scholarly journals Human Apolipoprotein a-I Decreases Platelet Activation Responses and Protects Mice from Arterial Thrombosis

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 4159-4159 ◽  
Author(s):  
Brian R. Branchford ◽  
Christopher Ramos ◽  
Wilbert Jones ◽  
Christine Brzezinski ◽  
Luke Law ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Platelet Function ◽  
Arterial Thrombosis ◽  
Aggregate Formation ◽  
Wild Type ◽  
Platelet Aggregate ◽  
Microfluidic Analysis ◽  
Induced Aggregation ◽  
Platelet Function Assays

Abstract Introduction: High-density lipoprotein (HDL) protects against thromboembolic coronary disease via reverse cholesterol transport mediated by HDL's primary lipoprotein subunit, apolipoprotein A-I (apoA-I), binding to scavenger receptor BI (SR-BI). Absence of SR-BI increases platelet aggregation and venous/arterial thrombosis in mice, consistent with an antithrombotic role of this signaling axis. To date, the effects of the isolated apoA-I subunit on platelet activation remain unknown. We hypothesize that the antithrombotic effect is mediated by apoA-I signaling through platelet-specific SR-BI. Methods: Platelet function assays were performed on samples from healthy human volunteers (n=3). ApoA-I levels were determined using an ELISA kit. Microfluidic analysis of platelet aggregate formation on collagen under physiologic flow conditions (650 sec-1) was carried out in whole blood samples. Light transmission platelet aggregation in response to collagen and ristocetin was conducted using both platelet rich plasma (PRP) and washed platelets (WP). Venous and arterial thromboses were induced in wild type C57Bl/6 mice in a collagen (0.3 mg/kg)/epinephrine (0.03 mg/kg)-induced pulmonary embolism model and a 6% ferric chloride (FeCl3)-induced carotid artery thrombosis model, respectively. Recombinant human apoA-I was used with a final concentration of 300 µg/mL. Results/Discussion: ApoA-I plasma levels in the subjects studied fell within previously published ranges. Microfluidic analysis of platelet aggregate formation on collagen under shear stress did not demonstrate significant difference in total surface area coverage (n=3 in quadruplicate), but apoA1-treated samples demonstrated a significantly greater proportion of small aggregates (3-10 platelets/aggregate, 40.2% vs 31.8%, p<0.0001), compared to controls which had a significantly greater proportion of large aggregates (51-100 plts/agg, 6.3 vs 2.3%, p<0.001 and >100 plts/agg, 2.3% vs 0.7%, p<0.05). Platelet aggregation studies (Fig. 1) revealed significantly decreased collagen-induced aggregation in platelets treated with apoA-I compared to vehicle-treated controls in both PRP and WP samples, suggesting direct action of apoA-I on platelets. The difference was less pronounced in ristocetin-induced aggregation, though still significant, consistent with decreased activation of VWF, which has been recently shown to bind apoA-I. Apoa-I-treated PRP samples also had significantly decreased dense granule (ATP) release compared to non-treated samples. Pre-treatment with native HDL had no significant effect, consistent with studies demonstrating that only oxidized HDL inhibits aggregation. Recombinant human SR-BI (rhSR-BI) alone had little effect in PRP but inhibited collagen-induced aggregation in WP. Addition of rhSR-BI and apoA-I together inhibited aggregation in both PRP and WP to a greater degree than either component alone. Rabbit anti-human SR-BI alone had little effect in PRP but inhibited collagen-induced aggregation in WP. FeCl3 applicationinduced initial arterial occlusion within 8.8 +/-0.8 minutes in apoA-I-treated mice treated (n=3) compared to 6.1 +/- 0.3 minutes in vehicle-treated controls (n=3, p<0.01, t-test). Systemic thrombosis resulting in pulmonary embolism was induced by injection of collagen/epinephrine. Median time of survival after injection in mice pre-treated with apoA-I (n=5) was 30 +/- 0 minutes, compared to 3.3 +/- 0.4 minutes in vehicle-treated controls (n=5, p<0.001, t-test). Experiments were stopped after 30 minutes, at which time all apoA-1-treated mice were still alive, compared to zero controls. Conclusion: Though the mechanism is not yet completely understood, the data show an inhibitory effect of isolated human apoA-I on human platelet activation and murine arterial/venous thrombosis, likely through platelet inhibition. The increased inhibition seen with rhSR-BI and apoA-I together is consistent with previous studies demonstrating SR-BI's regulatory role in platelet function. Ongoing studies include increasing sample size for human platelet function assays and murine venous/arterial thrombosis models in wild type mice with and without infusion of apoA-I compared to SR-BI-/- and apoA-I-/- mice with and without infusion of apoA-I. Figure 1: Maximum aggregation values (mean +/- SEM) following addition of collagen or ristocetin Figure 1:. Maximum aggregation values (mean +/- SEM) following addition of collagen or ristocetin Disclosures No relevant conflicts of interest to declare.

Inhibition of PAR4 Signaling in Ethanol-Attenuation of Platelet Function.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3892-3892
Author(s):  
Shogo Kasuda ◽  
Yoshihiko Sakurai ◽  
Midori Shima ◽  
Masahiro Takeyama ◽  
Katsuhiko Hatake ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Phospholipase A2 ◽  
Platelet Activation ◽  
Platelet Function ◽  
Phase Curve ◽  
Peak Time ◽  
Dependent Manner ◽  
Low Concentrations ◽  
High Concentrations ◽  
Induced Aggregation

Abstract Background: Moderate consumption of alcohol beverages reduces the morbidity from coronary heart disease. Previous studies describing of inhibitory activity of ethanol (EtOH) on platelet function have substantiated this observation. However, the effects of EtOH on thrombin-related platelet activation remains to be fully elucidated, though platelet activation by thrombin is essential for normal hemostasis as well as relevant to pathophysiological conditions of thrombosis. Objectives: The aim of this study is to elucidate the effect of EtOH on α-thrombin-related platelet function by measuring platelet aggregation and intracellular calcium ([Ca2+]i). Materials and Methods: A dual-wavelength spectrofluorometer was used for measurement. α-thrombin, PAR1-activating peptide (AP) (10 μM) or PAR4-AP (25 μM) was added to fura2-AM loaded washed platelet preincubated with or without EtOH (40, 80, 160 and 320 mM). Results and Interpretations: First, the effects of EtOH on 0.5 nM of thrombin-induced platelet activation was assessed. The concentration 0.5 nM used is conceived to activate platelets only via PAR-1. EtOH did not affect platelet aggregation. EtOH inhibited rise of [Ca2+]i dose-dependently. [Ca2+]i peak time at which maximal rise of [Ca2+]i delayed in a dose-dependent manner. Secondly, 10 nM of thrombin was used as an agonist. Stimulation by high concentrations of thrombin (〉 5nM) results in cleavage of both PAR1 and PAR4. The changes in [Ca2+]i showed double-phase curve composed of transient spike and prolonged peak in the absence of EtOH. Although EtOH inhibited neither platelet aggregation nor the first phase of [Ca2+]i increasing, it reduced the second prolonged elevation of [Ca2+]i dose-dependently. To elucidate the inhibiting mechanism of EtOH more precisely, the effects of EtOH on PAR1-AP-induced platelet function were examined. Rise of [Ca2+]i gave a spike form and was almost unchanged even in the presence of high concentrations of EtOH, whereas platelet aggregation was reduced and dissociated in the presence of EtOH. Lastly, the effects of EtOH on PAR4-AP-induced platelet function was examined. Aggregation of PRP was quenched by high concentrations of EtOH but dissociation was not observed contrary to that observed in PAR1-AP-induced aggregation. Further, EtOH inhibited [Ca2+]i rise and delayed [Ca2+]i peak time dose-dependently. Our results provided a possible mechanism by which EtOH inhibits platelet activation. Reduction of the prolonged elevation of [Ca2+]i by high concentrations of thrombin suggested that EtOH inhibits PAR4 signaling not PAR1 since the second prolonged phase of [Ca2+]i is mediated by PAR4. Inhibition of PAR4-induced aggregation and [Ca2+]i elevation by EtOH supported the findings and EtOH might reduce Ca2+ influx through inhibition of PAR4. Furethermore, the difference between the platelet activation mechanisms of low concentrations of thrombin and PAR1-AP was suggested. PAR1-AP can aggregate platelets at least but might fail to activate phospholipase A2 required for sustaining stable aggregation since EtOH abolishes phospholipase A2 and thereby reduces thromboxane A2 generation. On the other, thrombin at low concentrations might have another pathway for activating platelet differently than PAR1-AP. Further characterization of the mechanisms involved in inhibition of platelet activation by EtOH may help develop new strategies to control thrombin-mediated platelet activation.

Genetic Evidence for a Predominant Role of PI3Kβ In ITAM— and Integrin-Mediated Signaling in Platelets

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 410-410
Author(s):  
Ilaria Canobbio ◽  
Lucia Stefanini ◽  
Lina Cipolla ◽  
Elisa Ciraolo ◽  
Cristian Gruppi ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Type I Collagen ◽  
Small Gtpase ◽  
Genetic Evidence ◽  
Type I ◽  
Wild Type ◽  
Wide Range ◽  
Induced Aggregation ◽  
Stimulation Of

Abstract Phosphoinositide 3-kinases (PI3K) have been implicated in platelet activation downstream of G protein-coupled receptors (GPCRs), ITAM-bearing receptors, as well as integrins. Among the multiple PI3K isoforms expressed in platelets, PI3Kγ and PI3Kβ are considered to be involved in GPCRs-mediated signaling leading to Rap1b activation and cell aggregation. However, a definitive genetic analysis of the relative contribution of PI3Kβ versus PI3Kγ in the different contexts of platelet activation has not been provided yet. PI3Kγ knockout mice have been widely investigated, but PI3Kγ has been recognized to mediate also kinase-independent effects. Moreover, deletion of p110β is embryonic lethal, and thus PI3Kβ knockout platelets have not been previously available for analysis. Knockin mice carrying a kinase dead form of either PI3Kβ or PI3Kγ have been recently generated by introducing K833R mutation in p110γ, or K805R mutation in p110β, respectively (Patrucco et al, Cell2004:118,375; Ciraolo et al, 2008, in press). In this study, we have compared platelet activation in mice homozygous for the kinase dead forms of PI3Kβ (PI3KβKD) or PI3Kγ (PI3KγKD). Platelet count and tail bleeding time were not altered in either PI3KγKD or PI3KβKD mice. Platelet aggregation induced by the thromboxane A2 analogue U46619, which stimulates GPCRs, was only slightly reduced in PI3KγKD platelets, and was not further inhibited by treatment with the selective PI3Kβ inhibitor TGX221. Accordingly, catalytic inactivation of PI3Kβ caused only a modest reduction of platelet aggregation induced by U46619. Similar results were also obtained upon stimulation of GPCRs for ADP or thrombin. ADP- and U46619-induced activation of the small GTPase Rap1b, a key regulator of platelet aggregation, was only reduced in PI3KγKD, but was almost completely suppressed in PI3KβKD, pointing to a major role for this isoform in the GPCRs-mediated regulation of the GTPase. Rap1b was normally activated upon stimulation of PI3KγKD platelets with the GPVI ligand convulxin (CVX), both in the absence and presence of the ADP scavenger apyrase. Moreover, CVX-induced aggregation of PI3KγKD platelets was only slightly reduced. By contrast, PI3KβKD platelets completely failed to aggregate in response to a wide range of concentrations of CVX, and accumulation of active GTP-bound Rap1b was almost undetectable. Accordingly, the PI3Kβ inhibitor TGX221 completely suppressed both Rap1b activation and aggregation induced by GPVI stimulation in either wild type or PI3KγKD platelets, both in the presence and absence of apyrase. PI3KγKD platelets adhered normally to immobilize type I collagen or fibrinogen, and displayed only a small defect of spreading on fibrinogen. Similarly, adhesion to collagen was comparable in PI3KβKD and wild type platelets. However, PI3KβKD revealed a severely impaired adhesion to fibrinogen. In addition, adherent cells completely failed to spread, and virtually no lamellipodia-forming platelets were observed. Altogether these results indicate both PI3Kγ and PI3Kβ are both involved in Rap1b activation and platelet aggregation upon stimulation of GPCRs, but provide the first genetic evidence that PI3Kβ, rather than PI3Kγ, plays an essential role in platelet activation downstream of the ITAM-bearing receptor GPVI, as well as in integrin αIIbβ3 outside-in signaling.

Modulation of Platelet Function by Recombinant Thrombomodulin Hematologic Implications.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 3898-3898
Author(s):  
Cafer Adiguzel ◽  
Omer Iqbal ◽  
Daniel Fareed ◽  
Debra Hoppensteadt ◽  
Walter Jeske ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Platelet Function ◽  
Whole Blood ◽  
Platelet Function Tests ◽  
Flow Cytometric ◽  
Platelet Release ◽  
Recombinant Thrombomodulin ◽  
Microparticle Formation ◽  
Induced Aggregation

Abstract Several recombinant thrombomodulin (r-TM) preparations have been developed for different clinical indications. While the in vitro effects of r-TM on blood coagulation parameters are extensively studied, the effect of this agent on platelet function tests such as the adhesion, aggregation, activation and secretion are not fully explored. The purpose of this study was to investigate the effect of a recombinant version of thrombomodulin (ART 123, Asahi, Pharmaceutical, Japan) on various platelet function tests. Platelet aggregation, platelet release and platelet activation by tissue factor (TF) utilizing flow cytometry studies were carried out. In the platelet aggregation studies, citrated whole blood was supplemented with graded amounts of r-TM, in a concentration range of 0–10 ug/ml in the blood of normal healthy volunteers (n=25). Platelet rich plasma (PRP) was prepared by controlled centrifugation (800g) for 15 minutes. Platelet count in the PRP was adjusted to 250,000/ul and aggregation studies were carried out using ADP (5 and 2.5 uM), and alpha thrombin (0.5 U/ml). Platelet activation studies were carried out using flow cytometric method utilizing citrated whole blood and recombinant TF and ADP as activators. In this procedure whole blood was supplemented with TM in a concentration range of 0–10 ug/ml and incubated. TF was then added and further incubated for an additional 2 minutes. Platelets were fixed and incubated with CD 61 and CD 62 antibodies and analyzed using the flow cytometer. Platelet release assays included the 14C serotonin release (SRA) from the washed platelets. In the platelet aggregation assay r-TM did not produce any significant inhibition of agonist induced aggregation with ADP and epinephrine, however, r-TM produced a strong concentration inhibition of thrombin induced aggregation with an IC 50 of 0.42 ug/ml. In the flow studies, r-TM produced an initial augmentation of the generation of microparticles at concentrations up to 0.31 ug/ml (ranges; 5–20%). However at concentrations &gt; 0.31 ug/ml r-TM produced a concentration dependent inhibition of the microparticle formation with an IC 50 of 2.5 ug/ml. At concentrations of &gt;5.0 ug/ml a complete inhibition of TF mediated microparticle formation was noted. Interestingly, r-TM did not produce any inhibition of the p-selectin expression at all concentrations studied. In the SRA, r-TM did not produce any release at concentrations up to 10 ug/ml. However, r-TM produced a strong inhibition of the alpha thrombin induced SRA release. These studies demonstrate that although in the agonist induced platelet aggregation studies r-TM does not produce any modulation of platelet aggregation responses with the exception of thrombin, in the flow cytometric studies it produces a biophasic response. In a concentration range of 0 - .31 ug/ml it produced a slight augmentation of the TF mediated platelet activation. However, at higher concentrations it produced an inhibition of the platelet microparticle formation. Interestingly, there was no effect of r-TM on p-selectin activation. These studies suggest that although r-TM does not produce any inhibition of the agonist induced aggregation of platelets, it can inhibit the TF mediated microparticle formation. Moreover, since r-TM at concentrations of up to 10 ug/ml does not produce any effect on p-selectin expression. It is unlikely to produce any primary hemostatic compromise in a therapeutic range of 2–6 ug/ml.

scholarly journals Linagliptin Regulates the Mitochondrial Respiratory Reserve to Alter Platelet Activation and Arterial Thrombosis

2020 ◽  
Vol 11 ◽  
Author(s):  
Yi Li ◽  
Rong Li ◽  
Ziqian Feng ◽  
Qin Wan ◽  
Jianbo Wu
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Arterial Thrombosis ◽  
Morphological Changes ◽  
High Dose ◽  
Chow Diet ◽  
Dipeptidyl Peptidase 4 ◽  
Consumption Rates ◽  
Normal Chow ◽  
Induced Aggregation

Background: The pharmacological inhibition of dipeptidyl peptidase-4 (DPP-4) potentiates incretin action, and DPP-4 is a drug target for type 2 diabetes and reducing cardiovascular risk. However, little is known about the non-enteroendocrine pathways by which DPP-4 might contribute to ischaemic cardiovascular events.Methods: We tested the hypothesis that inhibition of DPP-4 can inhibit platelet activation and arterial thrombosis by preventing platelet mitochondrial dysfunction and release. The effects of pharmacological DPP-4 inhibition on carotid artery thrombosis, platelet aggregation, and platelet mitochondrial respiration signaling pathways were studied in mice.Results: Platelet-dependent arterial thrombosis was significantly delayed in mice treated with high dose of linagliptin, a potent DPP-4 inhibitor, and fed normal chow diet compared to vehicle-treated mice. Thrombin induced DPP-4 expression and activity, and platelets pretreated with linagliptin exhibited reduced thrombin-induced aggregation. Linagliptin blocked phosphodiesterase activity and contrained cyclic AMP reduction when thrombin stimulates platelets. Linagliptin increases the inhibition of platelet aggregation by nitric oxide. The bioenergetics profile revealed that platelets pretreated with linagliptin exhibited decreased oxygen consumption rates in response to thrombin. In transmission electron microscopy, platelets pretreated with linagliptin showed markedly reversed morphological changes in thrombin-activated platelets, including the secretion of α-granules and fewer mitochondria.Conclusion: Collectively, these findings identify distinct roles for DPP-4 in platelet function and arterial thrombosis.

Platelet Activation and Aggregation Induced by Recombinant von Willebrand Factors Reproducing Four Type 2B von Willebrand Disease Missense Mutations

1998 ◽  
Vol 79 (01) ◽  
pp. 211-216 ◽  
Author(s):  
Lysiane Hilbert ◽  
Claudine Mazurier ◽  
Christophe de Romeuf
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Von Willebrand Disease ◽  
Platelet Glycoprotein ◽  
Missense Mutations ◽  
Inhibit Platelet Aggregation ◽  
Wild Type ◽  
A1 Domain ◽  
Von Willebrand ◽  
Willebrand Factor

SummaryType 2B of von Willebrand disease (vWD) refers to qualitative variants with increased affinity of von Willebrand factor (vWF) for platelet glycoprotein Ib (GPIb). All the mutations responsible for type 2B vWD have been located in the A1 domain of vWF. In this study, various recombinant von Willebrand factors (rvWF) reproducing four type 2B vWD missense mutations were compared to wild-type rvWF (WT-rvWF) for their spontaneous binding to platelets and their capacity to induce platelet activation and aggregation. Our data show that the multimeric pattern of each mutated rvWF is similar to that of WT-rvWF but the extent of spontaneous binding and the capacity to induce platelet activation and aggregation are more important for the R543Q and V553M mutations than for the L697V and A698V mutations. Both the binding of mutated rvWFs to platelets and platelet aggregation induced by type 2B rvWFs are inhibited by monoclonal anti-GPIb and anti-vWF antibodies, inhibitors of vWF binding to platelets in the presence of ristocetin, as well as by aurin tricarboxylic acid. On the other hand, EDTA and a monoclonal antibody directed against GPIIb/IIIa only inhibit platelet aggregation. Furthermore, the incubation of type 2B rvWFs with platelets, under stirring conditions, results in the decrease in high molecular weight vWF multimers in solution, the extent of which appears correlated with that of plasma vWF from type 2B vWD patients harboring the corresponding missense mutation. This study supports that the binding of different mutated type 2B vWFs onto platelet GPIb induces various degrees of platelet activation and aggregation and thus suggests that the phenotypic heterogeneity of type 2B vWD may be related to the nature and/or location of the causative point mutation.

On The Mechanism Of Heparin-Induced Potentiation Of Platelet Aggregation

1981 ◽  
Author(s):  
M Yamamoto ◽  
K Watanabe ◽  
Y Ando ◽  
H Iri ◽  
N Fujiyama ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Coagulation Factors ◽  
Marked Enhancement ◽  
Matrix Bound ◽  
Induced Aggregation ◽  
Aggregation Of Platelets ◽  
Plasma Heparin

It has been suggested that heparin caused potentiation of aggregation induced by ADP or epinephrine. The exact mechanism of heparin-induced platelet activation, however, remained unknown. In this paper, we have investigated the role of anti-thrombin III ( AT ) in heparin-induced platelet activation using purified AT and AT depleted plasma. When ADP or epinephrine was added to citrated PRP one minute after addition of heparin ( 1 u/ml, porcine intestinal mucosal heparin, Sigma Co. USA ), marked enhancement of platelet aggregation was observed, compared with the degree of aggregation in the absence of heparin. However, in platelet suspensions prepared in modified Tyrode’s solution, heparin exhibited no potentiating effect on platelet aggregation induced by epinephrine or ADP. Potentiation of epinephrine- or ADP-induced platelet aggregation by heparin was demonstrated when purified AT was added to platelet suspensions at a concentration of 20 μg/ml. AT depleted plasma, which was prepared by immunosorption using matrix-bound antibodies to AT, retained no AT, while determination of α1-antitrypsinα2- macroglobulin and fibrinogen in AT depleted plasma produced values which corresponded to those of the original plasma when dilution factor was taken into account. The activities of coagulation factors were also comparable to those of the original plasma. Heparin exhibited potentiating effect on ADP- or epinephrine-induced aggregation of platelets in original plasma, but no effect in AT depleted plasma. When purified AT was added back to AT depleted plasma at a concentration of 20 μg/ml, potentiation of platelet aggregation by heparin was clearly demonstrated.Our results suggest that effect of heparin on platelet aggregation is also mediated by anti-thrombin III.

Enhanced platelet aggregation and activation under conditions of hypothermia

2007 ◽  
Vol 98 (12) ◽  
pp. 1266-1275 ◽  
Author(s):  
Ruben Xavier ◽  
Ann White ◽  
Susan Fox ◽  
Robert Wilcox ◽  
Stan Heptinstall
Keyword(s):  
Cardiac Surgery ◽  
Platelet Aggregation ◽  
Platelet Function ◽  
Whole Blood ◽  
Platelet Rich Plasma ◽  
Baseline Levels ◽  
Spontaneous Aggregation ◽  
High Concentrations ◽  
Induced Aggregation ◽  
P2y12 Antagonist

SummaryThe effects on platelet function of temperatures attained during hypothermia used in cardiac surgery are controversial. Here we have performed studies on platelet aggregation in whole blood and platelet-rich plasma after stimulation with a range of concentrations of ADP, TRAP, U46619 and PAF at both 28°C and 37°C. Spontaneous aggregation was also measured after addition of saline alone. In citrated blood, spontaneous aggregation was markedly enhanced at 28°C compared with 37°C. Aggregation induced by ADP was also enhanced. Similar results were obtained in hirudinised blood. There was no spontaneous aggregation in PRP but ADP-induced aggregation was enhanced at 28°C. The P2Y12 antagonist AR-C69931 inhibited all spontaneous aggregation at 28°C and reduced all ADP-induced aggregation responses to small, reversible responses. Aspirin had no effect. Aggregation was also enhanced at 28°C compared with 37°C with low but not high concentrations of TRAP and U46619. PAF-induced aggregation was maximal at all concentrations when measured at 28°C, but reversal of aggregation was seen at 37°C. Baseline levels of platelet CD62P and CD63 were significantly enhanced at 28°C compared with 37°C. Expression was significantly increased at 28°C after stimulation with ADP, PAF and TRAP but not after stimulation with U46619. Overall, our results demonstrate an enhancement of platelet function at 28°C compared with 37°C, particularly in the presence of ADP.

scholarly journals Cysteine sulfenylation by CD36 signaling promotes arterial thrombosis in dyslipidemia

2020 ◽  
Vol 4 (18) ◽  
pp. 4494-4507 ◽  
Author(s):  
Moua Yang ◽  
Wei Li ◽  
Calvin Harberg ◽  
Wenjing Chen ◽  
Hong Yue ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Hydrogen Peroxide ◽  
Platelet Aggregation ◽  
Platelet Activation ◽  
Arterial Thrombosis ◽  
Reactive Oxygen ◽  
Src Family Kinases ◽  
Oxygen Species ◽  
Carbon Nucleophiles ◽  
Platelet Proteins

Abstract Arterial thrombosis in the setting of dyslipidemia promotes clinically significant events, including myocardial infarction and stroke. Oxidized lipids in low-density lipoproteins (oxLDL) are a risk factor for athero-thrombosis and are recognized by platelet scavenger receptor CD36. oxLDL binding to CD36 promotes platelet activation and thrombosis by promoting generation of reactive oxygen species. The downstream signaling events initiated by reactive oxygen species in this setting are poorly understood. In this study, we report that CD36 signaling promotes hydrogen peroxide flux in platelets. Using carbon nucleophiles that selectively and covalently modify cysteine sulfenic acids, we found that hydrogen peroxide generated through CD36 signaling promotes cysteine sulfenylation of platelet proteins. Specifically, cysteines were sulfenylated on Src family kinases, which are signaling transducers that are recruited to CD36 upon recognition of its ligands. Cysteine sulfenylation promoted activation of Src family kinases and was prevented by using a blocking antibody to CD36 or by enzymatic degradation of hydrogen peroxide. CD36-mediated platelet aggregation and procoagulant phosphatidylserine externalization were inhibited in a concentration-dependent manner by a panel of sulfenic acid–selective carbon nucleophiles. At the same concentrations, these probes did not inhibit platelet aggregation induced by the purinergic receptor agonist adenosine diphosphate or the collagen receptor glycoprotein VI agonist collagen-related peptide. Selective modification of cysteine sulfenylation in vivo with a benzothiazine-based nucleophile rescued the enhanced arterial thrombosis seen in dyslipidemic mice back to control levels. These findings suggest that CD36 signaling generates hydrogen peroxide to oxidize cysteines within platelet proteins, including Src family kinases, and lowers the threshold for platelet activation in dyslipidemia.

scholarly journals Cigarette Smoke Extract Inhibits Platelet Aggregation by Suppressing Cyclooxygenase Activity

TH Open ◽  
2017 ◽  
Vol 01 (02) ◽  
pp. e122-e129
Author(s):  
Hitoshi Kashiwagi ◽  
Koh-ichi Yuhki ◽  
Yoshitaka Imamichi ◽  
Fumiaki Kojima ◽  
Shima Kumei ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Cigarette Smoke ◽  
Platelet Function ◽  
Inhibitory Effect ◽  
Cigarette Smoke Extract ◽  
Human Platelets ◽  
Receptor Subtypes ◽  
Ic50 Value ◽  
Induced Aggregation ◽  
Cox 1

AbstractThe results of studies that were performed to determine whether cigarette smoking affects platelet function have been controversial, and the effects of nicotine- and tar-free cigarette smoke extract (CSE) on platelet function remain to be determined. The aim of this study was to determine the effect of CSE on platelet aggregation and to clarify the mechanism by which CSE affects platelet function. CSE inhibited murine platelet aggregation induced by 9,11-dideoxy-9α,11α-methanoepoxy-prosta-5Z,13E-dien-1-oic acid (U-46619), a thromboxane (TX) A2 receptor agonist, and that induced by collagen with respective IC50 values of 1.05 ± 0.14% and 1.34 ± 0.19%. A similar inhibitory action of CSE was also observed in human platelets. CSE inhibited arachidonic acid–induced TXA2 production in murine platelets with an IC50 value of 7.32 ± 2.00%. Accordingly, the inhibitory effect of CSE on collagen-induced aggregation was significantly blunted in platelets lacking the TXA2 receptor compared with the inhibitory effect in control platelets. In contrast, the antiplatelet effects of CSE in platelets lacking each inhibitory prostanoid receptor, prostaglandin (PG) I2 receptor and PGE2 receptor subtypes EP2 and EP4, were not significantly different from the effects in respective control platelets. Among the enzymes responsible for TXA2 production in platelets, the activity of cyclooxygenase (COX)-1 was inhibited by CSE with an IC50 value of 1.07 ± 0.15% in an uncompetitive manner. In contrast, the activity of TX synthase was enhanced by CSE. The results indicate that CSE inhibits COX-1 activity and thereby decreases TXA2 production in platelets, leading to inhibition of platelet aggregation.

COMPARISON OF THE PLATELET AGGREGATION INDUCED BY THREE THROMBIN-LIKE ENZYMES OF SNAKE VENOMS AND THROMBIN

1987 ◽  
Author(s):  
C M Teng ◽  
F N Ko
Keyword(s):  
Platelet Aggregation ◽  
Platelet Activation ◽  
Creatine Phosphokinase ◽  
Antithrombin Iii ◽  
Atp Release ◽  
Creatine Phosphate ◽  
Snake Venoms ◽  
Rabbit Platelets ◽  
Bothrops Atrox ◽  
Induced Aggregation

Acutin was isolated from Agkistrodon acutus venom and batroxobin and thrombocytin were isolated from Bothrops atrox venom. These three thrombin-like enzymes had different specificity for platelet activation and fibrinogen clotting. The clotting activities were 700, 170 and 7 μ/mg for batroxobin, acutin and thrombocytin, respectively. They induced aggregation and ATP release of washed rabbit platelets. The aggregating activities were 102, 104 and 105 times less than that of thrombin for thrombocytin, acutin and batroxobin, respectively basing on the clotting unit. The platelet -activating potency was correlated with their effectiveness on the retractility and elasticity of the clots. Platelet aggregation induced by thrombin or thrombocytin could be inhibited by heparin with antithrombin III while that by acutin or batroxobin could not. The thrombin-like enzymes did not induce aggregation of thrombin-degranulated platelets even fibrinogen was added. Indomethacin showed weak inhibition on the aggregation while the ADP - scavenging system, creatine phosphate/creatine phosphokinase, or apyrase inhibited the aggregation induced by the three thrombin-like enzymes but not that by thrombin. In the presence of EGTA, only thrombin could induce ATP release from platelets. It is concluded that the aggregation induced by thrombin-like enzymes is different from that of thrombin and mainly due to ADP released from platelets.

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