Optimization of Multiplate® whole blood platelet aggregometry in the Beagle dog and Wistar rat for ex vivo drug toxicity testing

2013 ◽  
Vol 65 (5) ◽  
pp. 637-644 ◽  
Author(s):  
Myriam Defontis ◽  
Serge Côté ◽  
Martina Stirn ◽  
David Ledieu
Keyword(s):  
Whole Blood ◽  
Drug Toxicity ◽  
Ex Vivo ◽  
Toxicity Testing ◽  
Wistar Rat ◽  
Blood Platelet ◽  
Beagle Dog ◽  
Platelet Aggregometry

Multiple electrode aggregometry: A new device to measure platelet aggregation in whole blood

2006 ◽  
Vol 96 (12) ◽  
pp. 781-788 ◽  
Author(s):  
Andreas Calatzis ◽  
Sandra Penz ◽  
Hajna Losonczy ◽  
Wolfgang Siess ◽  
Orsolya Tóth
Keyword(s):  
Platelet Aggregation ◽  
Whole Blood ◽  
Blood Platelet ◽  
Platelet Inhibition ◽  
New Device ◽  
Platelet Aggregometry ◽  
Spontaneous Platelet Aggregation ◽  
Induced Aggregation ◽  
Blood Platelet Aggregation ◽  
Multiple Electrode

SummarySeveral methods are used to analyse platelet function in whole blood. A new device to measure whole blood platelet aggregation has been developed, called multiple electrode platelet aggregometry (MEA). Our aim was to evaluate MEA in comparison with the single platelet counting (SPC) method for the measurement of platelet aggregation and platelet inhibition by aspirin or apyrase in diluted whole blood. Platelet aggregation induced by different concentrations of ADP, collagen and TRAP-6 and platelet inhibition by apyrase or aspirin were determined in citrateor hirudin-anticoagulated blood by MEA and SPC. MEA indicated that spontaneous platelet aggregation was lower, and stimulated platelet aggregation was higher in hirudin- than citrate-anticoagulated blood. In hirudin-anticoagulated, but not citrate-anticoagulated blood, spontaneous platelet aggregation measured by MEA was inhibited by apyrase. For MEA compared with SPC the dose response-curves of agonist-induced platelet aggregation in citrate- and hirudin-blood showed similar EC50 values for TRAP, and higher EC50 values for ADP (non-significant) and collagen (p<0.05). MEA and the SPC method gave similar results concerning platelet-inhibition by apyrase and aspirin. MEA was more sensitive than SPC to the inhibitory effect of aspirin in collagen-induced aggregation. In conclusion, MEA is an easy, reproducible and sensitive method for measuring spontaneous and stimulated platelet aggregation, and evaluating antiplatelet drugs in diluted whole blood. The use of hirudin as an anticoagulant is preferable to the use of citrate. MEA is a promising technique for experimental and clinical applications.

scholarly journals Determination of Clopidogrel Resistance by Whole Blood Platelet Aggregometry and Inhibitors of the P2Y12 Receptor

2006 ◽  
Vol 52 (3) ◽  
pp. 383-388 ◽  
Author(s):  
Boris T Ivandic ◽  
Philipp Schlick ◽  
Peter Staritz ◽  
Kerstin Kurz ◽  
Hugo A Katus ◽  
...  
Keyword(s):  
Whole Blood ◽  
Adenosine Monophosphate ◽  
Blood Platelet ◽  
P2y12 Receptor ◽  
Fisher Exact Test ◽  
Exact Test ◽  
Clopidogrel Resistance ◽  
Impedance Aggregometry ◽  
Platelet Aggregometry ◽  
Increased Risk

Abstract Background: Inhibition of platelet aggregation by clopidogrel may be insufficient in up to 30% of users. These nonresponders carry an increased risk of cardiovascular events. We reported here a simple assay to study clopidogrel responsiveness. Methods: Electrical impedance aggregometry was performed in diluted whole blood in the presence of 5 and 20 μmol/L ADP. Some samples were incubated with 0.1 mmol/L methyl-S-adenosine monophosphate (MeSAMP), a P2Y12 receptor blocker, to maximize inhibition of aggregation before aggregometry. To validate the assay, we analyzed 6-min impedance in 21 healthy probands and 244 patients with coronary artery disease (CAD). Results: At 5 μmol/L ADP, the imprecision of the assay was 11%. Mean (SD) impedance of the healthy cohort was 12.2 (2.2) Ω. The mean − 3 SD was used to define the cutoff for clopidogrel responsiveness: responders and nonresponders exhibited a 6-min impedance ≤5 Ω and &gt;5 Ω, respectively. Samples from nonresponders were incubated with MeSAMP and analyzed again to distinguish pharmacokinetic and pharmacodynamic types of resistance. Sixteen percent of CAD patients were classified as nonresponders (38 and 2 cases of pharmacokinetic and pharmacodynamic resistance, respectively). Female sex was strongly associated with clopidogrel resistance (P = 0.0002, Fisher exact test). A higher clopidogrel loading dose (P = 0.0353, Mann–Whitney U-test) was given to responders (median, 450 mg) than nonresponders (median, 300 mg). Age and cardiovascular diagnosis showed no significant associations. Conclusions: Impedance aggregometry using 5 μmol/L ADP is a useful tool for studying clopidogrel responsiveness. MeSAMP allows characterization of responsiveness “on treatment” and may be useful for optimizing clopidogrel dosing.

COMPARISON OF THE EFFECT OF ASPIRIN (ASA) AND CHOLINE MAGNESIUM TRISALICYLATE (CMT) ON PLATELET AGGREGATION IN WHOLE BLOOD EX-VIVO

1987 ◽  
Author(s):  
B J Z Danesh ◽  
A R Saniabadi ◽  
R I Russell ◽  
G D O Lowe ◽  
C D Forbes
Keyword(s):  
Platelet Aggregation ◽  
Whole Blood ◽  
Therapeutic Potential ◽  
Ex Vivo ◽  
Random Order ◽  
Blood Platelet ◽  
Bleeding Complications ◽  
Double Blind ◽  
Inhibition Of Platelet Aggregation ◽  
Spontaneous Aggregation

Suppression of platelet aggregation by ASA limits the therapeutic use of this drug as an analgesic in patients with bleeding tendencies. CMT is a non-acetylated salicylate derivative with analgesic and anti-inflammatory effect similar to that of ASA. We compared platelet aggregation in human whole blood ex-vivo, three hours after ingestion of ASA and. CMT. Using a whole blood platelet counter, platelet aggregation was quantified by measuring the fall in the number of single platelets at peak aggregation in response to collagen (lμg/ml) arachidonic acid (AA, 0.5 mM) as well as spontaneous aggregation. In double blind and random order, 12 healthy volunteers received a single oral dose of ASA and CMT containing 500 mg equivalent salicylate, on two separate occasions, 10 days apart. Despite a comparable absorption of salicylic acid from the two drugs, ingestion of ASA resulted in a marked inhibition of platelet aggregation induced by collagen, AA and spontaneous aggregation, whereas such effects were not observed after CMT ingestion.We suggest that CMT may have therapeutic potential as an alternative to aspirin when inhibition of platelet aggregation can induce bleeding complications.

scholarly journals An introduction to point-of-care testing in extracorporeal circulation and LVADs

Hematology ◽  
2018 ◽  
Vol 2018 (1) ◽  
pp. 516-521 ◽  
Author(s):  
Rachel Sara Bercovitz
Keyword(s):  
Whole Blood ◽  
Point Of Care ◽  
Blood Platelet ◽  
Ventricular Assist Devices ◽  
Bleeding Complications ◽  
Future Research ◽  
Ventricular Assist ◽  
Assist Devices ◽  
Platelet Aggregometry ◽  
Coagulation Tests

Abstract There is a delicate balance between bleeding and clotting in patients on circuits such as ventricular assist devices or extracorporeal membrane oxygenation. Traditional coagulation tests, prothrombin time, activated partial thromboplastin time, and anti-factor Xa levels, are used to monitor patients on these devices. However, turnaround times and inability to assess global hemostasis, including platelets and fibrinogen have contributed to a recognition that faster, accurate, and more informative coagulation tests are needed. Activated clotting time is used to monitor heparin in patients on circuits and has the advantages of being a near-patient point-of-care test. However, its utility is limited to heparin monitoring. Viscoelastic tests (thromboelastometry and thromboelastography) are global, whole-blood coagulation tests, and whole-blood platelet aggregometry evaluates platelet function. Ideally, these tests can ensure that patients are within the therapeutic range of their antithrombotic medications, identify patients at risk for hemorrhagic or thrombotic complications, and guide management of acute bleeding complications. This ideal is currently hampered by a lack of studies that delineate clear ranges that are clinically relevant. Future research is needed to better understand the optimal use of point-of-care coagulation testing in patients on extracorporeal circuits and ventricular assist devices.

Comparison Of The Effects Of Aspirin In Humans Ex Vivo In Platelet-Rich Plasma And Whole Blood

1981 ◽  
Author(s):  
Barbara Nunn
Keyword(s):  
Whole Blood ◽  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Trisodium Citrate ◽  
Blood Platelet ◽  
Maximal Response ◽  
Blood Samples ◽  
Platelet Concentration

The effect of aspirin on human platelet function is usually assessed using platelet-rich plasma (PRP). Some preliminary results in vitro suggested that the effect of aspirin appears to be greater in PRP than whole blood. To explore this possibility further, a comparison of the effect of aspirin in humans ex vivo has been made taking measurements simultaneously in whole blood and PRP at 2 platelet concentrations. Blood samples (36ml) were drawn from 7 male volunteers after a light breakfast. Each took 300mg soluble aspirin and blood samples were drawn again 2 hours later. Blood was mixed with 0.1 volumes 129nM trisodium citrate. Some (30ml) was then centrifuged to prepare PRP and platelet -poor plasma (PPP) by standard techniques. Platelet concentration of some PRP was adjusted with PPP to equal that of the corresponding blood sample; the rest was adjusted to 350,000 per μl. Aggregation in response to collagen (Horm, Munich) was measured photometrically at 37°. Aggregation in 0.5ml aliquots of whole blood was measured after 4 min stirring with 154mM NaCl (control) or collagen at 37° as the fall in single platelet count determined using an Ultraflo- 100 whole blood platelet counter (Clay Adams). The concentrations of collagen producing a 50% maximal response (EC50) in PRP and blood were determined. Dose-ratios for each volunteer were calculated by dividing the EC50 obtained after aspirin by the corresponding value obtained before aspirin.The effect of aspirin was significantly (p<0.001) less in blood than PRP. Whether or not the results in whole blood more closely reflect the effect of aspirin in vivo remains to be determined.

Platelet Deposition Induced by Severely Damaged Vessel Wall Is Inhibited by a Boroarginine Synthetic Peptide with Antithrombin Activity

1994 ◽  
Vol 71 (04) ◽  
pp. 511-516 ◽  
Author(s):  
J J Badimon ◽  
D Weng ◽  
J H Chesebro ◽  
V Fuster ◽  
L Badimon
Keyword(s):  
Platelet Aggregation ◽  
Whole Blood ◽  
Synthetic Peptide ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Blood Platelet ◽  
Flow Conditions ◽  
Local Flow ◽  
Platelet Deposition ◽  
Damaged Vessel

SummaryThrombin plays a key role in platelet activation and thrombosis. Specific inhibition of thrombin appears to be one of the best approaches to prevent thrombus formation. We have studied the effects of a synthetic a-aminoboronic acid derivative - [Ac, (D) Phe-Pro-Boro-Arg-Hydrocloric acid] - on platelet deposition on severely damaged arterial wall. Platelet deposition was evaluated under well characterized rheological conditions in an original perfusion chamber and detected by autologous mIn-labeled platelets. The study was performed “in vivo” in a porcine model of arterial thrombosis triggered by severely damaged vessel wall at blood flow conditions mimicking mild stenosis (1690 s−1) and patent (212 s−1) vessels. In addition, ex-vivo platelet aggregation activity was evaluated by whole blood impedance aggregometry using collagen, ADP and thrombin as agonists. The synthetic a-aminoboronic peptide was intravenously administered as a bolus followed by continuous infusion. Ex vivo thrombin-induced whole blood platelet aggregation was totally abolished, while ADP- and Collagen-induced whole blood platelet aggregation was not modified. The effects of the synthetic antithrombin on platelet deposition were evaluated in native blood (non-anticoagulated) conditions and in combination with heparin. Under both experimental conditions, the synthetic peptide significantly inhibited platelet deposition at local flow conditions of both high (1690 s−1) and low (212s−1) shear rates. Our results suggest that specific inhibition of locally generated thrombin might be a good strategy to prevent platelet dependent arterial thrombus formation independently of the local flow shear rate of the area at risk.

A Prostacyclin-sparing Effect of Indobufen vs. Aspirin

1996 ◽  
Vol 75 (03) ◽  
pp. 510-514 ◽  
Author(s):  
R De Caterina ◽  
D Giannessi ◽  
W Bernini ◽  
G Lazzerini ◽  
M Lavezzari ◽  
...  
Keyword(s):  
Arachidonic Acid ◽  
Platelet Aggregation ◽  
Whole Blood ◽  
Ex Vivo ◽  
Blood Platelet ◽  
Tolerability Profile ◽  
Double Blind ◽  
Baseline Serum ◽  
Cyclooxygenase Inhibition ◽  
Blood Platelet Aggregation

SummaryIndobufen ((±)-2-[p-(l-oxo-2-insoindolinyl)-phenyI]-butyric acid, indo) is a drug inhibiting platelet function by a reversible block of the arachidonic acid metabolism at the level of cyclooxygenase. Since tolerability profile of such drugs is mostly linked to extra-platelet cyclooxygenase inhibition, we prospectively evaluated the extent of platelet and extra-platelet cyclooxygenase inhibition by in vivo administration of indo in comparison with ASA. We assessed the effects of the two drugs on the ex vivo generation of TXB2 and 6-keto-PGFlΑ in whole blood, as indices of the production of TXA2 and PGI2 (prostacyclin), respectively, either after spontaneous clotting at 37° C for 1 h (Study 1) or after the addition of 2 Μg/ml collagen (Study 2). Generation of 6-keto-PGFlΑ in whole blood is a mixed index of platelet and extra-platelet cyclooxygenase activity, deriving from both platelet and white blood cell arachidonic acid metabolization. Fifteen patients with ischemic heart disease and baseline serum TXB2 levels > 300 ng/ml were allocated to receiving one single administration of either indobufen 200 mg (n = 6) or aspirin 500 mg (n = 9). Whole blood prostanoid generation was assessed at 0,1,2,4,6, 8,12 and 24 h after drug administration (Study 1). Ten healthy male volunteers were allocated to a double-blind, randomized crossover comparison of indo 200 mg b.i.d. vs. ASA 300 mg/d for 7 days (Study 2). Prostanoid generation and whole blood platelet aggregation were performed before and at the end of each study period (Day 0 and Day 7). At each time-point after single dose administration (Study 1), indobufen caused less % inhibition of whole blood 6-keto-PGFlΑ than of TXB2. At 2 h, TXB2 was reduced to a similar extent after ASA (98 ± 4%) and indo (97 ± 6%) (p = N.S.), while inhibition of 6-keto-PGFla was clearly different (> 98% after ASA, 81 ± 2.5% after indo, p < 0.01). After one week of ASA or indo (Study 2) the maximum extent of whole blood platelet aggregation was similarly inhibited (from 17.2 ± 1.4 ohms to 3.6 ± 1.3 ohms with ASA; from 18.3 ± 1.0 ohms to 1.6 ± 0.7 ohms with indo (p ASA vs. indo = N. S.). Despite equal inhibition of whole blood TX production after collagen (from 49.0 ± 4.3 ng/ml to 1.1 ± 0.6 ng/ml with ASA, from 49.8 ± 1.3 ng/ml to 1.4 ± 0.6 ng/ml with indo), again, however, 6-keto-PGFlΑ production was less affected by indo than by ASA (from 409 ± 30 pg/ml to 37 ± 13 pg/ml with ASA, inhibition = 91%; from 396 ± 35 to 318 ± 40 with indo, inhibition = 20%). These differential effects of indo and ASA might lead to a better platelet selectivity, tolerability and benefit/risk profile of indo vs. ASA, which are worthy of further assessment.

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