J W C M Jansen
Keyword(s):  
Platelet Aggregation ◽  
Guinea Pig ◽  
Species Differences ◽  
Platelet Rich Plasma ◽  
Phosphodiesterase Inhibitors ◽  
Human Platelets ◽  
Cyclooxygenase Inhibitors ◽  
Pharmacological Studies ◽  
Aggregation Of Platelets ◽  
Rat Platelets

One approach to the development of antithrombotics is inhibition of platelet aggregation. The pharmacological approach often used is to test compounds on collagen induced platelet aggregation measured in platelet rich plasma. Therefore we have compared inhibitors with different mechanism of action on aggregation of platelets from six different species commonly used in pharmacological studies. Aggregation was induced with submaximal amounts of collagen (Hormone Chemie).Inhibitors of the cyclooxygenase system, aspirin and indomethacin, were very potent in inhibiting aggregation of platelets from humans guinea pig and dog (IC50 20-60 and 1-3 ¼M resp.). Aggregation of pig and rat platelets was poorly inhibited by both of these compounds (IC5: 700-900 ¼M), whereas platelets from mice showed intermediate sensitivety (IC50 ca.100 ¼M).The combined lipoxygenase/cyclooxygenase inhibitor BW755C, was extremely active on platelets of guinea pig (IC50 1 ¼M) and was poorly active in mice platelets (IC50 300 ¼M). In the other species the inhibitory activity ranged from 20-80 ¼M.The phosphodiesterase inhibitors, papaverine and BL3459 inhibited aggregation in all species (IC50 50-100 and 1-5 ¼M resp.). Dipyridamole inhibited aggregation also in all species but with lower activity (IC50 > 100 ¼M).Conclusion: remarkable species differences are present with respect to inhibition of collagen induced platelet aggregation by the various compounds e.g. rat and porcine platelet aggregation was hardly inhibited by cyclooxygenase inhibitors. The effects of the compounds on human platelets are comparable to the effects on canine plateletes.

scholarly journals In Vitro Spontaneous Platelet Aggregation in Cerebrovascular Disease

1977 ◽  
Author(s):  
J.W.ten Cate
Keyword(s):  
Platelet Aggregation ◽  
Room Temperature ◽  
Platelet Rich Plasma ◽  
Divalent Cations ◽  
Normal Range ◽  
Refractory State ◽  
Spontaneous Platelet Aggregation ◽  
Second Wave ◽  
Aggregation Tendency

Fifty patients from a group of 130 patients with transient ischemic attacks or cerebral infarction were found to demonstrate in vitro spontaneous platelet aggregation (SPA). This phenomenon occurred within 15 minutes when platelet-rich plasma samples (PRP) of these patients were stirred at 37°C in an aggregometer.In addition all patients showing SPA also demonstrated a lowthreshold concentration for the onset of ADP-induced second wave aggregation (ADP f.c. < 0.25 uM; normal range 0.75 + 0.2 uM). Of the remaining 80 patients 25 patients were found to be sensitive tolow concentrations of ADP.SPA remained present in samples of 8 patients studied when stored at room temperature for two hours. SPA was found to be dependant upon the presence of divalent cations and could be prevented by adenosine, phentolamine and aspirin. The following additional findings point towards a possible platelet defect:1. Platelets from 10 patients with SPA when isolated and resuspended in normal plasma still demonstrated SPA while isolated normal platelets in patients did not.2. Platelets demonstrating SPA showed an increased aggregation tendency upon incubation with ADP while normal platelets developed the expected refractory state for ADP.

scholarly journals Relationship Between Aggregation and Prostaglandin (PG)-Biosynthesis in Rat Platelet Rich Plasma (PRP)

1977 ◽  
Author(s):  
H. Bult ◽  
P. C. Bragt ◽  
I. L. Bonta
Keyword(s):  
Arachidonic Acid ◽  
Fatty Acid ◽  
Essential Fatty Acid ◽  
Platelet Rich Plasma ◽  
Thromboxane A2 ◽  
Essential Fatty Acid Deficient ◽  
Normal Rat ◽  
Threshold Doses ◽  
High Doses ◽  
Rat Platelets

The importance of PG biosynthesis in aggregation has been studied by comparing PRP from normal and essential fatty acid deficient (EFAD) rats. The latter show a marked lack of the PG precursor, arachidonic acid (AA).No differences were observed in aggregation with collagen (from 10 μg/ml), ADP (from 0.8 μM) or the PG-endoperoxide PGH2 (1 μg/ml). PGH2 induced only a small, transient aggregation. However, with a collagen dose that produced a 50% response in normal PRP, the aggregation of EFAD PRP was 94% reduced. This correlated with reduced (84–89%) biosynthesis of thromboxane A2 and PGE-like material (PGE), both assessed by bioassay. The PG synthetase of EFAD PRP was unaltered, since PGE and malondialdehyde production were normal when exogenous AA was added. AA, up to 0.33 mM, did not aggregate normal PRP, but high amounts of PGE were formed.These results indicate that internal PG biosynthesis is only essential for aggregation with threshold doses of collagen. At higher collagen doses endogenous PG production does not seem to be important for aggregation. It is unlikely that residual formation of PG-endoperoxides in EFAD PRP, at high doses of collagen, is sufficient for a normal aggregation, since normal rat platelets form PG-endoperoxides from exogenous AA without aggregation.

ATP- and ADP-Induced Rat Platelet Aggregation: Significance of Plasma in ATP-Induced Aggregation

1979 ◽  
Vol 42 (05) ◽  
pp. 1580-1588 ◽  
Author(s):  
Ethan J Haskel ◽  
Kailash C Agarwal ◽  
Robert E Parks
Keyword(s):  
Creatine Kinase ◽  
Platelet Aggregation ◽  
Human Plasma ◽  
Platelet Rich Plasma ◽  
Creatine Phosphate ◽  
Rat Plasma ◽  
Human Platelets ◽  
Induced Aggregation ◽  
Aggregation Of Platelets ◽  
Rat Platelets

SummaryATP caused platelet aggregation in rat platelet-rich plasma (PRP) but in contrast strongly inhibited ADP-induced human platelet aggregation. ADP-induced aggregation of rat platelets suspended in human plasma was strongly inhibited by ATP, whereas human platelets in rat plasma were aggregated by ADP. The ATP analog β,γ-methylene ATP which is not dephosphorylated did not induce aggregation in rat PRP. Adenosine, AMP, 2- chloroadenosine, α,β-methylene ADP and β,γ-methylene ATP each inhibited ATP-induced aggregation of platelets in rat PRP to a similar extent as ADP-induced aggregation. A solution containing creatine kinase and creatine phosphate (which converts ADP to ATP) rapidly reversed both ADP- and ATP-induced aggregation in rat PRP; preincubation with this solution completely inhibited rat platelet aggregation induced by both ADP and ATP. Adenosine-8-14C-triphosphate ([14C]-ATP) conversion to [14C]-ADP was about five-fold faster in rat plasma than in human plasma. Addition of creatine phosphate to rat PRP strongly inhibited ATP-induced aggregation, while creatine or creatine kinase slightly potentiated aggregation by ATP. Creatine phosphate, creatine, or creatine kinase individually had minimal and varying effects on ADP-induced rat platelet aggregation. These results suggest that the observed phenomenon of ATP-induced aggregation in rat PRP is caused by a higher activity of creatine kinase in rat plasma than in human plasma, which converts the added ATP to ADP, a potent aggregator.

Functional Characterization of PM6/13, a β3-specific (GPIIIa/CD61) Monoclonal Antibody that Shows Preferential Inhibition of Fibrinogen Binding over Fibronectin Binding to Activated Human Platelets

1998 ◽  
Vol 79 (01) ◽  
pp. 177-185 ◽  
Author(s):  
Ashia Siddiqua ◽  
Michael Wilkinson ◽  
Vijay Kakkar ◽  
Yatin Patel ◽  
Salman Rahman ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
Functional Characterization ◽  
Human Platelets ◽  
Western Blots ◽  
Activated Platelets ◽  
Reducing Conditions ◽  
Fibronectin Binding

SummaryWe report the characterization of a monoclonal antibody (MAb) PM6/13 which recognises glycoprotein IIIa (GPIIIa) on platelet membranes and in functional studies inhibits platelet aggregation induced by all agonists examined. In platelet-rich plasma, inhibition of aggregation induced by ADP or low concentrations of collagen was accompanied by inhibition of 5-hydroxytryptamine secretion. EC50 values were 10 and 9 [H9262]g/ml antibody against ADP and collagen induced responses respectively. In washed platelets treated with the cyclooxygenase inhibitor, indomethacin, PM6/13 inhibited platelet aggregation induced by thrombin (0.2 U/ml), collagen (10 [H9262]g/ml) and U46619 (3 [H9262]M) with EC50 = 4, 8 and 4 [H9262]g/ml respectively, without affecting [14C]5-hydroxytryptamine secretion or [3H]arachidonate release in appropriately labelled cells. Studies in Fura 2-labelled platelets revealed that elevation of intracellular calcium by ADP, thrombin or U46619 was unaffected by PM6/13 suggesting that the epitope recognised by the antibody did not influence Ca2+ regulation. In agreement with the results from the platelet aggregation studies, PM6/13 was found to potently inhibit binding of 125I-fibrinogen to ADP activated platelets. Binding of this ligand was also inhibited by two other MAbs tested, namely SZ-21 (also to GPIIIa) and PM6/248 (to the GPIIb-IIIa complex). However when tested against binding of 125I-fibronectin to thrombin stimulated platelets, PM6/13 was ineffective in contrast with SZ-21 and PM6/248, that were both potent inhibitors. This suggested that the epitopes recognised by PM6/13 and SZ-21 on GPIIIa were distinct. Studies employing proteolytic dissection of 125I-labelled GPIIIa by trypsin followed by immunoprecipitation with PM6/13 and analysis by SDS-PAGE, revealed the presence of four fragments at 70, 55, 30 and 28 kDa. PM6/13 did not recognize any protein bands on Western blots performed under reducing conditions. However Western blotting analysis with PM6/13 under non-reducing conditions revealed strong detection of the parent GP IIIa molecule, of trypsin treated samples revealed recognition of an 80 kDa fragment at 1 min, faint recognition of a 60 kDa fragment at 60 min and no recognition of any product at 18 h treatment. Under similar conditions, SZ-21 recognized fragments at 80, 75 and 55 kDa with the 55kDa species persisting even after 18 h trypsin treatment. These studies confirm the epitopes recognised by PM6/13 and SZ-21 to be distinct and that PM6/13 represents a useful tool to differentiate the characteristics of fibrinogen and fibronectin binding to the GPIIb-IIIa complex on activated platelets.

Phospholipase C from Clostridium Perfringens Induces Human Platelet Aggregation in Plasma

1988 ◽  
Vol 59 (02) ◽  
pp. 236-239 ◽  
Author(s):  
Giovanna Barzaghi ◽  
Chiara Cerletti ◽  
Giovanni de Gaetano
Keyword(s):  
Platelet Aggregation ◽  
Receptor Antagonist ◽  
Phospholipase C ◽  
Clostridium Perfringens ◽  
Human Platelet ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Platelet Activating Factor ◽  
Inhibitory Effect ◽  
Thromboxane Receptor Antagonist

SummaryWe studied the aggregating effect of different concentrations of phospholipase C (PLC) (extracted from Clostridium perfringens) on human platelet-rich plasma (PRP). PRP was preincubated with PLC for 3 min at 37° C and the platelet aggregation was followed for 10 min. The threshold aggregating concentration (TAG) of PLC was 3-4 U/ml.We also studied the potentiation of PLC with other stimuli on platelet aggregation. Potentiating stimuli, such as arachidonic acid (AA), ADP. Platelet Activating Factor (PAF) and U-46619 (a stable analogue of cyclic endoperoxides) were all used at subthreshold concentrations. We also studied the possible inhibitory effect of aspirin, apyrase, TMQ, a prostaglandin endoper- oxide/thromboxane receptor antagonist and BN-52021, a PAF receptor antagonist. Only aspirin and apyrase were able to reduce aggregation induced by PLC alone and PLC + AA and PLC + ADP respectively. TMQ and BN-52021 were inactive. In ex vivo experiments oral aspirin (500 mg) partially inhibited platelet aggregation induced by PLC alone, PLC + AA and PLC + ADP 2 and 24 h after administration. Aspirin 20 mg for 7 days also reduced aggregation induced by PLC + AA.

Effect of Temperature on ADP-Induced Platelet Aggregation

1973 ◽  
Vol 29 (01) ◽  
pp. 183-189
Author(s):  
C. A Praga ◽  
E. M Pogliani
Keyword(s):  
Platelet Aggregation ◽  
Incubation Period ◽  
Room Temperature ◽  
Important Variable ◽  
Practical Significance ◽  
Effect Of Temperature ◽  
Induce Platelet Aggregation ◽  
Cuvette Holder ◽  
Exact Temperature

SummaryTemperature represents a very important variable in ADP-induced platelet aggregation.When low doses of ADP ( < 1 (μM) are used to induce platelet aggregation, the length of the incubation period of PRP in the cuvette holder of the aggregometer, thermostatted at 37° C, is very critical. Samples of the same PRP previously kept at room temperature, were incubated for increasing periods of time in the cuvette of the aggregometer before adding ADP, and a significant decrease of aggregation, proportional to the length of incubation, was observed. Stirring of the PRP during the incubation period made these changes more evident.To measure the exact temperature of the PRP during incubation in the aggre- gometer, a thermocouple device was used. While the temperature of the cuvette holder was stable at 37° C, the PRP temperature itself increased exponentially, taking about ten minutes from the beginning of the incubation to reach the value of 37° C. The above results have a practical significance in the reproducibility of the platelet aggregation test in vitro and acquire particular value when the effect of inhibitors of ADP induced platelet aggregation is studied.Experiments carried out with three anti-aggregating agents (acetyl salicyclic acid, dipyridamole and metergoline) have shown that the incubation conditions which influence both the effect of the drugs on platelets and the ADP breakdown in plasma must be strictly controlled.

Enhancement of ADP-induced Platelet Aggregation by Adrenaline in Vivo and Its Prevention

1973 ◽  
Vol 29 (02) ◽  
pp. 490-498 ◽  
Author(s):  
Hiroh Yamazaki ◽  
Itsuro Kobayashi ◽  
Tadahiro Sano ◽  
Takio Shimamoto
Keyword(s):  
Platelet Count ◽  
Platelet Aggregation ◽  
Platelet Rich Plasma ◽  
The Other ◽  
Endothelial Surface ◽  
Important Interaction ◽  
Blood Coagulability ◽  
The Difference

SummaryThe authors previously reported a transient decrease in adhesive platelet count and an enhancement of blood coagulability after administration of a small amount of adrenaline (0.1-1 µg per Kg, i. v.) in man and rabbit. In such circumstances, the sensitivity of platelets to aggregation induced by ADP was studied by an optical density method. Five minutes after i. v. injection of 1 µg per Kg of adrenaline in 10 rabbits, intensity of platelet aggregation increased to 115.1 ± 4.9% (mean ± S. E.) by 10∼5 molar, 121.8 ± 7.8% by 3 × 10-6 molar and 129.4 ± 12.8% of the value before the injection by 10”6 molar ADP. The difference was statistically significant (P<0.01-0.05). The above change was not observed in each group of rabbits injected with saline, 1 µg per Kg of 1-noradrenaline or 0.1 and 10 µg per Kg of adrenaline. Also, it was prevented by oral administration of 10 mg per Kg of phenoxybenzamine or propranolol or aspirin or pyridinolcarbamate 3 hours before the challenge. On the other hand, the enhancement of ADP-induced platelet aggregation was not observed in vitro, when 10-5 or 3 × 10-6 molar and 129.4 ± 12.8% of the value before 10∼6 molar ADP was added to citrated platelet rich plasma (CPRP) of rabbit after incubation at 37°C for 30 second with 0.01, 0.1, 1, 10 or 100 µg per ml of adrenaline or noradrenaline. These results suggest an important interaction between endothelial surface and platelets in connection with the enhancement of ADP-induced platelet aggregation by adrenaline in vivo.

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