Anfibatide, a novel GPIb complex antagonist, inhibits platelet adhesion and thrombus formation in vitro and in vivo in murine models of thrombosis

2014 ◽  
Vol 111 (02) ◽  
pp. 279-289 ◽  
Author(s):  
Yan Hou ◽  
Hui Zhou ◽  
Yiming Wang ◽  
Alexandra Marshall ◽  
Chaofan Liang ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Snake Venom ◽  
Platelet Adhesion ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
High Shear ◽  
Antithrombotic Agent ◽  
Vessel Occlusion ◽  
Induced Aggregation

SummaryPlatelet adhesion and aggregation at the sites of vascular injury are key events for thrombosis and haemostasis. It has been well demonstrated that interaction between glycoprotein (GP) Ib and von Willebrand factor (VWF) initiates platelet adhesion and contributes to platelet aggregation, particularly at high shear. GPIb has long been suggested as a desirable antithrombotic target, but anti-GPIb therapy has never been successfully developed. Here, we evaluated the antithrombotic potential of Anfibatide, a novel snake venom-derived GPIb antagonist. We found Anfibatide inhibited washed murine platelet aggregation induced by ristocetin and recombinant murine VWF. It also blocked botrocetin-induced binding of murine plasma VWF to recombinant human GPIb . Interestingly, Anfibatide did not inhibit botrocetin- induced aggregation of platelet-rich plasma, indicating that its binding site may differ from other snake venom-derived GPIb antagonists. Anfibatide strongly inhibited platelet adhesion, aggregation, and thrombus formation in perfusion chambers at high shear conditions and efficiently dissolved preformed thrombi. Anfibatide also inhibited thrombus growth at low shear conditions, though less than at high shear. Using intravital microscopy, we found that Anfibatide markedly inhibited thrombosis in laser-injured cremaster vessels and prevented vessel occlusion in FeCl3-injured mesenteric vessels. Importantly, Anfibatide further inhibited residual thrombosis in VWF-deficient mice, suggesting that Anfibatide has additional antithrombotic effect beyond its inhibitory role in GPIb-VWF interaction. Anfibatide did not significantly cause platelet activation, prolong tail bleeding time, or cause bleeding diathesis in mice. Thus, consistent with the data from an ongoing clinical trial, the data from this study suggests that Anfibatide is a potent and safe antithrombotic agent.

Abstract 33: Platelet 12-lipoxygenase is a Key Regulator of Platelet Reactivity and Thrombus Formation in vivo

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Reheman Adili ◽  
Katherine Mast ◽  
Theodore R Holman ◽  
Michael Holinstat
Keyword(s):  
Platelet Aggregation ◽  
Ex Vivo ◽  
Platelet Reactivity ◽  
Thrombus Formation ◽  
High Shear ◽  
Vessel Occlusion ◽  
12 Lipoxygenase ◽  
Induced Aggregation

Background: Platelet reactivity is required to maintain hemostasis, however high platelet reactivity leads to thrombus formation, myocardial infarction, and stroke. Platelet 12-lipoxygenase (12-LOX) has been demonstrated by our lab and others to regulate agonist-mediated platelet reactivity suggesting a role for 12-LOX in regulation of in vivo thrombosis. The ability to target 12-LOX in vivo has not been established to date. Therefore, we sought to determine if 12-LOX regulates platelet reactivity and thrombus formation in vivo using the selective 12-LOX inhibitor ML355 to determine whether platelet 12-LOX is an effective target for anti-platelet therapeutics. Methods: ML355 effects on human platelet function was assessed in vitro by platelet aggregometry, ex vivo by perfusion chamber, and in vivo by thrombus formation and vessel occlusion in small and large vessels in 12-LOX -/- , WT mice, and mice treated with ML355 via intravital microscopy using the FeCl 3 and laser injury models. Results: In in vitro platelet aggregation, ML355 dose-dependently inhibited agonist-induced aggregation. In ex vivo flow chamber assays, platelet adhesion and thrombus formation on collagen-coated surfaces at high shear was attenuated in both mouse and human whole blood after incubation with ML355. Further, platelet aggregation and thrombus growth in 12-LOX -/- mice were impaired in both laser and FeCl 3 -induced mesenteric, carotid artery and cremaster arteriole thrombosis models. Thrombi in 12-LOX -/- mice were unstable and frequently formed emboli, which resulted in impaired vessel occlusion or reopening. Additionally, thrombus formation and vessel occlusion was impaired in ML355 treated WT mice. Conclusions: The 12-LOX inhibitor ML355 inhibits platelet aggregation induced by a number of platelet agonists. Ex vivo high shear conditions in both mice and human was attenuated in the presence of ML355. Thrombus formation and vessel occlusion were impaired in mice deficient in 12-LOX. Finally, ML355 attenuates thrombus formation and prevents vessel occlusion in vivo . Our data strongly indicates 12-LOX is an important determinant of platelet reactivity and inhibition of platelet 12-LOX may represent a new target for anti-platelet therapeutics.

Novel 12-LOX Inhibitor ML355 Attenuates Platelet Reactivity and Impairs Thrombus Growth, Stability and Vessel Occlusion In Vivo

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 3442-3442 ◽  
Author(s):  
Reheman Adili ◽  
Theodore R Holman ◽  
Michael Holinstat
Keyword(s):  
Platelet Aggregation ◽  
Platelet Function ◽  
Platelet Adhesion ◽  
Ex Vivo ◽  
Platelet Reactivity ◽  
Thrombus Formation ◽  
Vessel Occlusion ◽  
Growth Stability

Abstract Background: Adequate platelet reactivity is required for platelet adhesion and aggregation at the site of vascular injury to maintain hemostasis. However, excessive platelet reactivity can also lead to the formation of occlusive thrombi, the predominate underlying cause of myocardial infarction and stroke. While current anti-platelet treatments limit platelet function, they often result in an increased risk of bleeding. 12-lipoxygenase (12-LOX), an oxygenase highly expressed in the platelet, has been demonstrated by our lab and others to regulate PAR4 and GPVI-mediated platelet reactivity suggesting a role of 12-LOX in regulation of vivo thrombosis. However, the ability to pharmacologically target 12-LOX in vivo has not been established to date. Aims: To determine how 12-LOX regulates thrombus formation in vivo and whether platelet 12-LOX is an effective target for anti-platelet therapeutics, wild-type (WT) or 12-LOX deficient (12-LOX-/-) mice were treated with or without the 12-LOX inhibitor, ML355, and were assessed for inhibitory effects on platelet activation in vitro, ex-vivo and in vivo. Methods: The effect of the novel 12-LOX inhibitor ML355 on human platelet function was assessed in vitro by platelet aggregometry, ex vivo by perfusion chamber. In vivo thrombus formation and vessel occlusion in small and large vessels were studied in 12-LOX-/-, WT mice and mice treated with ML355 using intravital microscopy using the FeCl3 injury models. Results: Using in vitro platelet aggregation assays, ML355 dose dependently inhibited thrombin, PAR1-AP, and PAR4-AP-induced aggregation in washed human platelets. Interestingly, the negative regulatory effects of ML355 inhibition of 12-LOX can be overcome by high concentration of thrombin. Additionally, ML355 was able to attenuate ADP-induced platelet aggregation both in platelet-rich-plasma and whole blood. In ex vivo flow chamber assays, platelet adhesion and thrombus formation on collagen-coated surfaces at high shear was attenuated in both mouse and human whole blood after incubation with ML355. Further, platelet aggregation and thrombus growth in 12-LOX-/- mice was impaired in FeCl3-induced mesenteric or carotid artery thrombosis models. Thrombi in 12-LOX-/- mice were unstable and frequently form emboli, which resulted in impaired vessel occlusion or reopening. Additionally, thrombus formation and vessel occlusion was impaired in ML355 treated WT mice. Conclusions: The highly selective 12-LOX inhibitor ML355 inhibits platelets aggregation induced by various platelet agonists and ML355 inhibition of platelet function is not agonist specific. Platelet function at high shear in ex vivo conditions in both mice and human was attenuated in the presence of ML355. Thrombus growth, stability, and vessel occlusion was impaired in mice deficient for 12-LOX. Finally, the highly selective 12-LOX inhibitor ML355 attenuates thrombus formation and prevents vessel occlusion in vivo. Our data strongly indicates 12- LOX is an important determinant of platelet reactivity and inhibition of platelet 12-LOX may represent a new target for anti-platelet therapeutics. Disclosures No relevant conflicts of interest to declare.

Apolipoprotein AIV (ApoA-IV) Is a Novel Ligand of Platelet β3 Integrin That Negatively Regulates Platelet Adhesion, Aggregation, and Thrombosis.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 156-156
Author(s):  
Christopher M. Spring ◽  
Wuxun Jin ◽  
Hong Yang ◽  
Adili Reheman ◽  
Guangheng Zhu ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Adhesion ◽  
Conflicts Of Interest ◽  
Thrombus Formation ◽  
Protein S ◽  
Vessel Occlusion ◽  
Platelet Adhesion And Aggregation ◽  
Β3 Integrin

Abstract Abstract 156 Platelet adhesion and aggregation at sites of vascular injury are key events required for haemostasis and thrombosis. It has been documented that von Willebrand factor (VWF) and fibrinogen (Fg) are required for platelet adhesion and aggregation. However, we previously showed that occlusive thrombi still form in mice deficient for both Fg and VWF (Fg/VWF−/−) via a β3 integrin-dependent pathway. Here, we have investigated novel, non-classical ligands of β3 integrin that may regulate platelet adhesion and aggregation. To identify potential ligand(s) of β3 integrin, latex beads were coated with purified human platelet β3 integrin and incubated with human plasma. Protein(s) specifically associated with β3 integrin were electrophoresed and apolipoprotein AIV (ApoA-IV) was identified by mass spectrometry. We found that ApoA-IV binds to the surface of stimulated platelets, but not to quiescent platelets or β3−/− platelets, and ApoA-IV/platelet association was blocked by the addition of a specific anti-β3 integrin monoclonal antibody. It appears that ApoA-IV binds to, but is not internalized by platelet β3 integrins. ApoA-IV-deficient (ApoA-IV−/−) mice exhibited enhanced platelet aggregation induced by ADP, Collagen, and TRAP in plasma (but not PIPES buffer) compared to wild type (WT) littermates. This enhancement was diminished when ApoA-IV−/− plasma was replaced by WT plasma, indicating that the reduction was due to plasma ApoA-IV and not an unrelated platelet effect. When platelets were incubated with FITC-Fg, ApoA-IV was able to reduce platelet/Fg association, indicating that ApoA-IV may act to displace pro-thrombotic β3 integrin ligand(s). In support of this, ApoA-IV reduced the number of adherent platelets on immobilized Fg in perfusion chamber assays and enhanced thrombus formation was observed when ApoA-IV−/− mouse blood was perfused over collagen. We found that addition of recombinant ApoA-IV inhibited platelet aggregation and thrombus formation in vitro, while the control apolipoprotein ApoA-I did not. Using intravital microscopy, we further demonstrated that early platelet deposition was increased, and the time for thrombus formation and vessel occlusion were shorter in ApoA-IV−/− mice, which can be corrected by recombinant ApoA-IV transfusion. Furthermore, recombinant ApoA-IV inhibited WT platelet aggregation, thrombus formation and enhanced thrombus dissolution both in vitro and in vivo. Our data demonstrate for the first time that ApoA-IV is a novel ligand of platelet β3 integrin that negatively regulates thrombosis. These new data are consistent with the reported association between ApoA-IV and reduced cardiovascular diseases, and establish the first link between ApoA-IV and thrombosis. Disclosures: No relevant conflicts of interest to declare.

Pharmacological Profile of DZ-697b, a Novel Anti-Platelet Agent -Selective Inhibitor of vWF- and Collagen-Induced Platelet Aggregation.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1875-1875 ◽  
Author(s):  
Yoshiyasu Ogihara ◽  
Sumie Muramatsu ◽  
Yuki Kaneda ◽  
Takako Iijima ◽  
Tomoko Shibutani ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Guinea Pigs ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Bleeding Risk ◽  
Selective Inhibitor ◽  
Pharmacological Profile ◽  
Induced Aggregation

Abstract Introduction: Bleeding risk accompanied with anti-platelet drugs is an ultimate dilemma in the treatment of thrombosis patient. Under high shear condition of blood flow, vWF- and collagen-induced signaling pathways are likely to trigger the platelet adhesion to the injured endothelium, which leads to the activation of platelets and arterial thrombus formation. Thus, the recent studies suggest that the selective inhibitor of these pathways is a new target of anti-platelet drugs with lower bleeding risk. We report here a pharmacological profile of DZ-697b, which selectively inhibits platelet aggregation evoked by ristocetin and collagen in vitro and ex vivo. Materials and methods: Human volunteers blood was processed platelet rich plasma (PRP) or washed platelets. PRP aggregation was induced by ristocetin and collagen. To reveal the selectivity, effect of DZ-697b on U46619 (TXA2 analogue), ADP, thrombin and TRAP induced aggregation in the washed platelets were examined. In guinea pigs and cynomolgus monkeys, effects of DZ-697b given orally were also examined on ex vivo PRP aggregation induced by collagen. To investigate the underlying mechanisms of DZ-697b, changes in phosphorylation of FcR γ chain, a common signaling pathway of both vWF- and collagen-induced platelet aggregation, were studied. Results: DZ-697b potently inhibited both ristocetin- and collagen-induced human PRP aggregation, the IC50 being 0.74 μM and 0.55 μM, respectively. In contrast, DZ-697b even at 50 μM did not show any influences on U46619, ADP, thrombin and TRAP induced platelet aggregation. DZ-697b did not affect ovine COX-1 and COX-2 activities at up to 300 μM. The bioavailability of this compound was more than 80% in monkeys. Oral administration of DZ-697b at 1–3 mg/kg significantly and persistently inhibited collagen induced PRP aggregation in monkeys and guinea pigs. Application of ristocetin, vWF, and collagen significantly increased the intensity of phosphorylation of FcR γ chain in washed platelets, which were inhibited by DZ-697b. Conclusion: DZ-697b is an orally active compound which selectively inhibits ristocetin- and collagen-induced platelet aggregation and seems to be promising as novel anti-platelet drug.

SELECTIVE ANTAGONISTS OF PAF: INTERFERENCE WITH PLATELET FUNCTION AND EFFECT ON THROMBOGENESIS INDUCED BY SUBENDOTHELIUM.

1987 ◽  
Author(s):  
P Hadvary ◽  
H R Baumgartner
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Blood Platelets ◽  
Rabbit Aorta ◽  
Constant Infusion ◽  
Induced Aggregation

Platelet activating factor (PAF) is a very potent excitatory agonist of blood platelets but the physiological importance of this mediator in platelet thrombus formation is not known. We investigated the effect of two chemically unrelated selective inhibitors of PAF-induced platelet aggregation on thrombogenesis induced by rabbit aorta subendothelium (SE) using an ex vivo perfusion system.Ro 19-3704 is a highly potent inhibitor structurally related to PAF. This compound inhibits PAF-induced aggregation of rabbit platelets in platelet rich plasma in vitro competitively. Against 4 nM PAF, a concentration resulting in submaximal platelet aggre-gregation velocity, the IC50 was 70 nM. Inhibition was highly selective for PAF-induced aggregation, since aggregation induced by collagen (HORM, 5 yg/ml), ADP (1 yM) or thrombin (0.4 U/ml) was not inhibited even at a concentration as high as 10 yM. Bro-tizolam, a triazolobenzodiazepine reported to be a selective inhibitor of PAF-induced platelet activation, had in our system an IC50 of 200 nM. The selective benzodiazepine antagonist Ro 151788 was without effect on inhibition of PAF-induced platelet activation by brotizolam.Ro 19-3704 was given intravenously to rabbits as a bolus of 0.2 mg/kg followed by constant infusion of 0.02 mg/kg/min. This dosage provoked ex vivo a constant right shift ratio of the dose response curve for PAF-induced aggregation (RSR[PAF]) by a factor of 25 to 35. Brotizolam was given orally at a dose of 100 mg/ kg together with 300 mg/kg of Ro 15-1788 (to antagonize the central effects) 90 minutes before starting the perfusion experiment, resulting in a RSR[PAF] of 35 to 135. ADP induced platelet aggregation was not impaired by either compound. SE was exposed to the non-anticoagulated blood withdrawn from the carotid artery for 3 min at 2600 s-1 and for 20 min at 200 s-1 shear rate. Quantitative morphometric evaluation showed that SE coverage by platelets and by fibrin, thrombus area and thrombus height were all unchanged by the PAF antagonists at low and at high shear rates despite a very substantial inhibition of PAF-induced platelet aggregation. Therefore a major role of PAF in SE-induced thrombogenesis seems unlikely.

ALX-0081 Nanobody™, an Engineered Bivalent Anti-Thrombotic Drug Candidate with Improved Efficacy and Safety as Compared to the Marketed Drugs.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 896-896
Author(s):  
Karen Silence ◽  
Heidi Jonckheere ◽  
Peter Casteels ◽  
Jan Roodt ◽  
Muriel Meiring ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Half Life ◽  
Platelet Adhesion ◽  
Thrombus Formation ◽  
Bleeding Complications ◽  
High Shear ◽  
Single Chain ◽  
A1 Domain ◽  
Low Shear

Abstract In patients with plaque rupture, platelets adhere, aggregate and form a thrombus. Current strategies to prevent thrombus formation consist of the use of Aspirin®, Plavix® and integrin αIIbβ3 blockers (e.g. Reopro®) in combination with Heparin®. These drugs are associated with high bleeding risk. Several in vivo experiments have shown that neutralizing the collagen von Willebrand Factor (vWF) platelet glycoprotein (GP)Ib-IX-V axis strongly inhibits arterial thrombosis without bleeding complications, therefore, these targets are of high interest to develop new anti-thrombotic drugs. Nanobodies are antibody-derived therapeutic proteins with the structural and functional properties of naturally occurring single-chain antibodies derived from camelids. ALX-0081 is a bivalent humanized Nanobody targeting the GPIb-IX-V binding site at the A1 domain of vWF. The precursor molecule was isolated from a llama immunized with the recombinant A1 domain of vWF and then humanized and engineered into a bivalent format to maximally benefit from the avid binding to vWF. In vitro, ALX-0081 can completely inhibit platelet adhesion to collagen at nanomolar concentrations. This inhibition is specific for the high shear rates relevant for coronary and carotid arteries whilst platelet adhesion and aggregation under low shear conditions is unaltered. The Nanobody also inhibits platelet adhesion to ultra large vWF (ULvWF) whilst it does not inhibit cleavage of ULvWF by ADAMTS-13. In a modified Folt’s model in baboons ALX-0081 inhibits thrombus formation more efficiently than a combination of Aspirin, Heparin and Plavix. Inhibition of thrombus formation is sustained in the presence of epinephrine and upon a new injury confirming the strong anti-thrombotic effect of ALX-0081. The Nanobody is effective at doses approximately 10–20 times lower than the dose required for Reopro. Ex vivo analysis of plasma samples after ALX-0081 administration in baboons in the ristocetin induced platelet aggregation (RIPA) assay reflects the efficacy seen in the Folt’s model. Therefore, this assay seems to be suited to predict effective ALX-0081 concentrations in vitro. In comparison to Reopro and Plavix, ALX-0081 is associated with less bleeding complications, even at doses exceeding the effective dose by a factor of 10 probably because of its selective inhibition of platelet aggregation under high shear but not under low shear conditions. After treatment with ALX-0081 no effect on other hematological parameters such as PT, aPTT, platelet count, VWF concentration and FVIII levels is seen and no immunogenicity is detected in baboons after repeated administration of ALX-0081. The terminal half-life of ALX-0081 in baboons is 8 hours, indicating that the molecule adopts the half-life of vWF. This high efficacy combined with an improved safety compared to the currently marketed drugs suggests that Ablynx’ drug development candidate ALX-0081 can become a powerful drug to treat acute thrombotic events in indications such as ACS, stroke, and TTP.

Effects of Bupranolol, a New β-Blocker, on Platelet Functions of Rabbit and Human in Vitro

1976 ◽  
Vol 36 (02) ◽  
pp. 376-387 ◽  
Author(s):  
Teruhiko Umetsu ◽  
Kazuko Sanai ◽  
Tadakatsu Kato
Keyword(s):  
Platelet Aggregation ◽  
Platelet Adhesion ◽  
Human Platelets ◽  
Rabbit Platelet ◽  
Rabbit Platelets ◽  
Β Blocker ◽  
Platelet Aggregates ◽  
Induced Aggregation ◽  
Platelet Functions

SummaryThe effects of bupranolol, a new β-blocker, on platelet functions were investigated in vitro in rabbits and humans as compared with propranolol, a well-known β-blocker. At first, the effect of adrenaline on ADP-induced rabbit platelet aggregation was studied because adrenaline alone induces little or no aggregation of rabbit platelets. Enhancement of ADP-induced rabbit platelet aggregation by adrenaline was confirmed, as previously reported by Sinakos and Caen (1967). In addition the degree of the enhancement was proved to be markedly affected by the concentration of ADP and to increase with decreasing concentration of ADP, although the maximum aggregation (percent) was decreased.Bupranolol and propranolol inhibited the (adrenaline-ADP-)induced aggregation of rabbit platelets, bupranolol being approximately 2.4–3.2 times as effective as propranolol. Bupranolol stimulated the disaggregation of platelet aggregates induced by a combination of adrenaline and ADP, but propranolol did not. Platelet adhesion in rabbit was also inhibited by the β-blockers and bupranolol was more active than propranolol. With human platelets, aggregation induced by adrenaline was inhibited by bupranolol about 2.8–3.3 times as effectively as propranolol.From these findings. We would suggest that bupranolol might be useful for prevention or treatment of thrombosis.

A Thiazole Compound with Potential Antithrombotic Activity

1982 ◽  
Vol 47 (02) ◽  
pp. 173-176 ◽  
Author(s):  
E E Nishizawa ◽  
A R Mendoza ◽  
T Honohan ◽  
K A Annis
Keyword(s):  
Platelet Aggregation ◽  
Potent Inhibitor ◽  
Platelet Rich Plasma ◽  
Development Program ◽  
Antithrombotic Agent ◽  
Antithrombotic Activity ◽  
Thiazole Derivative ◽  
Cyclooxygenase Activity ◽  
Drug Development Program

SummaryA thiazole derivative, 4,5-bis(p-methoxyphenyl)-2-(trifluoromethyl)-thiazole was found to be a potent inhibitor of collagen-induced platelet aggregation, in vitro, using platelets from at least six species, including man. It was active in human platelet-rich plasma at a concentration of 1 ng/ml. While its antiplatelet activity was greater than that of flurbiprofen, its cyclooxygenase activity was equivalent to that of flurbiprofen. Also, compared to flurbiprofen, the thiazole had less anti-inflammatory activity in the hind-paw edema test. The thiazole derivative inhibited platelet aggregation following oral administration in five laboratory species. In the guinea pig it was active at 0.5 mg/kg. The LD50 in mice was greater than 1000 mg/kg (i.p.). This compound, which was designed through a systematic drug development program, may have high potential as an antithrombotic agent.

Dipyridamole Inhibits Platelet Aggregation in Whole Blood

1983 ◽  
Vol 50 (04) ◽  
pp. 852-856 ◽  
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 ◽  
Induced Aggregation

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

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 ◽  
Induced Aggregation

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.

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