scholarly journals Epigenetic regulation of PAR4-related platelet activation: mechanistic links between environmental exposure and cardiovascular disease

2018 ◽  
Author(s):  
Laura J. Corbin ◽  
Amy E. Taylor ◽  
Stephen J. White ◽  
Christopher M. Williams ◽  
Kurt Taylor ◽  
...  
Keyword(s):  
Cardiovascular Disease ◽  
Dna Methylation ◽  
Platelet Activation ◽  
Cardiovascular Health ◽  
Platelet Reactivity ◽  
Dna Hypomethylation ◽  
Enhancer Activity ◽  
Exon 2 ◽  
Short Term Exposure

AbstractProtease-activated receptor 4 (PAR4) is a potent thrombin receptor. Epigenetic control of theF2RL3locus (which encodes for PAR4) via DNA methylation is associated with both smoking and cardiovascular disease. We examined the association between DNA hypomethylation atF2RL3and risk of cardiovascular disease, focusing on acute myocardial infarction (AMI) (n=853 cases / 2,352 controls). We usedin vitrocell models to dissect the role of DNA methylation in regulating expression ofF2RL3.We investigated the interplay betweenF2RL3DNA methylation and platelet function in human (n=41). Lastly, we used Mendelian randomization to unify observational and functional work by assessing evidence for causal relationships using data from UK Biobank (n=407,141) and CARDIoGRAMplusC4D (n=184,305). Observationally, one standard deviation (SD) decrease in DNA methylation atF2RL3was associated with a 25% increase in the odds of AMI.In vitro, short-term exposure of cells to cigarette smoke reducedF2RL3DNA methylation and increased gene expression. Transcriptional assays flagged a role for a CEBP recognition sequence in modulating the enhancer activity ofF2RL3exon 2. Lower DNA methylation atF2RL3was associated with increased platelet reactivity in human. The estimated casual odds ratio of ischaemic heart disease was 1.03 (95% CI: 1.00, 1.07) per 1 SD decrease inF2RL3DNA. In conclusion, we show that DNA methylation-dependent platelet activation is part of a complex system of features contributing to cardiovascular health. Tailoring therapeutic intervention to new knowledge ofF2RL3/PAR4 function should be explored to ameliorate the detrimental effects of this risk factor on cardiovascular health.One sentence summaryDNA methylation-dependent platelet activation is a likely causal contributor to cardiovascular health.

scholarly journals Epigenetic Regulation of F2RL3 Associates with Myocardial Infarction and Platelet Function

2022 ◽  
Author(s):  
Laura Jayne Corbin ◽  
Stephen J White ◽  
Amy Taylor ◽  
Christopher Michael Williams ◽  
Kurt Taylor ◽  
...  
Keyword(s):  
Gene Expression ◽  
Myocardial Infarction ◽  
Cardiovascular Disease ◽  
Dna Methylation ◽  
Platelet Reactivity ◽  
Atherosclerotic Cardiovascular Disease ◽  
Dna Hypomethylation ◽  
Exon 2 ◽  
Increased Risk

Background: DNA hypomethylation at the F2RL3 locus has been associated with both smoking and atherosclerotic cardiovascular disease; whether these smoking-related associations form a pathway to disease is unknown. F2RL3 encodes protease-activated receptor 4, a potent thrombin receptor expressed on platelets. Given the role of thrombin in platelet activation and the role of thrombus formation in myocardial infarction, alterations to this biological pathway could be important for ischemic cardiovascular disease. Methods: We conducted multiple independent experiments to assess whether DNA hypomethylation at F2RL3 in response to smoking is associated with risk of myocardial infarction via changes to platelet reactivity. Using cohort data (N=3,205), we explored the relationship between smoking, DNA hypomethylation at F2RL3 and myocardial infarction. We compared platelet reactivity in individuals with low versus high DNA methylation at F2RL3 (N=41). We used an in vitro model to explore the biological response of F2RL3 to cigarette smoke extract (CSE). Finally, a series of reporter constructs were used to investigate how differential methylation could impact F2RL3 gene expression. Results: Observationally, DNA methylation at F2RL3 mediated an estimated 34% of the smoking effect on increased risk of myocardial infarction. An association between methylation group (low/high) and platelet reactivity was observed in response to PAR4 stimulation. In cells, CSE exposure was associated with a 4.9 to 9.3% reduction in DNA methylation at F2RL3 and a corresponding 1.7 (95% CI: 1.2, 2.4, p=0.04) fold increase in F2RL3 mRNA. Results from reporter assays suggest the exon 2 region of F2RL3 may help control gene expression. Conclusions: Smoking-induced epigenetic DNA hypomethylation at F2RL3 appears to increase PAR4 expression with potential downstream consequences for platelet reactivity. Combined evidence here not only identifies F2RL3 DNA methylation as a possible contributory pathway from smoking to cardiovascular disease risk, but from any feature potentially influencing F2RL3 regulation in a similar manner.

scholarly journals The JUUL E-Cigarette Elevates the Risk of Thrombosis and Potentiates Platelet Activation

2020 ◽  
Vol 25 (6) ◽  
pp. 578-586
Author(s):  
Jean E. Montes Ramirez ◽  
Zubair A. Karim ◽  
Ahmed B. Alarabi ◽  
Keziah R. Hernandez ◽  
Ziyad Ben Taleb ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Platelet Function ◽  
Cardiovascular Health ◽  
Electronic Cigarettes ◽  
The United States ◽  
Short Term ◽  
Short Term Exposure ◽  
Clean Air

Background: Smoking is the main preventable cause of death in the United States and worldwide and is associated with serious cardiovascular health consequences, including thrombotic diseases. Recently, electronic cigarettes (e-cigarettes) and, in particular JUUL, have attained wide popularity among smokers, nonsmokers, pregnant females, and even the youth, which is alarming. Interestingly, there is/are no information/studies regarding the effect of JUUL on cardiovascular diseases, specifically in the context of modulation of platelet activation. Thus, it is important to discern the cardiovascular disease health risks associated with JUUL. Methods and Results: We used a passive e-vape vapor inhalation system where C57BL/6J mice (10-12 weeks old) were exposed to JUUL e-cigarette vape. Menthol flavored JUUL pods containing 5% nicotine by weight were used as the e-liquid. Mice were exposed to a total of 70 puffs daily for 2 weeks; 3-second puff duration, and 25-second puff interval. The effects of JUUL relative to clean air were analyzed, on mouse platelet function in vitro (eg, aggregation) and in vivo (eg, FeCl3-induced carotid artery injury thrombosis model). Our results indicate that short-term exposure to JUUL e-cigarette causes hyperactivation of platelets and shortens the thrombus occlusion as well as hemostasis/bleeding times, relative to clean air (medians of 14 vs. 200 seconds, P < .01 and 35 vs. 295 seconds, P < .001, respectively). Conclusion: Our findings document—for the first time—that short-term exposure to the JUUL e-cigarette increases the risk of thrombotic events, in part by modulating platelet function, such as aggregation and secretion, in mice.

The Effect of Heparin vs. Citrate on the Interaction of Platelets with Vascular Graft Materials

1985 ◽  
Vol 54 (04) ◽  
pp. 842-848 ◽  
Author(s):  
Kandice Kottke-Marchant ◽  
James M Anderson ◽  
Albert Rabinovitch ◽  
Richard A Huskey ◽  
Roger Herzig
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Vascular Graft ◽  
Time Course ◽  
Platelet Reactivity ◽  
Polymer Surfaces ◽  
In Vitro Perfusion ◽  
Citrate Anticoagulation ◽  
Platelet Release

SummaryHeparin is known to affect platelet function in vitro, but little is known about the effect of heparin on the interaction of platelets with polymer surfaces in general, and vascular graft materials in particular. For this reason, the effect of heparin vs. citrate anticoagulation on the interaction of platelets with the vascular graft materials expanded polytetrafluoroethylene (ePTFE), Dacron Bionit (DB) and preclotted Dacron Bionit (DB/PC) was studied in a recirculating, in vitro perfusion system. Platelet activation, as shown by a decrease in platelet count, an increase in platelet release and a decrease in platelet aggregation, was observed for all vascular graft materials tested using heparin and was greater for Dacron and preclotted Dacron than for ePTFE. Significant differences between heparin and citrate anticoagulation were seen for platelet release, platelet aggregation and the relative ranking of material platelet-reactivity. However, the trends and time course of platelet activation were similar with both heparin and citrate for the materials tested.

scholarly journals Erythrocytes metabolically enhance collagen-induced platelet responsiveness via increased thromboxane production, adenosine diphosphate release, and recruitment

Blood ◽  
1991 ◽  
Vol 78 (1) ◽  
pp. 154-162 ◽  
Author(s):  
J Valles ◽  
MT Santos ◽  
J Aznar ◽  
AJ Marcus ◽  
V Martinez-Sales ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Therapeutic Potential ◽  
Platelet Reactivity ◽  
Adenosine Diphosphate ◽  
Stimulus Response ◽  
Absolute Requirement ◽  
Thromboxane Production ◽  
And Function ◽  
Aspirin Ingestion

Abstract Erythrocytes promoted platelet reactivity in a plasma medium, as demonstrated in an in vitro system that independently evaluated the biochemistry of platelet activation and recruitment. The prothrombotic erythrocyte effects were metabolically regulated, as evidenced by lack of activity of ATP-depleted or glutaraldehyde-fixed erythrocytes. They occurred in the absence of cell lysis as verified by lactate dehydrogenase assays, and had an absolute requirement for platelet activation. The presence of erythrocytes induced a twofold increase in platelet thromboxane B2 (TXB2) synthesis upon collagen stimulation, indicating that erythrocytes modulated platelet eicosanoid formation. Cell-free releasates from stimulated platelet-erythrocyte suspensions, which exhibited increased recruiting capacity, contained 6.9-fold more ADP and 4.9-fold more ATP than releasates from stimulated platelets alone. Following aspirin ingestion, TXB2 formation was blocked, but erythrocyte promotion of platelet reactivity persisted at those doses of collagen that reinduced platelet activation. Moreover, when platelet mixtures consisted of as little as 10% obtained before aspirin plus 90% obtained post-aspirin ingestion, significant erythrocyte enhancement of platelet reactivity occurred, even at low agonist concentrations. These erythrocyte effects would decrease the therapeutic potential of inhibition of platelet cyclooxygenase by aspirin. The erythrocyte- induced modulation of platelet biochemistry and function emphasizes the importance of cell-cell interactions in stimulus-response coupling.

scholarly journals RGS10 and RGS18 differentially limit platelet activation, promote platelet production, and prolong platelet survival

Blood ◽  
2020 ◽  
Vol 136 (15) ◽  
pp. 1773-1782 ◽  
Author(s):  
Daniel DeHelian ◽  
Shuchi Gupta ◽  
Jie Wu ◽  
Chelsea Thorsheim ◽  
Brian Estevez ◽  
...  
Keyword(s):  
Platelet Count ◽  
G Protein ◽  
Platelet Activation ◽  
Platelet Reactivity ◽  
Rgs Proteins ◽  
Platelet Recovery ◽  
Platelet Production ◽  
Platelet Survival

Abstract G protein–coupled receptors are critical mediators of platelet activation whose signaling can be modulated by members of the regulator of G protein signaling (RGS) family. The 2 most abundant RGS proteins in human and mouse platelets are RGS10 and RGS18. While each has been studied individually, critical questions remain about the overall impact of this mode of regulation in platelets. Here, we report that mice missing both proteins show reduced platelet survival and a 40% decrease in platelet count that can be partially reversed with aspirin and a P2Y12 antagonist. Their platelets have increased basal (TREM)-like transcript-1 expression, a leftward shift in the dose/response for a thrombin receptor–activating peptide, an increased maximum response to adenosine 5′-diphosphate and TxA2, and a greatly exaggerated response to penetrating injuries in vivo. Neither of the individual knockouts displays this constellation of findings. RGS10−/− platelets have an enhanced response to agonists in vitro, but platelet count and survival are normal. RGS18−/− mice have a 15% reduction in platelet count that is not affected by antiplatelet agents, nearly normal responses to platelet agonists, and normal platelet survival. Megakaryocyte number and ploidy are normal in all 3 mouse lines, but platelet recovery from severe acute thrombocytopenia is slower in RGS18−/− and RGS10−/−18−/− mice. Collectively, these results show that RGS10 and RGS18 have complementary roles in platelets. Removing both at the same time discloses the extent to which this regulatory mechanism normally controls platelet reactivity in vivo, modulates the hemostatic response to injury, promotes platelet production, and prolongs platelet survival.

scholarly journals Naturally occurring methylation inhibitor: DNA hypomethylation and hemoglobin synthesis in human K562 cells.

1987 ◽  
Vol 7 (5) ◽  
pp. 1759-1763 ◽  
Author(s):  
S B Lyon ◽  
L Buonocore ◽  
M Miller
Keyword(s):  
Dna Methylation ◽  
Genomic Dna ◽  
Dna Methyltransferase ◽  
Globin Gene ◽  
K562 Cells ◽  
Gene Probe ◽  
Dna Hypomethylation ◽  
Naturally Occurring ◽  
Methylation Inhibitor

A naturally occurring methylation inhibitor isolated from rabbit liver and named methinin inhibits a number of methyltransferases. Methinin is a low-molecular-weight compound (1,400) that has an active amine group. This compound inhibits the DNA methyltransferase of human erythroleukemia cells (K562) in vitro. When the K562 cells were grown in medium containing methinin, fetal hemoglobin was produced. Small but detectable amounts of adult hemoglobin were also produced. Methinin was not toxic to these cells. The overall rate of genomic DNA methylation was reduced by 60% in cells grown in medium containing methinin. Southern blots of genomic DNA from methinin-treated cells and untreated cells hybridized to a 32P-labeled globin gene probe showed that one site in the globin gene region was hypomethylated. Methinin is a naturally occurring compound which inhibits DNA methylation both in vitro and in vivo.

scholarly journals Genome-scale oscillations in DNA methylation during exit from pluripotency

2018 ◽  
Author(s):  
Steffen Rulands ◽  
Heather J Lee ◽  
Stephen J Clark ◽  
Christof Angermueller ◽  
Sébastien A Smallwood ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Single Cell ◽  
Epigenetic Mark ◽  
List Type ◽  
Dna Hypomethylation ◽  
Cell Fate Decisions ◽  
Oscillatory Dynamics ◽  
Genome Scale

SummaryPluripotency is accompanied by the erasure of parental epigenetic memory with naïve pluripotent cells exhibiting global DNA hypomethylation both in vitro and in vivo. Exit from pluripotency and priming for differentiation into somatic lineages is associated with genome-wide de novo DNA methylation. We show that during this phase, coexpression of enzymes required for DNA methylation turnover, DNMT3s and TETs, promotes cell-to-cell variability in this epigenetic mark. Using a combination of single-cell sequencing and quantitative biophysical modelling, we show that this variability is associated with coherent, genome-scale, oscillations in DNA methylation with an amplitude dependent on CpG density. Analysis of parallel single-cell transcriptional and epigenetic profiling provides evidence for oscillatory dynamics both in vitro and in vivo. These observations provide fresh insights into the emergence of epigenetic heterogeneity during early embryo development, indicating that dynamic changes in DNA methylation might influence early cell fate decisions.HighlightsCo-expression of DNMT3s and TETs drive genome-scale oscillations of DNA methylationOscillation amplitude is greatest at a CpG density characteristic of enhancersCell synchronisation reveals oscillation period and link with primary transcriptsMultiomic single-cell profiling provides evidence for oscillatory dynamics in vivo

scholarly journals Rivaroxaban Reduces Arterial Thrombosis by Inhibition of FXa-Driven Platelet Activation via Protease Activated Receptor-1

2020 ◽  
Vol 126 (4) ◽  
pp. 486-500 ◽  
Author(s):  
Tobias Petzold ◽  
Manuela Thienel ◽  
Lisa Dannenberg ◽  
Philipp Mourikis ◽  
Carolin Helten ◽  
...  
Keyword(s):  
Atrial Fibrillation ◽  
Platelet Activation ◽  
Arterial Thrombosis ◽  
Platelet Reactivity ◽  
Thrombus Formation ◽  
Vitamin K Antagonists ◽  
Factor Xa ◽  
Antiplatelet Effect ◽  
Protease Activated Receptor 1

Rationale: A reduced rate of myocardial infarction has been reported in patients with atrial fibrillation treated with FXa (factor Xa) inhibitors including rivaroxaban compared with vitamin K antagonists. At the same time, low-dose rivaroxaban has been shown to reduce mortality and atherothrombotic events in patients with coronary artery disease. Yet, the mechanisms underlying this reduction remain unknown. Objective: In this study, we hypothesized that rivaroxaban’s antithrombotic potential is linked to a hitherto unknown rivaroxaban effect that impacts on platelet reactivity and arterial thrombosis. Methods and Results: In this study, we identified FXa as potent, direct agonist of the PAR-1 (protease-activated receptor 1), leading to platelet activation and thrombus formation, which can be inhibited by rivaroxaban. We found that rivaroxaban reduced arterial thrombus stability in a mouse model of arterial thrombosis using intravital microscopy. For in vitro studies, atrial fibrillation patients on permanent rivaroxaban treatment for stroke prevention, respective controls, and patients with new-onset atrial fibrillation before and after first intake of rivaroxaban (time series analysis) were recruited. Platelet aggregation responses, as well as thrombus formation under arterial flow conditions on collagen and atherosclerotic plaque material, were attenuated by rivaroxaban. We show that rivaroxaban’s antiplatelet effect is plasma dependent but independent of thrombin and rivaroxaban’s anticoagulatory capacity. Conclusions: Here, we identified FXa as potent platelet agonist that acts through PAR-1. Therefore, rivaroxaban exerts an antiplatelet effect that together with its well-known potent anticoagulatory capacity might lead to reduced frequency of atherothrombotic events and improved outcome in patients.

scholarly journals Comparative proteomics reveals unexpected quantitative phosphorylation differences linked to platelet activation state

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
G. J. Schmidt ◽  
C. M. Reumiller ◽  
H. Ercan ◽  
U. Resch ◽  
E. Butt ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Platelet Activation ◽  
Platelet Reactivity ◽  
Fold Increase ◽  
Surface Expression ◽  
Discrimination Power ◽  
Vasp Phosphorylation ◽  
Platelet Proteome ◽  
Quantitative Changes

AbstractThere is a need to assess platelet activation in patients with thrombotic disorders. P-selectin and activated integrin αIIbβ3 are usually quantified by flow cytometry to measure platelet activation. Monitoring changes in vasodilator-stimulated phosphoprotein (VASP) phosphorylation is an established method to determine the platelet-reactivity status. To study disruptions of platelet reactivity more comprehensively, we compared the human non-secretory platelet proteome after in-vitro -activation and –inhibition with their respective untreated controls using unbiased fluorescence two-dimensional differential in-gel electrophoresis. The non-secretory platelet proteome was more severely affected during inhibition than during activation. Strikingly, while VASP reached a 1.3-fold increase in phosphorylation levels in inhibited platelets, other protein kinase A targets showed several-fold stronger inhibition-induced phosphorylation levels, including LIM and SH3 domain protein 1 (6.7-fold), Src kinase-associated phosphoprotein 2 (4.6-fold), and Ras-related protein Rap1b (4.1-fold). Moreover, phosphorylation of integrin-linked protein kinase (ILK) and pleckstrin (PLEK) species was associated with P-selectin surface expression. The discrimination power between activation and inhibition was more pronounced for dephosphorylated ILK (3.79 Cohen’s d effect size) and phosphorylated PLEK (3.77) species than for P-selectin (2.35). These data reveal new insights into the quantitative changes of the platelet reactivity proteome and suggest powerful alternatives to characterise their activation and inactivation potential.

In vivo platelet activation and platelet hyperreactivity in abacavir-treated HIV-infected patients

2013 ◽  
Vol 110 (08) ◽  
pp. 349-357 ◽  
Author(s):  
Barbara Belfiori ◽  
Eleonora Petito ◽  
Giuseppe Guglielmini ◽  
Lisa Malincarne ◽  
AnnaMaria Mezzasoma ◽  
...  
Keyword(s):  
Platelet Activation ◽  
Cardiovascular Events ◽  
Light Transmission ◽  
Ex Vivo ◽  
Platelet Reactivity ◽  
Retrospective Case ◽  
Activation Markers ◽  
Induced Aggregation

SummaryAbacavir (ABC) has been associated with ischaemic cardiovascular events in HIV-infected patients, but the pathogenic mechanisms are unknown. Aim of our study was to assess whether ABC induces in vivo platelet activation and ex vivo platelet hyper-reactivity. In a retrospective, case-control study, in vivo platelet activation markers were measured in 69 HIV-infected patients, before starting therapy and after 6–12 months of either ABC (n=35) or tenofovir (TDF) (n=34), and compared with those from 20 untreated HIV-infected patients. A subgroup of patients was restudied after 28–34 months for ex vivo platelet reactivity. In vivo platelet activation markers were assessed by ELISA or flow cytometry, ex vivo platelet reactivity by light transmission aggregometry (LTA) and PFA-100®. The in vitro effects of the ABC metabolite, carbovir triphosphate, on aggregation and intra-platelet cGMP were also studied. sPLA2, sPsel and sGPV increased significantly 6–12 months after the beginning of ABC, but not of TDF or of no treatment. Ex vivo platelet function studies showed enhanced LTA, shorter PFA-100® C/ADP closure time and enhanced platelet expression of P-sel and CD40L in the ABC group. The intake of ABC blunted the increase of intraplatelet cGMP induced by nitric oxide (NO) and acutely enhanced collagen-induced aggregation. Preincubation of control platelets with carbovir triphosphate in vitro enhanced platelet aggregation and blunted NO-induced cGMP elevation. In conclusion, treatment with ABC enhances in vivo platelet activation and induces platelet hyperreactivity by blunting the inhibitory effects of NO on platelets. These effects may lead to an increase of ischaemic cardiovascular events.

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