Assessment of a flow cytometry technique for studying signaling pathways in platelets: Monitoring of VASP phosphorylation in clinical samples

Mallouk, Nora; Varvat, Jérôme; Berger, Anne‐Emmanuelle; Epinat, Magali; Accassat, Sandrine; Garcin, Arnauld; Montmartin, Aurélie; Li, G.; Garnier, P.; Mismetti, Patrick; Lambert, Claude

doi:10.1016/j.plabm.2018.02.002

Cited by 9 publications

(12 citation statements)

References 27 publications

(36 reference statements)

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“…The status of phosphorylation of the platelet vasodilator stimulated phosphoprotein (VASP) was measured using the PLT VASP/P2Y 12 kit (BioCytex, Marseille, France) according to the manufacturer’s instructions as previously described [23]. Phosphorylation of VASP closely correlates with inhibition of the P2Y 12 receptor and platelet aggregation [24].…”

Section: Methodsmentioning

confidence: 99%

Ticagrelor induces paraoxonase-1 (PON1) and better protects hypercholesterolemic mice against atherosclerosis compared to clopidogrel

et al. 2019

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Ticagrelor (TIC), a P2Y purinoceptor 12 (P2Y 12 )-receptor antagonist, has been widely used to treat patients with acute coronary syndrome. Although animal studies suggest that TIC protects against atherosclerosis, it remains unknown whether it does so through its potent platelet inhibition or through other pathways. Here, we placed hypercholesterolemic Ldlr -/- Apobec1 -/- mice on a high-fat diet and treated them with either 25 mg/kg/day of clopidogrel (CLO) or 180 mg/kg/day of TIC for 16 weeks and evaluated the extent of atherosclerosis. Both treatments equally inhibited platelets as determined by ex vivo platelet aggregation assays. The extent of atherosclerosis, however, was significantly less in the TIC group than in the CLO group. Immunohistochemical staining and ELISA showed that TIC treatment was associated with less macrophage infiltration to the atherosclerotic intima and lower serum levels of CCL4, CXCL10, and TNFα, respectively, than CLO treatment. Treatment with TIC, but not CLO, was associated with higher serum activity and tissue level of paraoxonase-1 (PON1), an anti-atherosclerotic molecule, suggesting that TIC might exert greater anti-atherosclerotic activity, compared with CLO, through its unique ability to induce PON1. Although further studies are needed, TIC may prove to be a viable strategy in the prevention and treatment of chronic stable human atherosclerosis.

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Section: Methodsmentioning

confidence: 99%

Ticagrelor induces paraoxonase-1 (PON1) and better protects hypercholesterolemic mice against atherosclerosis compared to clopidogrel

et al. 2019

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“…VASP‐P was recognized as an efficacy indicator of P2Y 12 receptor inhibition. 19 , 20 , 37 Besides, cilostazol was also found to have the ability to phosphorylate VASP through promoting PKA activation and further to inhibit platelet aggregation. 38 It turned out that when cilostazol was combined with P2Y 12 receptor inhibitors, similar phosphorylation of VASP was reached in two groups, indicating that there might be no cumulative effect of PDE III inhibitor and P2Y 12 receptor inhibitors.…”

Section: Discussionmentioning

confidence: 99%

Cilostazol combined with P2Y₁₂ receptor inhibitors: A substitute antiplatelet regimen for aspirin‐intolerant patients undergoing percutaneous coronary stent implantation

Zhao

Zhou

Gao

et al. 2022

Clinical Cardiology

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Background: Cilostazol combined with P2Y 12 receptor inhibitor has been used as a substitute regimen for aspirin-intolerant patients undergoing percutaneous coronary stent implantation on a small scale. Its exact impact on platelet functions and clinical benefits of aspirin-intolerant patients is unknown.Hypothesis: Cilostazol combined with P2Y 12 receptor inhibitors could be used as a substitute antiplatelet regimen for aspirin-intolerant patients undergoing percutaneous coronary stent implantation.Methods: In this multicenter prospective cohort trial, patients undergoing elective percutaneous coronary stent implantation were assigned to the cilostazol group (cilostazol plus P2Y 12 receptor inhibitors), based on aspirin intolerance criteria, or the aspirin group (aspirin plus P2Y 12 receptor inhibitors). Platelet PAC-1, CD62p, and vasodilator-stimulated phosphoprotein phosphorylation (VASP-P) were detected by flow cytometry. The primary endpoints were major adverse cardiovascular and cerebrovascular events (MACCE) including all-cause death, acute myocardial infarction, emerging arrhythmia, nonfatal stroke, and heart failure. The secondary endpoints were the Bleeding Academic Research Consortium (BARC) bleeding events.Results: One hundred and fifty-four aspirin-intolerant percutaneous coronary stent implantation patients and 154 matched aspirin-tolerant patients from a total of 2059 percutaneous coronary stent implantation patients were enrolled. The relative activation level of PAC-1, CD62p, and platelet reaction index reflected by the VASP-P test were similar in the two groups (p > .05). After 12 months of follow-up, the incidence of all-cause death was 1.9% in the cilostazol group and 1.3% in the

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“…To monitor a specific response of blood platelets to ADP, we used the VASP/P2Y 12 flow cytometric kit (BioCytex, Marseille, France) [ 82 ]. Under the study conditions, the phosphorylation of VASP correlates with the inhibition of the P2Y 12 receptor, whereas in the non-phosphorylated state, it is associated with the active form of the P2Y 12 receptor.…”

Section: Methodsmentioning

confidence: 99%

The Molecular Aspects of Disturbed Platelet Activation through ADP/P2Y12 Pathway in Multiple Sclerosis

Dziedzic

Miller

Saluk‐Bijak

et al. 2021

IJMS

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Epidemiological studies confirm a high risk of ischemic events in secondary-progressive multiple sclerosis (SP MS) patients, directly associated with an increased level of pro-thrombotic activity of platelets. Our work aimed to verify potential molecular abnormalities of the platelet P2Y12 receptor expression and functionality as a cause of an increased risk of thromboembolism observed in the course of MS. We have demonstrated an enhanced platelet reactivity in response to adenosine diphosphate (ADP) in SP MS relative to controls. We have also shown an increased mRNA expression for the P2RY12 gene in both platelets and megakaryocytes, as well as enhanced density of these receptors on the platelet surface. We postulate that one of the reasons for the elevated risk of ischemic events observed in MS may be a genetically or phenotypically reinforced expression of the platelet P2Y12 receptor. In order to analyze the effect of the PAR1 (protease activated receptor type 1) signaling pathway on the expression level of P2Y12, we also analyzed the correlation parameters between P2Y12 expression and the markers of platelet activation in MS induced by selective PAR1 agonist (thrombin receptor activating peptide-6, TRAP-6). Identifying the molecular base responsible for the enlarged pro-thrombotic activity of platelets in SP MS could contribute to the implementation of prevention and targeted treatment, reducing the development of cardiovascular disorders in the course of the disease.

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Assessment of a flow cytometry technique for studying signaling pathways in platelets: Monitoring of VASP phosphorylation in clinical samples

Cited by 9 publications

References 27 publications

Ticagrelor induces paraoxonase-1 (PON1) and better protects hypercholesterolemic mice against atherosclerosis compared to clopidogrel

Ticagrelor induces paraoxonase-1 (PON1) and better protects hypercholesterolemic mice against atherosclerosis compared to clopidogrel

Cilostazol combined with P2Y₁₂ receptor inhibitors: A substitute antiplatelet regimen for aspirin‐intolerant patients undergoing percutaneous coronary stent implantation

The Molecular Aspects of Disturbed Platelet Activation through ADP/P2Y12 Pathway in Multiple Sclerosis

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Assessment of a flow cytometry technique for studying signaling pathways in platelets: Monitoring of VASP phosphorylation in clinical samples

Cited by 9 publications

References 27 publications

Ticagrelor induces paraoxonase-1 (PON1) and better protects hypercholesterolemic mice against atherosclerosis compared to clopidogrel

Ticagrelor induces paraoxonase-1 (PON1) and better protects hypercholesterolemic mice against atherosclerosis compared to clopidogrel

Cilostazol combined with P2Y12 receptor inhibitors: A substitute antiplatelet regimen for aspirin‐intolerant patients undergoing percutaneous coronary stent implantation

The Molecular Aspects of Disturbed Platelet Activation through ADP/P2Y12 Pathway in Multiple Sclerosis

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Cilostazol combined with P2Y₁₂ receptor inhibitors: A substitute antiplatelet regimen for aspirin‐intolerant patients undergoing percutaneous coronary stent implantation