ArticleLiterature Review

Platelet Function in Health and Disease: from Molecular Mechanisms, Redox Considerations to Novel Therapeutic Opportunities

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Abstract

Abstract Increased oxidative stress appears to be of fundamental importance in the pathogenesis and development of several disease processes. Indeed, it is well known that reactive oxygen species (ROS) exert critical regulatory functions within the vascular wall, and it is, therefore, plausible that platelets represent a relevant target for their action. Platelet activation cascade (including receptor-mediated tethering to the endothelium, rolling, firm adhesion, aggregation, and thrombus formation) is tightly regulated. In addition to already well-defined platelet regulatory factors, ROS may participate in the regulation of platelet activation. It is already established that enhanced ROS release from the vascular wall can indirectly affect platelet activity by scavenging nitric oxide (NO), thereby decreasing the antiplatelet properties of endothelium. On the other hand, recent data suggest that platelets themselves generate ROS, which may evoke pro-thrombotic responses, triggering many biological processes participating in atherosclerosis initiation, progression, and complication. That oxidative stress may alter platelet function is conceivable when considering that antioxidants play a role in the prevention of cardiovascular disease, although the precise mechanism accounting for changes attributable to antioxidants in atherosclerosis remains unknown. It is possible that the effects of antioxidants may be a consequence of their enhancing or promoting the antiplatelet effects of NO derived from both endothelial cells and platelets. This review focuses on current knowledge regarding ROS-dependent regulation of platelet function in health and disease, and summarizes in vitro and in vivo evidence for their physiological and potential therapeutic relevance. Antioxid. Redox Signal. 17, 1447-1485.

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... There is a growing recognition that platelets may facilitate inflammation and leukocyte recruitment to sites of vascular injury and may also modulate immune responses (5). Upon activation, platelets release from the α-granules a number of cytokines and chemokines, and many of them have been identified in atherosclerotic lesions (6). The release of inflammatory mediators boosts the inflammatory milieu at sites of platelet activation (e. g. vascular injury) leading to cell activation and cell migration/recruitment (7). ...
... However, there is no clear evidence of in vivo platelet activation in the setting of IBD and of the possible influence of anti-inflammatory therapy on modulating platelet activation in this setting. Our group has previously demonstrated a clear relationship between inflammatory stimuli, lipid peroxidation and in vivo platelet activation in several clinical settings (5,6,11). Thus, we hypothesised that a similar vicious cycle may be implicated in IBD. ...
... Intravascular platelet aggregates have been also detected in mucosal biopsies of UC patients (24) Moreover, enhanced platelet P-selectin expression and increased number of circulating platelet aggregates have been found in the mesenteric venous circulation draining the inflamed bowel in UC, indicating that platelet activation may occur in the intestinal microcirculation (24). Platelets regulate a variety of inflammatory responses, shedding and releasing key proteins able to control inflammatory processes (5,6,25). Among them, CD40L, a potent immune-regulator and pro-inflammatory molecule, is shed into circulation by CD40L-positive platelets (5). ...
... There is a growing recognition that platelets may facilitate inflammation and leukocyte recruitment to sites of vascular injury and may also modulate immune responses (5). Upon activation, platelets release from the α-granules a number of cytokines and chemokines, and many of them have been identified in atherosclerotic lesions (6). The release of inflammatory mediators boosts the inflammatory milieu at sites of platelet activation (e. g. vascular injury) leading to cell activation and cell migration/recruitment (7). ...
... However, there is no clear evidence of in vivo platelet activation in the setting of IBD and of the possible influence of anti-inflammatory therapy on modulating platelet activation in this setting. Our group has previously demonstrated a clear relationship between inflammatory stimuli, lipid peroxidation and in vivo platelet activation in several clinical settings (5,6,11). Thus, we hypothesised that a similar vicious cycle may be implicated in IBD. ...
... Intravascular platelet aggregates have been also detected in mucosal biopsies of UC patients (24) Moreover, enhanced platelet P-selectin expression and increased number of circulating platelet aggregates have been found in the mesenteric venous circulation draining the inflamed bowel in UC, indicating that platelet activation may occur in the intestinal microcirculation (24). Platelets regulate a variety of inflammatory responses, shedding and releasing key proteins able to control inflammatory processes (5,6,25). Among them, CD40L, a potent immune-regulator and pro-inflammatory molecule, is shed into circulation by CD40L-positive platelets (5). ...
Article
Patients with inflammatory bowel disease (IBD) are at higher risk of venous thromboembolism and coronary artery disease despite having a lower burden of traditional risk factors. Platelets from IBD patients release more soluble CD40 ligand (CD40L), and this has been implicated in IBD platelet hyper-activation. We here measured the urinary F2-isoprostane 8-iso-prostaglandin (PG)2α (8-iso-PGF2α), urinary 11–dehydro–thromboxane (TX) B2 (11-dehydro–TXB2) and plasma CD40L in IBD patients, and explored the in vitro action of anti-tumour necrosis factor (TNF)–α antibody infliximab on IBD differentiating megakaryocytes. Urinary and blood samples were collected from 124 IBD patients and 37 healthy subjects. Thirteen IBD patients were also evaluated before and after 6–week infliximab treatment. The in vitro effect of infliximab on patient-derived megakaryocytes was evaluated by immunoflorescence microscopy and by flow cytometry. IBD patients had significantly (p<0.0001) higher urinary 8–iso–PGF2α and 11–dehydro–TXB2 as well as plasma CD40L levels than controls, with active IBD patients displaying higher urinary and plasma values when compared to inactive patients in remission. A 6-week treatment with infliximab was associated with a significant reduction of the urinary excretion of 8–iso–PGF2α and 11–dehydro–TXB2 (p=0.008) and plasma CD40L (p=0.001). Infliximab induced significantly rescued pro-platelet formation by megakaryocytes derived from IBD patients but not from healthy controls. Our findings provide evidence for enhanced in vivo TX–dependent platelet activation and lipid peroxidation in IBD patients. Anti-TNF–α therapy with infliximab down-regulates in vivo isoprostane generation and TX biosynthesis in responder IBD patients. Further studies are needed to clarify the implication of infliximab induced-proplatelet formation from IBD megakaryocytes. Supplementary Material to this article is available online at www.thrombosis-online.com.
... Several enzymatic pathways contribute to ROS production in the vascular bed ( Fig. 7) (106,227,273,285,355) and the connection between oxidant stress and hemostatic balance as mediated by endothelial dysfunction is well established. Thus, as demonstrated in other clinical settings, an increased ROS production in migraine could be responsible for clotting activation and consequent prothrombotic switch (134). ...
... Thus, as demonstrated in other clinical settings, an increased ROS production in migraine could be responsible for clotting activation and consequent prothrombotic switch (134). In addition, an unbalanced redox status could also provoke platelet activation, as platelets are a prime target for ROS that are produced in or released from the vascular lumen (106). However, platelets themselves generate ROS and release proinflammatory cytokines and/or procoagulant factors, which makes them capable of sustaining a vicious cycle that creates an increased thrombophilic predisposition in migraine (79). ...
... 2. Effects on platelet activation. It is widely acknowledged that increased oxidative stress may promote platelet aggregation (106). Furthermore, activated platelets can produce both ROS (e.g., O 2 $-and H 2 O 2 ) (Fig. 7) (80,229,276) and NO (123). ...
Article
Significance: Migraine represents the third most prevalent and the seventh most disabling human disorder. Approximately 30% of migraine patients experience transient, fully reversible, focal neurological symptoms (aura) preceding the attack. Recent Advances: Awareness of the hypothesis that migraine actually embodies a spectrum of illnesses-ranging from episodic to chronic forms-is progressively increasing and poses novel challenges for clarifying the underlying pathophysiological mechanisms of migraine as well as for the development of novel therapeutic interventions. Several theories have evolved to the current concept that a combination of genetic, epigenetic, and environmental factors may play a role in migraine pathogenesis, although their relative importance is still being debated. Critical issues: One critical issue that deserves a particular attention is the role of oxidative stress in migraine. Indeed, potentially harmful oxidative events occur during the migraine attack and long-lasting or frequent migraine episodes may increase brain exposure to oxidative events that can lead to chronic transformation. Moreover, a wide variety of dietary, environmental, physiological, behavioral, and pharmacological migraine triggers may act through oxidative stress, with clear implications for migraine treatment and prophylaxis. Interestingly, almost all current prophylactic migraine agents exert antioxidant effects. Future directions: Increasing awareness of the role of oxidative stress and/or decreased antioxidant defenses in migraine pathogenesis and progression to a chronic condition lays the foundations for the design of novel prophylactic approaches, which, by reducing brain oxidative phenomena, could favorably modify the clinical course of migraine. Antioxid. Redox Signal. 00, 000-000.
... The negative effect of these radicals is neutralized by diverse oxidant scavengers, Fe 2+ and Cu + chelators and a large series of antioxidant enzymes (31). The maintenance of an optimal balance between the oxygen reactive species and the corresponding protective molecules is critical for cell survival and the preservation of healthy tissues (32). Recent evidence supports the idea that ozone might be used to increase the antioxidant capacity of blood and its derivates. ...
... Carbohydrates, enzymes, DNA and RNA can also be affected by ozone overdose. The high ozone concentration may have altered platelet structure and the configuration of some key proteins involved in the plasma coagulation process, resulting in its disruption (32). ...
Article
Background and Objective Until now, ozone has been used in a rather empirical way. This in-vitro study investigates, for the first time, whether different ozone treatments of plasma rich in growth factors (PRGF) alter the biological properties and outcomes of this autologous platelet-rich plasma.Material and Methods Human plasma rich in growth factors was treated with ozone using one of the following protocols: a continuous-flow method; or a syringe method in which constant volumes of ozone and PRGF were mixed. In both cases, ozone was added before, during and after the addition of calcium chloride. Three ozone concentrations, of the therapeutic range 20, 40 and 80 μg/mL, were tested. Fibrin clot properties, growth factor content and the proliferative effect on primary osteoblasts and gingival fibroblasts were evaluated.ResultsOzone treatment of PRGF using the continuous flow protocol impaired formation of the fibrin scaffold, drastically reduced the levels of growth factors and significantly decreased the proliferative potential of PRGF on primary osteoblasts and gingival fibroblasts. In contrast, treatment of PRGF with ozone using the syringe method, before, during and after the coagulation process, did not alter the biological outcomes of the autologous therapy.Conclusion These findings suggest that ozone dose and the way that ozone combines with PRGF may alter the biological potential and therapeutic outcomes of PRGF.
... Two important mediators of the responses to Ifnγ are ROS and NO [19][20][21]. The role of ROS in contributing to platelet aggregation, for example, has been well documented [29]. However, Ifnγ induced ROS played a negligible role during aggregation of APECs. ...
... However, Ifnγ induced ROS played a negligible role during aggregation of APECs. On the other hand, platelet aggregation is inhibited by NO generated by both platelets and endothelial cells [29]. Also, NO generated in response to Ifnγ inhibits T cell adhesion to the endothelium in the presence of Tnfα [30]. ...
Article
Full-text available
Interferon-gamma (Ifnγ), a key macrophage activating cytokine, plays pleiotropic roles in host immunity. In this study, the ability of Ifnγ to induce the aggregation of resident mouse adherent peritoneal exudate cells (APECs), consisting primarily of macrophages, was investigated. Cell-cell interactions involve adhesion molecules and, upon addition of Ifnγ, CD11b re-localizes preferentially to the sites of interaction on APECs. A functional role of CD11b in enhancing aggregation is demonstrated using Reopro, a blocking reagent, and siRNA to Cd11b. Studies with NG-methyl-L-arginine (LNMA), an inhibitor of Nitric oxide synthase (Nos), NO donors, e.g., S-nitroso-N-acetyl-DL-penicillamine (SNAP) or Diethylenetriamine/nitric oxide adduct (DETA/NO), and Nos2-/- mice identified Nitric oxide (NO) induced by Ifnγ as a key regulator of aggregation of APECs. Further studies with Nos2-/- APECs revealed that some Ifnγ responses are independent of NO: induction of MHC class II and CD80. On the other hand, Nos2 derived NO is important for other functions: motility, phagocytosis, morphology and aggregation. Studies with cytoskeleton depolymerizing agents revealed that Ifnγ and NO mediate the cortical stabilization of Actin and Tubulin which contribute to aggregation of APECs. The biological relevance of aggregation of APECs was delineated using infection experiments with Salmonella Typhimurium (S. Typhimurium). APECs from orally infected, but not uninfected, mice produce high amounts of NO and aggregate upon ex vivo culture in a Nos2-dependent manner. Importantly, aggregated APECs induced by Ifnγ contain fewer intracellular S. Typhimurium compared to their single counterparts post infection. Further experiments with LNMA or Reopro revealed that both NO and CD11b are important for aggregation; in addition, NO is bactericidal. Overall, this study elucidates novel roles for Ifnγ and Nos2 in regulating Actin, Tubulin, CD11b, motility and morphology during the aggregation response of APECs. The implications of aggregation or “group behavior” of APECs are discussed in the context of host resistance to infectious organisms.
... 4 Noteworthy, oxidant stress and enhanced lipid peroxidation are also key players in the initiation of persistent platelet activation both in T2D and in obesity. [5][6][7] Platelets, in turn, play a fundamental role in cancer progression, as strongly suggested by recent experimental and epidemiological studies in human cancers, [8][9][10] including BC. 11 Thus, based on the available body of evidence, we hypothesized that increased oxidative stress in BC patients could induce enhanced lipid peroxidation, which, in turn, could contribute to platelet activation and poor clinical outcome. ...
... Nonetheless, these observations are consistent with previous findings from our group demonstrating that oxidant stress and enhanced lipid peroxidation are key players in the initiation of persistent platelet activation in various clinical settings, including T2D and obesity. [5][6][7] Here we provide evidence of the occurrence of a pro-oxidant status (as reflected by increased 8-iso-PGF 2a urinary excretion levels) in BC women, which was independently associated to insulin resistance (a key feature of T2D and obesity) and hormone receptor status. While insulin resistance has been proposed as a pathogenetic mechanism linking obesity, T2D, and BC, 2 the finding of enhanced lipid peroxidation in BC is in agreement with previous findings demonstrating the occurrence of elevated levels of urinary isoprostanes in patients with invasive BC. 30 Furthermore, higher oxidative stress was found in ER(1) compared with ER(2) BC patients, as previously suggested. ...
Article
The hypothesis that increased oxidative stress in breast cancer (BC) patients could induce enhanced lipid peroxidation, which, in turn, would contribute to platelet activation and poor clinical outcome is attractive. To address this issue, we investigated pre-surgical urinary 8-iso-prostaglandin (PG)F2α (marker of in vivo oxidative stress) and 11-dehydro-thromboxane (TX)B2 (marker of in vivo platelet activation) levels in patients with primary BC (n=115) compared with control women paired for co-morbidities and their association with patients' metabolic profile and clinical prognostic factors. The results obtained showed that pre-surgical urinary excretion of both biomarkers was enhanced in BC patients compared to controls and was associated with patients' estrogen receptor (ER) expression, but not HER2 status or Ki67 proliferation index. Accordingly, both urinary biomarkers were increased in patients with luminal-like BC molecular subtypes compared with triple negative or HER2-enriched tumors. ER status was an independent predictor of 8-iso-PGF2α urinary levels, which, in turn, significantly predicted 11-dehydro-TXB2 urinary levels together with disease stage and ER status. The prognostic value of 11-dehydro-TXB2 was then evaluated showing a significant correlation with BC pathological response to neoadjuvant treatment. Furthermore, among relapsing patients, those with elevated urinary biomarker levels were more likely to develop distant metastasis rather than local recurrence. In conclusion, we may speculate that enhanced oxidative stress due to estrogen-related mechanisms might cause a condition of persistent platelet activation capable of sustaining BC growth and progression through the release of bioactive stored molecules, ultimately contributing to tumor invasiveness. Further studies specifically addressing this hypothesis are presently needed. This article is protected by copyright. All rights reserved.
... 1). Platelets activate in response to agonists, forming a loose aggregate after downstream activation of integrin αIIbβ3, which allows platelet-platelet interaction via ligands including fibrinogen, fibrin, von Willebrand factor (VWF), and fibronectin [11][12][13][14] in the process of platelet aggregation. In response to strong agonists, a subset of activated platelets expose phosphatidylserine (PS) on their surface and are able Fig. 1 The role of platelets in thrombus formation in acute coronary syndromes. ...
... 14,81 This triggers the conformation change of the extracellular domain of αIIbβ3 to its high affinity state, allowing engagement of fibrinogen or VWF, which is necessary for platelet aggregation. 11,13,14 As platelets aggregate, occupied αIIbβ3 integrins cluster and trigger outside-in signaling that stabilizes the aggregate. 82 Development of monoclonal antibodies such as PAC-1 and LIBS1 that are specific for the active state of αIIbβ3 enable us to distinguish between platelets that retain a resting integrin versus those with the activated integrin capable of mediating aggregation. ...
Article
Primary and secondary prevention of cardiovascular disease remain a public health priority. Effective risk stratification of patients is a central requisite for effective preventative care and several scoring systems incorporating biomarkers have been used for prognostication in patients to guide intervention decisions. Thrombosis of atherosclerotic coronary arteries is the main mechanism behind the acute coronary syndromes and since platelets play a pivotal role in the pathogenesis of thrombosis, atherogenesis, and angiogenesis, platelet-derived biomarkers are an attractive concept. This review assesses the potential and the limitations of a range of platelet-based assays as biomarkers for coronary artery disease. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.
... Of fundamental importance in this system is EC-derived NO, which is a potent inhibitor of both platelet aggregation and adhesion to the EC monolayer (181). NO causes a decrease in platelet cytosolic Ca 2+ , which influences platelet glycoprotein activation, and therefore, platelet adhesion (63). NO can also inhibit fibrin deposition, further attenuating thrombus formation (137). ...
... The ability of NO to regulate the expression of adhesion molecules also plays a key role in the NO-mediated inhibition of coagulation and thrombus formation. It has long been appreciated that NO is a potent inhibitor of platelet aggregation and adhesion to ECs (181) by increasing intracellular cGMP, resulting in a subsequent decrease in intracellular Ca 2+ (63). This suppression of intracellular Ca 2+ influx inhibits a conformational change in the platelet glycoprotein IIb/IIIa, essentially decreasing the number of fibrinogen binding sites on the platelet surface, thereby limiting platelet cross-linking and clot strength (41,154). ...
Article
The pulmonary endothelium represents a heterogeneous cell monolayer covering the luminal surface of the entire lung vasculature. As such, this cell layer lies at a critical interface between the blood, airways, and lung parenchyma, and must act as a selective barrier between these diverse compartments. Lung endothelial cells are able to produce and secrete mediators, display surface receptor, and cellular adhesion molecules, and metabolize circulating hormones to influence vasomotor tone, both local and systemic inflammation, and coagulation functions. In this review, we will explore the role of the pulmonary endothelium in each of these systems, highlighting key regulatory functions of the pulmonary endothelial cell, as well as novel aspects of the pulmonary endothelium in contrast to the systemic cell type. The interactions between pulmonary endothelial cells and both leukocytes and platelets will be discussed in detail, and wherever possible, elements of endothelial control over physiological and pathophysiological processes will be examined. © 2015 American Physiological Society. Compr Physiol 5: 531-559, 2015.
... Platelets are the discoid anuclear blood cells derived from megakaryocytes with a short a lifespan of 7-9 days, and play a critical role in hemostasis and thrombosis [1]. In the physiological system, the number of circulating platelets is precisely managed by an antagonistic balance between their production and destruction. ...
... Thus, it is postulated that oxidative stress-induced pathologies trigger altered platelet function/apoptosis. Of late, it is demonstrated that the oxidative stress and high shear stress inside stenotic atherosclerotic arteries trigger apoptosis in platelets causing the release of microparticles (MPs), into extracellular space [1][2][3]. Several studies suggest that the MPs shed from apoptotic platelets provide a catalytic phospholipid surface for the assembly of blood coagulation factors, thereby promoting the coagulation cascade, inflammation and cardiovascular disease progression [4]. Apart from this, it is also shown that several clinically significant therapeutic agents which include anti-inflammatory drugs (NSAIDS), cardiac medications, and anti-cancerous drugs are known to induce oxidative stress-mediated platelet apoptosis [5][6][7][8][9]. ...
Article
Full-text available
Oxidative stress-induced platelet apoptosis is one among the many causes for the development and progression of many disorders like cardiovascular diseases, arthritis, Alzheimer's disease and many chronic inflammatory responses. Many studies have demonstrated the less optimal effect of N-acetyl cysteine (NAC) in oxidative stress-induced cellular damage. This could be due to its less lipophilicity which makes it difficult to enter the cellular membrane. Therefore in the present study, lipophilic sila-amide derivatives (6a and 6b) synthesized through the reaction of NAC with 3-Aminopropyltrimethylsilane and aminomethyltrimethylsilane were used to determine their protective property against oxidative stress-induced platelet apoptosis. At a concentration of 10 µM, compound 6a and 6b were able to significantly inhibit Rotenone/H2O2 induced platelet apoptotic markers like reactive oxygen species, intracellular calcium level, mitochondrial membrane potential, cytochrome c release from mitochondrial to the cytosol, caspase-9 and -3 activity and phosphatidylserine externalization. Therefore, the compounds can be extrapolated as therapeutic agents to protect platelets from oxidative stress-induced platelet apoptosis and its associated complications.
... ONOO − may react with tyrosyl residues to form 3-NO 2 -Tyr, and with thiols to generate S-nitrosothiols, reducing the inhibitory effect of nitrogen oxide. ONOO− may also act as a substrate for cyclooxygenase peroxidase, causing prostaglandin endoperoxide H synthase activation and increased prostaglandin synthesis, 67 and thus, thromboxane A 2 synthesis. 68 ONOO− may also induce the formation of 8-isoprostaglandin F2α 69 and, as the effect might increase calcium release from intracellular stores, induce changes in platelet morphology and amplify platelet aggregation in response to agonists. ...
... 68 ONOO− may also induce the formation of 8-isoprostaglandin F2α 69 and, as the effect might increase calcium release from intracellular stores, induce changes in platelet morphology and amplify platelet aggregation in response to agonists. 27,67 Together, these complex interactions support the possibility that oxidative stress has a proatherogenic role 70 and increases cardiovascular risk. Also, we can conclude about association between ROS and the important role of cardiovascular comorbidities in COPD. ...
Article
Full-text available
Background: COPD represents a major global health issue, which is often accompanied by cardiovascular diseases. A considerable body of evidence suggests that cardiovascular risk is elevated by the activation of blood platelets, which in turn is exacerbated by inflammation. As reactive oxygen species are believed to be an important factor in platelet metabolism and functioning, the aim of our study was to perform a complex assessment of mitochondrial function in platelets in chronic smoke exposed animals with COPD-like lung lesions. Materials and methods: Eight-week-old, male Dunkin Hartley guinea pigs (the study group) were exposed to the cigarette smoke from commercial unfiltered cigarettes (0.9 mg/cig of nicotine content) or to the air without cigarette smoke (control group), using the Candela Constructions® exposure system. The animals were exposed for 4 hours daily, 5 days a week, with 2×70 mL puff/minute, until signs of dyspnea were observed. The animals were bled, and isolated platelets were used to monitor blood platelet respiration. The mitochondrial respiratory parameters of the platelets were monitored in vitro based on continuous recording of oxygen consumption by high-resolution respirometry. Results: An elevated respiration trend was observed in the LEAK-state (adjusted for number of platelets) in the smoke-exposed animals: 6.75 (5.09) vs 2.53 (1.28) (pmol O 2 /[s ⋅ 1×10 ⁸ platelets]); bootstrap-boosted P 1α =0.04. The study group also demonstrated lowered respiration in the ET-state (normalized for protein content): 12.31 (4.84) vs 16.48 (1.72) (pmol O 2 /[s ⋅ mg of protein]); bootstrap-boosted P 1α =0.049. Conclusion: Our results suggest increased proton and electron leak and decreased electron transfer system capacity in platelets from chronic smoke-exposed animals. These observations may also indicate that platelets play an important role in the pathobiology of COPD and its comorbidities and may serve as a background for possible therapeutic targeting. However, these preliminary outcomes should be further validated in studies based on larger samples.
... Platelets play a crucial role in the vascular homeostasis and in the onset of thrombotic complications in different atherosclerotic diseases [1]. Several physiologic and pathologic factors are able to induce platelet activation [2]. Among others, oxidative stress has been proposed as a regulator of platelet function, representing a common feature of many cardiovascular diseases. ...
... The purpose of this review is to summarize available evidence on (1) data supporting the role of Nox in platelet ROS formation and platelet activation; (2) atherosclerotic diseases in which Nox up-regulation is implicated; (3) interventional trials with specific and non-specific inhibitors of Nox, including statins, polyphenols, apocynin and more recent evidence on some nutrients, such as olive oil. ...
... (7) Hyperactivation of platelets in patients with alcoholism results in higher oxidative stress. (8) Platelet activation generate reactive oxygen species (ROS), (9) express CD40L, and releases its soluble form (sCD40L), which acts as an inflammatory mediator. (10) Increase in CD40L promotes plateletleucocyte aggregation. ...
... Activated platelets release alpha/dense granules, which play a major role in hemostasis, leucocyte recruitment, repair, and regeneration. (9) Granule release is the key function of platelets, which controls the vascular integrity and regulates hemostasis and inflammation. (33) Granule secretion requires fusion of plasma membrane t-SNAREs (Syntaxin and SNAP-23) with vesicular membrane v-SNARES (VAMP-8, VAMP-3) in the presence of MUNC13-4, RAB27b, along with other small guanosine triphosphatases. ...
Article
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Hyperoxidized albumin promotes inflammation and modulates several immune cells in severe alcoholic hepatitis (SAH). Platelets mediate inflammation by interacting with immune cells, endothelium, and other cells. The role of hyperoxidized albumin in platelet activation and alteration of platelet phenotype/functions is not known. Quantitative platelet proteomics performed in 10 patients with SAH was compared with 10 patients with alcoholic cirrhosis and 10 healthy controls, respectively. Dysregulated pathways were identified and validated in a separate cohort (n = 40). Healthy platelets were exposed to patient plasma or purified albumin or ex vivo modified albumin (human‐mercaptalbumin, humannonmercaptalbumin‐1, and human nonmercaptalbumin 2) in the presence or absence of CD36 blockade, and platelet secretome was analyzed. Two hundred and two up‐regulated proteins linked to platelet activation, complement regulation, lipid transportation, and 321 down‐regulated proteins related to platelet hemostasis and coagulation (fold change ± 1.5, P 0.3, P
... This particular feature is commonly observed for commercial NSAIDs such as ASA and indomethacin, which provides improved outcome for patients undergoing acute coronary syndrome due to the specific inhibition of thromboxane A 2 (TXA 2 ) production by human platelets. 19 The lack of inhibitory activity in platelet aggregation induced by the remaining tested agonists also rule-out the impairment of common biochemical steps downstream to each stimulated G-protein coupled receptor (GPCR), such as the activation of Src kinases and the expression of the fibrinogen-binding integrin αIIbβ3 (GpIIb/IIIa), which are related to late phase platelet functions such as clot retraction 14,[20][21][22] . No significant inhibition of clot retraction (p<0,05 ANOVA-Tukey test) was observed for human whole blood in the presence of the molecules (Fig. 3A), reinforcing their selectivity towards AA-induced aggregation and suggesting the maintenance of PGH 2 levels, which is required for clot retraction and is found altered after treatment with indomethacin 23 . ...
... The results showed that all active pyrazolopyridine derivatives fulfilled the Lipinski rule of five (Tab. 3), which suggest the bioavailability of these compounds, reinforcing their promising profile for further in vivo experimental investigations as leading compounds to design new antiplatelet drugs against thromboembolic diseases.ConclusionsOverall, in this work we showed that the series of N'-benzylidene-carbohydrazide-1Hpyrazolo[3,4-b]pyridine derivatives(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22) includes six fluorescent TXS inhibitors derivatives (4, plasma coagulation factors. Active membrane binding to human platelet membranes was demonstrated as well as their high lipophilicity, which had direct correlation to the antiplatelet profile of the active hits. ...
Article
Abstract Cardiovascular diseases (CVDs) account for over 17 million deaths globally each year, with atherosclerosis as the underlying cause of most CVDs. Herein we describe the synthesis and in vitro mechanistic evaluation of novel N’-benzylidene-carbohydrazide-1H-pyrazolo[3,4-b]pyridines (3-22) designed as non-anionic antiplatelet agents and presenting a 30-fold increase in potency compared to aspirin. The mechanism underlying their antiplatelet activity was elucidated by eliminating potential targets through a series of in vitro assays including light transmission aggregometry, clot retraction, and quantitative ELISA, further identifying the reduction in biosynthesis of thromboxane B2 as their main mechanism of action. The intrinsic fluorescence of the compounds permits their binding to platelet membranes to be readily monitored. In silico structure-activity relationship, molecular docking and dynamics studies support the biological profile of the series revealing the molecular basis of their activity and their potential as future molecular therapeutic agents.
... The most common agonists include thrombin, collagen and Arachidonic Acid (AA). Thrombin stimulates mitochondrial membrane depolarization and endogenous generation of hydrogen peroxide [18]; moreover, it could potentially activate NADPH oxidase via Protein kinase C (PKC) phosphorylation [19]. Collagen binds to the platelet-specific receptor, GPVI/FcR , causing a rapid, transient disulfide-dependent homodimerization, and the production of intracellular ROS generated by the NADPH oxidase linked to GPVI via TNF receptorassociated factor 4 (TRAF4) [20]. ...
Article
Experimental and clinical studies provided evidence that formation of intra-platelet reactive oxidant species (ROS) is implicated in the process of thrombosis. Animal models demonstrated that enhanced ROS formation was associated with serious thrombotic complications and death. In recent years, nutritional and therapeutic approaches were tested to modulate ROS mediated thrombus formation. The use of a nutritional approach stems from the observation that foods rich in antioxidant elements, such as polyphenols, were able to modulate ROS formation. Similarly, some drugs used for different diseases (i.e. statins) showed the ability to modulate oxidative stress. Aim of this review is to summarize current evidences supporting the role of nutrients rich in polyphenols, such as olive oil and cocoa, and of some drugs, such as statins as antiplatelet agents interfering with the Nicotinamide Adenine Dinucleotide Phosphate (NADPH) Oxidase signaling. Indeed, for nutrients and statins, the antiplatelet activity seems to be dependent, at least in part, upon the inhibition of platelet NADPH oxidase-derived ROS formation, resulting in down-regulation of isoprostanes, which are pro-aggregating molecules, and up-regulation of nitric oxide, which is a platelet inhibitor.
... Ongoing inflammatory conditions lead to increased proliferation in the megakaryocytic series and relative thrombocytosis (19). Previous studies demonstrated the relationship of adverse cardiovascular outcomes with both increased platelet and decreased lymphocyte counts (20)(21)(22). ...
Article
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OBJECTIVE: In this study, we aimed to investigate the relation of platelet to lymphocyte ratio (PLR) in saphenous vein graft disease (SVGD) in patients with stable angina pectoris after coronary artery bypass graft surgery. METHODS: A total of 455 patients were included in the study. There were 210 patients with SVGD and 245 patients without SVGD. The effects of different variables on SVGD were computed in logistic regression analysis. RESULTS: The platelet count, lymphocyte count, PLR, high-density lipoprotein (HDL), Na, and ALT were significantly associated with SVGD. In multivariate regression analysis, HDL and PLR were found to be significantly associated with SVGD. CONCLUSION: To the best of our knowledge, this is the first study showing the significant association of PLR with SVGD. This study suggests that PLR can be used as a marker of SVGD because it is an easily available and inexpensive test.
... Instead, they possess complicated structural features that are related to their functional and biochemical activities. [19] Platelets contain granules which are an important source of inflammatory mediators. They have surface adhesion molecules which enable them to bind to the walls of damaged blood vessels and contribute to the inflammatory response. ...
Article
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Immunity is divided into two parts – the innate and adaptive responses. The innate immune system relies upon limited receptors to detect invading pathogens but compensates by targeting conserved microbial components that are shared by the large groups of pathogens. Various cells involved in innate immune responses include neutrophils, macrophages, dendritic cells, mast cell, neutrophils, natural killer cells, and eosinophils. The innate immune responses are the first line of defense against invading pathogens. They are also required to initiate specific adaptive immune responses. The second kind of protection is adaptive immunity which develops throughout our lives. It uses two basic strategies – Humoral immunity which works to eliminate antigens that are extracellular and cellular immunity which deals with antigen residing within a host cell. T- and B-lymphocytes are the main self-defense weapons of the adaptive immune system. Adaptive immunity relies upon a clonal system with each T-cell and B-cell expressing its own unique receptor.
... At the histological level, immunoglobulins, T lymphocytes, plasma cells, and the presence of antigen-antibody complex in the atheroma plaque suggest the importance of inflammation in the formation and progression of plaque [10]. Ongoing inflammatory conditions lead to megakaryocytic proliferation and thrombocytosis [11]. Platelets are 2e4 mm in diameter. ...
Article
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Carotid artery stenosis (CAS) is primarily caused by atherosclerotic plaque. Progressive inflammation may contribute to the rupture of an atherosclerotic plaque. The platelet-to-lymphocyte ratio (PLR) is a new and simple marker that indicates inflammation. In this study, we aimed to investigate the use of the PLR to determine the severity of CAS. One hundred forty patients were chosen from among patients who underwent carotid angiography in our institution. Symptomatic patients with stenosis >50% in the carotid arteries and asymptomatic patients with stenosis >80% were diagnosed via carotid angiography as having critical stenosis. Patients were classified into two groups. Group 1 included patients who had critical CAS, whereas Group 2 included patients with noncritical CAS, as determined by carotid angiography. Correlations between the PLR and the severity of CAS were analyzed. There were no significant differences in sex and age between the two groups. The PLR was 162.5 ± 84.7 in the noncritical CAS group patients and 94.9 ± 60.3 in the critical CAS group patients (p < 0.0001). The PLR value of 117.1 had 89% sensitivity and 68% specificity for CAS [95% confidence interval, 0.043–0.159; area under the curve, 0.101 ± 0.03)]. In this study, we have shown that PLR values may be associated with critical stenosis in at least one of the carotid arteries. Furthermore, PLR values may be used to predict critical stenosis in the carotid arteries.
... Critical Limb Ischemia (CLI), the most advanced stage of peripheral artery disease (PAD), is characterized by ischemic rest pain or tissue loss as well as a profound risk for cardiovascular complications and mortality [1,2]. Abnormal platelet function with an increased tendency to aggregate is implicated in the pathogenesis of atherosclerosis [3] and development of superimposed acute ischemic events [4][5][6][7]. Antiplatelet therapy reduces the risk for future cardiovascular events (CVE) in patients with previous cardiovascular disease and is therefore the cornerstone of medical therapy in PAD [8]. ...
Article
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Patients with critical limb ischemia (CLI) have a high risk to develop cardiovascular events (CVE). We hypothesized that in CLI patients platelets would display increased baseline activation and reactivity. We investigated baseline platelet activation and platelet reactivity in patients with CLI. In this study baseline platelet activation and platelet reactivity in response to stimulation of all major platelet activation pathways were determined in 20 CLI patients (11 using aspirin and 9 using vitamin K-antagonists) included in the Juventas-trial (clinicaltrials.gov NCT00371371) and in 17 healthy controls. Platelet activation was quantified with flow cytometric measurement of platelet P-selectin expression and fibrinogen binding. CLI patients not using aspirin showed higher baseline platelet activation compared to healthy controls. Maximal reactivity to stimulation of the collagen and thrombin activation pathway was decreased in CLI patients compared to healthy controls. In line, attenuated platelet reactivity to stimulation of multiple activation pathways was associated with several traditional risk factors for cardiovascular disease. Baseline platelet activation was increased in CLI patients, whereas the reactivity of circulating platelets to several stimulatory agents is decreased. Reactivity of platelets was inversely correlated with cardiovascular risk factors.
... This in turns leads to increased expression of endothelin-1, a vasoconstrictor, resulting in vasoconstriction and vaso-oclusion. Interestingly, it has been previously demonstrated that NOS gene therapy down-regulates the expression of ICAM1, VCAM1, E-and P-selectin, and inhibits the activation of platelets [50,51]. This then suggests a role for NO in endothelial inactivation and further strengthens the fact that reduced bioavailability of NO in SCD may contribute to platelet activation and increased expression of adhesion molecules. ...
Article
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Sickle cell disease (SCD) is a haematological disease that affects multiple organs, thus eliciting episodes of chronic pain, acute anaemia and infection, due to a single nucleotide mutation in the β-globin gene, which results in the substitution of a glutamic acid residue in place of valine on the β-globin chain of the resultant haemoglobin protein molecule, the sickle haemoglobin (HbS). SCD is a major cause of morbidity and mortality characterized by episodes of vaso-occlusive crises, pain syndromes and end organ dysfunctions. Its global prevalence is highest in Sub-Saharan Africa with 75% of global births living in this region, of which Nigeria has the highest number of SCD patients with about 100.000 births each year. The burden of SCD in the sub-Saharan region of Africa is enormous. Emotional, financial and total healthcare costs are monumental. An understanding of the mechanism underlying the vaso-occlusive crises, pain syndromes, inflammatory conditions and other sequelae of SCD appears to be essential in providing more rational treatments. The present review discuses the prevalence of SCD in Africa, molecular mechanisms underlying SCD episodic crises including vaso-occlusive syndrome, anaemia and infection. Current available treatments modalities in Sub-Saharan Africa and possible new treatment methods that cure SCD are re-examined in light of these mechanisms.
... The cGMP/PKG-dependant signalling mechanism also promotes a potent inhibition of platelet aggregation [161]. NO's anti-thombotic properties also include inhibition of platelet activation and adhesion to the endothelium, TF expression and thrombin formation [58]. NO is also an important antagonist of inflammation [47]. ...
Article
Full-text available
Sickle cell disease and β-thalassaemia are inherited haemoglobinopathies resulting in structural and quantitative changes in the β-globin chain. These changes lead to instability of the generated haemoglobin or to globin chain imbalance, which in turn impact the oxidative environment both intracellularly and extracellularly. The ensuing oxidative stress and the inability of the body to adequately overcome it are, to a large extent, responsible for the pathophysiology of these diseases. This article provides an overview of the main players and control mechanisms involved in the establishment of oxidative stress in these haemoglobinopathies. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.
... These conditions are accompanied by an increase in free radicals, which can damage lipids and DNA both directly and indirectly and concur to promote oxidative stress and to amplify the inflammatory process [53] (Figure 1). A condition of oxidative stress or of altered redox system is established following an unbalance between the production of reactive oxygen species (ROS) and/or reactive nitrogen species (RNS) and their removal by endogenous antioxidants [54]. ROS, like superoxide radical (O 2 − ), the hydroxyl radical (HO − ), and the nonradical hydrogen peroxide (H 2 O 2 ), which normally participate to cell signalling, at high concentrations cause cell and tissue injury and damage. ...
Article
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Metabolic disorders, especially type 2 diabetes and its associated complications, represent a growing public health problem. Epidemiological findings indicate a close relationship between diabetes and many types of cancer (including breast cancer risk), which regards not only the dysmetabolic condition, but also its underlying risk factors and therapeutic interventions. This review discusses the advances in understanding of the mechanisms linking metabolic disorders and breast cancer. Among the proposed mechanisms to explain such an association, a major role is played by the dysregulated glucose metabolism, which concurs with a chronic proinflammatory condition and an associated oxidative stress to promote tumour initiation and progression. As regards the altered glucose metabolism, hyperinsulinaemia, both endogenous due to insulin-resistance and drug-induced, appears to promote tumour cell growth through the involvement of innate immune activation, platelet activation, increased reactive oxygen species, exposure to protumorigenic and proangiogenic cytokines, and increased substrate availability to neoplastic cells. In this context, understanding the relationship between metabolic disorders and cancer is becoming imperative, and an accurate analysis of these associations could be used to identify biomarkers able to predict disease risk and/or prognosis and to help in the choice of proper evidence-based diagnostic and therapeutic protocols.
... One commonality among many of the prothrombotic conditions is an increase in vascular oxidative stress, 5 which may generate excess reactive oxygen species (ROS). In vitro studies have suggested that ROS such as superoxide can have prothrombotic effects on vascular and blood cells, including enhanced platelet activation, 6 increased expression or activity of tissue factor (TF), 7 and dysregulation of anticoagulant pathways, 8 all of which may predispose to arterial and/or venous thrombosis. ...
Article
Clinical evidence suggests an association between oxidative stress and vascular disease, and in vitro studies have demonstrated that reactive oxygen species can have prothrombotic effects on vascular and blood cells. It remains unclear, however, whether elevated levels of reactive oxygen species accelerate susceptibility to experimental thrombosis in vivo. Using a murine model with genetic deficiency in superoxide dismutase-1 (SOD1), we measured susceptibility to carotid artery thrombosis in response to photochemical injury. We found that SOD1-deficient (Sod1(-/-)) mice formed stable arterial occlusions significantly faster than wild-type (Sod1(+/+)) mice (P<0.05). Sod1(-/-) mice also developed significantly larger venous thrombi than Sod1(+/+) mice after inferior vena cava ligation (P<0.05). Activation of protein C by thrombin in lung was diminished in Sod1(-/-) mice (P<0.05 versus Sod1(+/+) mice), and generation of activated protein C in response to infusion of thrombin in vivo was decreased in Sod1(-/-) mice (P<0.05 versus Sod1(+/+) mice). SOD1 deficiency had no effect on the expression of thrombomodulin, endothelial protein C receptor, or tissue factor in lung or levels of protein C in plasma. Exposure of human thrombomodulin to superoxide in vitro caused oxidation of multiple methionine residues, including critical methionine 388, and a 40% decrease in thrombomodulin-dependent activation of protein C (P<0.05). SOD and catalase protected against superoxide-induced methionine oxidation and restored protein C activation in vitro (P<0.05). SOD prevents thrombomodulin methionine oxidation, promotes protein C activation, and protects against arterial and venous thrombosis in mice. © 2015 American Heart Association, Inc.
... CF-LVADs employ a high-speed rotating impeller to draw blood from the left ventricle and to pump it to the aorta. Non-physiological high-shear stresses exist in some region within these devices [15]. Exposure of blood to elevated shear stresses can not only result in platelet activation, but also trigger apoptotic processes, including mitochondrial transmembrane potential depolarization and phosphatidylserine exposure [16]. ...
Article
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Abstract Non-surgical bleeding (NSB) is the most common clinical complication among heart failure (HF) patients supported by continuous-flow left ventricular assist devices (CF-LVADs). Understanding the role of platelet functionality contributing to NSB after CF-LVAD implantation is crucial for prevention and management of this adverse event. The aim of this study was to examine the role of intraplatelet reactive oxygen species (ROS) and platelet damage on the incidence of bleeding events after CF-LVAD implantation in HF patients. We recruited 25 HF patients implanted with CF-LVADs and 11 healthy volunteers as the control. Intraplatelet ROS generation, platelet mitochondrial damage and platelet apoptosis were quantified by flow cytometry. Among 25 patients, 8 patients developed non-surgical bleeding within one month after CF-LVAD implantation. Intraplatelet ROS, depolarized and apoptotic platelet were found to be pre-existing conditions in all baseline samples of the 25 HF patients when compared to the healthy volunteers. There was no significant difference in the levels of ROS between the non-bleeder and the bleeder groups prior to CF-LVAD implantation, although we noticed 2-fold and 1.5-fold rise in depolarized and apoptotic platelets, respectively, in the bleeder group compared to those in the non-bleeder group. Post implant levels of intraplatelet ROS, depolarized and apoptotic platelets increased and remained elevated in the bleeder group, whereas periodic decreases were noticed in the non-bleeder group, suggesting the potential role of platelet damage on bleeding incidence. ROS generation after CF-LVAD implantation positively associated with platelet apoptosis (ρ = 0.4263, p = 0.0023) and depolarized platelets (ρ = 0.4774, p = 0.0002), especially the latter. In conclusion, elevated intraplatelet ROS and platelet damage may be linked to the NSB among HF patients supported by CF-LVAD. These results provide mechanistic insights into the bleeding complication in patients with CF-LVAD support.
... 1,2 ROS serve as second messengers at physiologic concentration and, as such, they behave as intracellular signals for cell activation. 1 Among the cells, platelets represent a typical example of ROS involvement in the activation process. 3 Thus, on stimulation by common agonists, platelets produce several types of ROS such as superoxide anion (O 2 À ) or hydrogen peroxide (H 2 O 2 ), which in turn contribute to the propagation of platelet aggregation. 4 There are several enzymatic pathways that elicit the formation of ROS into the cells, including NADPH oxidase (NOx), myeloperoxidase, xanthine oxidase, and uncoupled nitric oxide synthase. ...
Article
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Background In vitro study showed that NADPH oxidase (NOx), the most important enzyme producing reactive oxygen species (ROS), plays a role in the process of platelet activation. However, it is unclear if changes in its activity affect platelet activation in vivo. Methods and Results In vivo and ex vivo experiments assessing platelet activation were investigated in healthy subjects, obese patients, and subjects with different low rates of NOx2 activity, namely X‐linked chronic granulomatous disease (X‐CGD) patients and X‐CGD carriers. We included 27 X‐CGD patients, 31 women carriers of hereditary deficiency of NOx2, 31 obese women, and 62 healthy subjects matched for sex and age. Plasma levels of soluble sCD40 L (sCD40L) and soluble P (sP)‐selectin, 2 markers of in vivo platelet activation, were reduced in X‐CGD patients (sCD40L=−55%; sP‐selectin=−51%, P<0.001) and in X‐CGD carriers (sCD40L=−41%; sP‐selectin=−57%, P<0.001) compared with respective controls. Conversely, obese women, who disclosed NOx2 upregulation, had significantly higher plasma levels of sCD40L (+47%, P<0.001) and sP‐selectin (+70%, P<0.001) compared with controls. Ex vivo study showed platelet isoprostane downexpression and enhanced platelet NO generation in both X‐CGD patients and X‐CGD carriers compared with controls; opposite findings were observed in obese patients. Correlation analysis showed that platelet NOx2 regulation was directly associated with plasma levels of sCD40L (R=0.336, P<0.001) and sP‐selectin (R=0.441; P<0.001). Conclusions The study provides the first evidence that in vivo platelet activation is significantly and directly associated with NOx2 activity. Platelet NOx2 may be a novel target for platelet activation inhibition.
... Galectin-8 pro-inflammatory activity in the endothelium mention that platelets are another source of Gal-8 during inflammation that can account for Gal-8 effects on the endothelium (Romaniuk et al. 2010). Platelets do not adhere to the endothelium under normal conditions, but they bind to activated endothelial cells under several circumstances (Ferroni et al. 2012;Jones et al. 2012). To evaluate whether extracellular Gal-8 promotes endothelial cell activation, we studied plateletbinding capacity to Gal-8-stimulated HMEC-1 cells. ...
Article
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Galectins (Gals), a family of mammalian lectins, play diverse roles under physiological and pathological conditions. Here, we analyzed the tandem-repeat Gal-8 synthesis, secretion and effects on the endothelium physiology. Gal-8M and Gal-8L isoforms were secreted under basal conditions by human microvascular endothelial cells (HMEC-1). However, expression and secretion of the Gal-8M isoform, but not Gal-8L, were increased in response to bacterial lipopolysaccharide (LPS) stimulus and returned to control values after LPS removal. Similarly, cell surface Gal-8 exposure was increased after stimulation with LPS. To evaluate Gal-8 effects on the endothelium physiology, HMEC-1 cells were incubated in the presence of recombinant Gal-8M. Pretreated HMEC-1 cells became proadhesive to human normal platelets, indicating that Gal-8 actually activates endothelial cells. This effect was specific for lectin activity as it was prevented by the simultaneous addition of lactose, but not by sucrose. Endothelial cells also increased their exposition of von Willebrand factor after Gal-8 treatment, which constitutes another feature of cell activation that could be, in turn, responsible for the observed platelet adhesion. Several pro-inflammatory molecules were abundantly produced by Gal-8 stimulated endothelial cells: CXCL1 (GRO-α), GM-CSF, IL-6 and CCL5 (RANTES), and in a lower degree CCL2 (MCP-1), CXCL3 (GRO-γ) and CXCL8 (IL-8). In agreement, Gal-8M induced nuclear factor kappa B phosphorylation. Altogether, these results not only confirm the pro-inflammatory role we have already proposed for Gal-8 in other cellular systems but also suggest that this lectin is orchestrating the interaction between leukocytes, platelets and endothelial cells. © The Author 2014. Published by Oxford University Press. All rights reserved. For permissions, please e-mail: [email protected] /* */
... At the histological level, immunoglobulins, T lymphocytes, plasma cells, and the presence of antigen-antibody complex in the atheroma plaque suggest the importance of inflammation in the formation and progression of plaque [10]. Ongoing inflammatory conditions lead to megakaryocytic proliferation and thrombocytosis [11]. Platelets are 2e4 mm in diameter. ...
Research
Full-text available
Abstract Carotid artery stenosis (CAS) is primarily caused by atherosclerotic plaque. Progressive inflammation may contribute to the rupture of an atherosclerotic plaque. The platelet-tolymphocyte ratio (PLR) is a new and simple marker that indicates inflammation. In this study, we aimed to investigate the use of the PLR to determine the severity of CAS. One hundred forty patients were chosen from among patients who underwent carotid angiography in our institution. Symptomatic patients with stenosis >50% in the carotid arteries and asymptomatic patients with stenosis >80% were diagnosed via carotid angiography as having critical stenosis. Patients were classified into two groups. Group 1 included patients who had critical CAS, whereas Group 2 included patients with noncritical CAS, as determined by carotid angiography. Correlations between the PLR and the severity of CAS were analyzed. There were no significant differences in sex and age between the two groups. The PLR was 162.5 � 84.7 in the noncritical CAS group patients and 94.9 � 60.3 in the critical CAS group patients (p < 0.0001). The PLR value of 117.1 had 89% sensitivity and 68% specificity for CAS [95% confidence interval, 0.043 e0.159; area under the curve, 0.101 � 0.03)]. In this study, we have shown that PLR values may be associated with critical stenosis in at least one of the carotid arteries. Furthermore, PLR values may be used to predict critical stenosis in the carotid arteries.
... As reported in the previous study, both TAVR and SAVR can induce systemic oxidative stress, although the former is associated with a significantly lower redox imbalance and faster recovery of antioxidant capacity [19]. As established experimentally and through observational studies, increased levels of endo-and exogenous reactive oxygen species (ROS) are important factors triggering platelet activation [20,21,22]. It is therefore important to understand whether there is an association between platelet-related oxidative stress and platelet function following TAVR and SAVR procedures. ...
Article
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Background: Intervention-induced platelet hypercoagulability may pose a risk of serious adverse events for patients. Aims: The aim of this study was to assess whether surgical (SAVR) and transcatheter aortic valve replacement (TAVR) differ in periprocedural platelet activity. Methods: The total number of 24 patients with a mean age (SD) of 71 (13) years who underwent SAVR (n=12) or TAVR (n=12) were recruited for the study. The following parameters were evaluated at four time-points: (i) platelet indices: total platelet count (PLT), platelet distribution width (PDW) and mean platelet volume (MPV), (ii) MPV/PLT ratio, (iii) platelet level of lipid peroxidation: malondialdehyde (MDA) content and MDA/PLT ratio. Eventually, percentage variations of PLT, PDW and MPV in relation to the baseline values were determined. Results: MPV/PLT ratio increased significantly after procedures in both groups (P = 0.01 in TAVI and P = 0.01 in SAVR). MDA concentrations were significantly higher when assessed directly post-procedure (P = 0.04) as well as 24h later (P = 0.01) in the SAVR and TAVI groups. The indirect parameter of platelet activity indexed for platelet counts (MDA/PLT) was comparable between both groups before and 48 hours after procedures, but was significantly higher in SAVR patients, particularly after 24h after interventions (P = 0.04; medians TAVR vs. SAVR, respectively). Conclusion: Standard surgical aortic valve replacement is associated with a more pronounced platelet reaction to intervention-induced injury, as compared to the transcatheter-based procedure. The importance of these laboratory findings requires further investigation focused on early and late clinical outcomes.
... AOPPs trigger reactive oxygen species (ROS) production in platelets. Activated platelets generate ROS, which promote a pro-thrombotic state 26 . Notably, exposure of platelets to AOPPs (100 μ g/mL) increased ROS/superoxide production to a similar extend when compared with collagen (10 μ g/ml), whereas control albumin showed no effect (Fig. 3a). ...
Article
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Plasma advanced oxidation protein products (AOPPs), a class of pro-inflammatory pathogenic mediators, accumulate in subjects with chronic kidney disease. Whether AOPPs contribute to coagulation abnormalities, which are frequently seen in uremic patients, is unknown. Here we report that AOPPs activate platelets via a CD36-mediated signaling pathway. Activation of signaling pathways by AOPP-platelet interaction resulted in the expression of several platelet activation markers and rapidly induced the expression of CD40 ligand, triggering platelet adhesion to endothelial cells and promoting endothelial tissue factor expression. AOPPs and serum tissue factor levels were considerably increased in end stage renal disease patients on hemodialysis and a significant correlation of AOPPs and serum tissue factor was found. Interestingly, serum levels of AOPPs and tissue factor were substantially lower in stable kidney transplant patients when compared with hemodialysis patients. Given that CD36 is known to transduce the effects of oxidized lipids into platelet hyperactivity, our findings reveal previously unknown pro-thrombotic activities of oxidized plasma albumin via a CD36 dependent pathway.
... In fact, O 2 can also modulate platelet function by interfering with NO and leading to the formation of ONOO -, a relevant radical that promotes lipid and protein oxidation and reduces NO bioavailability [132,133]. ONOOcan lead to the production of 8-iso-PGF2a from AA [27], whereas 8iso-PGF2a may increase Ca 2+ release from intraplatelet stores, induce platelet shape change, and promote platelet aggregation in response to agonists [134]. Furthermore, ONOOis able to oxidize the prosthetic haem group of sGC to its NO insensitive Fe 3+ state [135][136][137] which lead to platelet hyperactivation. ...
Article
Enhanced platelet activation and thrombosis are linked to various cardiovascular diseases. Among other mechanisms, oxidative stress seems to play a pivotal role in platelet hyperactivity. Indeed, upon stimulation by physiological agonists, human platelets generate and release several types of reactive oxygen species (ROS) such as O2-, H2O2 or OH- , further amplifying the platelet activation response via various signalling pathways, including, formation of isoprostanes, Ca2+ mobilization and NO inactivation. Furthermore, excessive platelet ROS generation, incorporation of free radicals from environment and/or depletion of antioxidants induce pro-oxidant, pro-inflammatory and platelet hyperaggregability effects, leading to the incidence of cardiovascular events. Here, we review the current knowledge regarding the effect of oxidative stress on platelet signaling pathways and its implication in CVD such as type 2 diabetes mellitus. We also summarize the role of natural antioxidants included in vegetables, fruits and medicinal herbs in reducing platelet function via an oxidative stress-mediated mechanism.
... The role of platelets in thrombosis is essential, and increasingly becoming well-understood. Given the complex content within platelets, researchers have recently begun to investigate platelet function beyond coagulation, and have implicated platelets in several processes including immunoregulation, infection, inflammation, and the pathogenesis of a growing list of diseases (neurodegenerative diseases, cardiovascular disease and cancer) (24)(25)(26). In the absence of a nucleus, the role of the platelet mitochondria in these processes has become a focus of intense studies, including how platelet dysfunction is associated with, contribute to, is affected by the disease pathologies (25). ...
Article
Full-text available
Platelets are abundant, small, anucleate circulating cells, serving many emerging pathophysiological roles beyond hemostasis; including active critical roles in thrombosis, injury response, and immunoregulation. In the absence of genomic DNA transcriptional regulation (no nucleus), platelets require strategic prepackaging of all the needed RNA and organelles from megakaryocytes, to sense stress (e.g., hyperglycemia), to protect themselves from stress (e.g., mitophagy), and to communicate a stress response to other cells (e.g., granule and microparticle release). Distinct from avian thrombocytes that have a nucleus, the absence of a nucleus allows the mammalian platelet to maintain its small size, permits morphological flexibility, and may improve speed and efficiency of protein expression in response to stress. In the absence of a nucleus, platelet lifespan of 7–10 days, is largely determined by the mitochondria. The packaging of 5–8 mitochondria is critical in aerobic respiration and yielding metabolic substrates needed for function and survival. Mitochondria damage or dysfunction, as observed with several disease processes, results in greatly attenuated platelet survival and increased risk for thrombovascular events. Here we provide insights into the emerging roles of platelets despite the lack of a nucleus, and the key role played by mitochondria in platelet function and survival both in health and disease.
... Reactive oxygen species (ROS) may participate in the regulation of platelet activation; therefore, factors involved in redox regulation can be tested for their antiplatelet effects (Ferroni et al., 2012). Examples include the oxygenase platelet-type 12-(S)-lipoxygenase (12-LOX) which has been shown to potentiate platelet activation and the selective 12-LOX inhibitor, ML355, which decreases thrombosis without prolonging hemostasis (Luci et al., 2010). ...
Article
Full-text available
Antiplatelet drugs serve as a first-line antithrombotic therapy for the management of acute ischemic events and the prevention of secondary complications in vascular diseases. Numerous antiplatelet therapies have been developed; however, currently available agents are still associated with inadequate efficacy, risk of bleeding, and variability in individual response. Understanding the mechanisms of platelet involvement in thrombosis and the clinical development process of antiplatelet agents is critical for the discovery of novel agents. The functions of platelets in thrombosis are regulated by two major mechanisms: the interaction between surface receptors and their ligands, and the downstream intracellular signaling pathways. Recently, most of the progress made in antiplatelet drug development has been achieved with P2Y receptor antagonists. Additionally, the usage of GP IIb/IIIa receptor antagonists has decreased, because it is associated with a higher risk of bleeding and thrombocytopenia. Agents targeting other platelet surface receptors such as PARs, TP receptor, EP3 receptor, GPIb-IX-V receptor, P-selectin, as well as intracellular signaling factors, such as PI3Kβ, have been evaluated in an attempt to develop the next generation of antiplatelet drugs, reduce or eliminate interpatient variability of drug efficacy and significantly lower the risk of drug-induced bleeding. The aim of this review is to describe the pathways of platelet activation in thrombosis, and summarize the development process of antiplatelet agents, as well as the preclinical and clinical evaluations performed on these agents.
... They also exhibit an irregular shape and function, primarily to maintain vascular integrity and prevent bleeding after vascular injury. In-depth studies of the physiological functions of platelets have revealed their key roles in human health and disease [1]. A study by Xu et al. [2] showed that activated platelets directly promoted the occurrence of reperfusion injury. ...
Article
Full-text available
Background: Platelet activation occurs upon ischemia/reperfusion and is related to the generation of reactive oxygen species (ROS) during this process. Vitamin C (VC) is a powerful antioxidant. VC scavenges ROS, reduces platelet activation, and attenuates reperfusion injury. However, the effects of VC on platelets undergoing hypoxia/reoxygenation (H/R) remain unclear. Objectives: Herein, we evaluated the effects of VC on platelets in vitro following H/R and the related mechanisms. Method: Fresh platelets were collected from 67 volunteers at the Blood Center of Hebei Province. Platelets were diluted with saline to a concentration of 2.00 × 10<sup>11</sup>/L. Aggregation and the curve slope were evaluated within 4 h with a whole-blood impedance analyzer. To determine the optimal experimental time, platelets were treated with hypoxia or reoxygenation for different times, and impedance aggregometry was carried out by measuring changes in electrical impedance induced by arachidonic acid (0.5 mM) and adenosine diphosphate (10 µM), thereby establishing the H/R model. Three antioxidants (VC, melatonin, and probucol) were used to treat platelets after H/R, and impedance aggregometry was used to determine their effects on platelet aggregation. The influence of VC on apoptosis-related indicators was detected. ROS and the mitochondrial membrane potential were observed by inverted fluorescence microscopy and flow cytometry, respectively. Related protein levels were detected by Western blotting. Results: ROS scavengers inhibited platelet activation and aggregation in a concentration-dependent manner. VC post-conditioning scavenged ROS, downregulated cytochrome C, Bax, and caspase-9 proteins, and upregulated Bcl-2 protein. These effects collectively blocked platelet apoptosis and inhibited platelet aggregation. Conclusions: VC inhibited platelet aggregation by blocking apoptosis. Thus, VC may have applications in the treatment of platelet-related diseases.
... In addition to canonical signalling pathways depending on protein kinase activity (2), platelets are regulated in a redox-dependent manner. Several lines of evidence suggest that platelets are modulated by extracellular reactive oxygen species (ROS) (3) and that platelet activation is essentially dependent on the generation of endogenous ROS (4)(5)(6). Therefore, the study of platelet regulation and haemostasis is shedding light on the interface between ROS biochemistry and cellular physiology. Superoxide anion (O 2 •− ) from exogenous sources or endogenously produced by platelets is shown to significantly increase platelet aggregation and thrombus formation (7). ...
Article
Full-text available
The regulation of platelets by oxidants is critical for vascular health and may explain thrombotic complications in diseases such as diabetes and dementia, but remains poorly understood. Here, we describe a novel technique combining electron paramagnetic resonance spectroscopy and turbidimetry, which has been utilized to monitor simultaneously platelet activation and oxygen radical generation. This technique has been used to investigate the redox-dependence of human and mouse platelets. Using selective peptide inhibitors of NADPH oxidases (NOXs) on human platelets and genetically modified mouse platelets (NOX1-/- or NOX2-/-), we discovered that: 1) intracellular but not extracellular superoxide anion generated by NOX is critical for platelet activation by collagen; 2) superoxide dismutation to hydrogen peroxide is required for thrombin-dependent activation; 3) NOX1 is the main source of oxygen radicals in response to collagen, while NOX2 is critical for activation by thrombin; 4) two platelet modulators, namely oxidized low density lipoproteins (oxLDL) and amyloid peptide β (Aβ), require activation of both NOX1 and NOX2 to pre-activate platelets. This study provides new insights into the redox dependence of platelet activation. It suggests the possibility of selectively inhibiting platelet agonists by targeting either NOX1 (for collagen) or NOX2 (for thrombin). Selective inhibition of either NOX1 or NOX2 impairs the potentiatory effect of tested platelet modulators (oxLDL and Aβ), but does not completely abolish platelet hemostatic function. This information offers new opportunities for the development of disease-specific antiplatelet drugs with limited bleeding side effects by selectively targeting one NOX isoenzyme.
... Increased inflammation results in megakaryocyte proliferation, which leads to relative thrombocytosis (20). High cytokine levels and increased inflammation may depress the myocardium and cause cardiac dysrhythmia and myocardial remodeling (21). ...
Article
Background: Acute heart failure is a heterogenous syndrome defined by a number of factors, such as its physiopathology, clinical picture, time of onset, and relation to acute coronary syndrome. Acute cardiogenic pulmonary edema (ACPE) constitutes approximately 10-20% of acute heart failure syndromes, and it is the most dramatic symptom of left heart failure. Platelet to lymphocyte ratio (PLR) is a relatively novel inflammatory marker that can be utilized for prognosis in various disease processes. Objective: In this study, we investigated the value of the PLR for the prediction of mortality in patients with ACPE. Methods: A total of 115 patients hospitalized with a diagnosis of ACPE were included in this study. The patients were divided into tertile groups according to their PLR values: high (PLR > 194.97), medium (98.3-194.97), and low tertile (PLR < 98.3). Results: We compared the PLR groups for in-hospital mortality and total mortality after discharge. Multivariate Cox regression analysis showed that PLR was independently associated with total mortality (hazard ratio 5.657; 95% confidence interval 2.467-12.969; p < 0.001). Survival analysis using the Kaplan-Meier curve showed that the high-PLR group had a significantly higher mortality rate than the other groups. Conclusions: We showed an association between high PLR and mortality in patients with ACPE. PLR, together with other inflammatory markers and clinical findings, may be used as an adjunctive parameter for the stratification of mortality risk, hospitalization, or discharge criteria scoring.
... Ultimately, PLTs spread into the subendothelial matrix to form a solid adhesion, complete PLT aggregation and clot retraction. [3]. ...
... Ferroni P et al. reporting mean platelet volume canbe used as a simple economical test in the monitoring of DM and thereby help to decrease the morbidity and mortality. Sustained hyper glycemia leads to a series of inter related alterations that can cause evident endothelial dysfunction and vascular lesions in diabetic complications [12]. Formation of advanced glycation end products and activation of protein kinase C are the possible mechanisms by which increased glucose induces vascular abnormalities. ...
... GSH, glutathione; GSGG, glutathione oxidized; ROS, reactive oxygen species. [54,115]. The amount of F 2 -isoprostanes is reflected by the urinary excretion of a PGF 2 isomer, 8-iso PGF 2 , a reliable marker of oxidative stress in vivo [36]. ...
Article
Background: Oxidative stress is involved in different pathophysiological states, such as aging, inflammatory, cardiovascular and neurodegenerative diseases, by damaging several cellular and tissue components including proteins, DNA and lipids. On the other hand, free radicals generated during physical activity are important modulators of muscle contraction, antioxidant protection, and oxidative damage repair. Indeed, ROS, generated during physical activity, are likely main mediators of antioxidant molecules upregulation, as reflected by increased glutathione reductase levels after exercise training. Methods: The aim of this review is to summarize the main mechanisms responsible for ROS-dependent adaptations to exercise training. Results: Regular moderate exercise seems to counteract oxidative stress-related detrimental changes and to promote a healthy lifestyle. Conversely, acute and strenuous exercise can generate an excess of free radicals production. Moreover, regular habitual physical activity is related to reduced risk of coronary heart disease and death, whereas vigorous exercise has been shown to favoursudden cardiac death in sedentary individuals with preexisting vascular disease. New specific markers of mitochondrial or ER dysfunction may be better clues of oxidative stress, and their application to clinical practice may help set up the optimal dose, intensity and modality of exercise training for every single subject. Conclusion: The relationship between exercise and oxidative stress is extremely complex, depending on the mode, intensity, and duration of exercise. These conflicting effects and outcomes may be explained by the hormesis theory, in which low doses of an agent that is detrimental at high doses, induces an adaptive beneficial effect on the cells or organism.
... The thrombin burst localizes fibrin formation to the platelet aggregate, thereby stabilizing the clot. Platelet aggregation and procoagulant activity combine to form a thrombus [2,3]. ...
Chapter
Flow cytometry is a powerful tool for rapid evaluation of multiple functional properties of large numbers of platelets in whole blood. In the following chapter, we provide a number of flow cytometry-based protocols broadly aimed at (1) assessment of constitutively expressed platelet membrane receptors to diagnose inherited platelet bleeding disorders and (2) investigation of basal and agonist-induced platelet functional responses including generation of platelet-leukocyte aggregates, alpha and dense granule release, calcium flux, and phosphatidylserine exposure.
... As demonstrated in recent years using both, in vitro and in vivo experimental models, important pathways through which platelet activation can be triggered involve endo-and exogenous reactive oxygen species (ROS) [27][28][29]. Moreover, in various cardiovascular disorders, augmented interactions of ROS with platelets and the release of ROS from platelets, have been observed [30][31][32]. ...
Article
Altered function of platelets can lead to cardiovascular complications in numerous disorders. Various studies aimed to investigate mechanisms triggering platelets activation cascade show a significant role of reactive oxygen species (ROS) in this matter. Moreover, ROS are known causal factor of oxidative stress that can result in DNA, lipid and protein damage. This review aims to comprehensively present the variety of methods that are potentially useful in assessment of platelets redox balance, such as intracellular concentration of particular ROS, activity of antioxidant enzymes, reduced/oxidized glutathione ratio, level of lipid peroxidation, Cu/Zn ratio, and molecular oxygen consumption. They may help to establish the platelet-related etiological factors in different disorders and to evaluate the antiplatelet therapies. The advantages and limitations of these methods are also discussed. The present paper highlights that clinicians may benefit from implementation of such tools and further encourages developing interdisciplinary evidence-based practice.
... AOPPs trigger reactive oxygen species (ROS) production in platelets. Activated platelets generate ROS, which promote a pro-thrombotic state 26 . Notably, exposure of platelets to AOPPs (100 μ g/mL) increased ROS/superoxide production to a similar extend when compared with collagen (10 μ g/ml), whereas control albumin showed no effect (Fig. 3a). ...
... Ongoing inflammatory conditions lead to increased proliferation in the megakaryocytic series and relative thrombocytosis (19). Previous studies demonstrated the relationship of adverse cardiovascular outcomes with both increased platelet and decreased lymphocyte counts (20)(21)(22). ...
Chapter
Evaluation of platelet function is important for understanding the physiology of hemostasis and thrombosis and is utilized in clinical practice to diagnose inherited and acquired platelet bleeding disorders. Flow cytometry is a powerful tool for rapid evaluation of multiple functional properties of large number of platelets in whole blood and offers many advantages over other traditional methods. Attention to pre-analytical factors is required to ensure biologically valid and robust results.
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Objective Using 3KO (triple NOX [NADPH oxidase] knockout) mice (ie, NOX1 −/− /NOX2 −/− /NOX4 −/− ), we aimed to clarify the role of this family of enzymes in the regulation of platelets in vitro and hemostasis in vivo. Approach and Results 3KO mice displayed significantly reduced platelet superoxide radical generation, which was associated with impaired platelet aggregation, adhesion, and thrombus formation in response to the key agonists collagen and thrombin. A comparison with single-gene knockouts suggested that the phenotype of 3KO platelets is the combination of the effects of the genetic deletion of NOX1 and NOX2, while NOX4 does not show any significant function in platelet regulation. 3KO platelets displayed significantly higher levels of cGMP—a negative platelet regulator that activates PKG (protein kinase G). The inhibition of PKG substantially but only partially rescued the defective phenotype of 3KO platelets, which are responsive to both collagen and thrombin in the presence of the PKG inhibitors KT5823 or Rp-8-pCPT-cGMPs, but not in the presence of the NOS (NO synthase) inhibitor L-NG-monomethyl arginine. In vivo, triple NOX deficiency protected against ferric chloride–driven carotid artery thrombosis and experimental pulmonary embolism, while hemostasis tested in a tail-tip transection assay was not affected. Procoagulatory activity of platelets (ie, phosphatidylserine surface exposure) and the coagulation cascade in platelet-free plasma were normal. Conclusions This study indicates that inhibiting NOXs has strong antithrombotic effects partially caused by increased intracellular cGMP but spares hemostasis. NOXs are, therefore, pharmacotherapeutic targets to develop new antithrombotic drugs without bleeding side effects.
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CalDAG-GEFI is a guanine nucleotide exchange factor, which actives small GTPase Rap1 and plays an important role in platelet aggregation. Our previous study has shown that CalDAG-GEFI contains redox-sensitive thiols, and its function can be inhibited by thiol modification. In the present study, the effect of CLL2-1, a 1,4-phenanthrenequinone, on CalDAG-GEFI and platelet functions was investigated. In human platelets, CLL2-1 prevented platelet aggregation caused by various stimulators. Flow cytometric analysis revealed that CLL2-1 inhibited GPIIb/IIIa activation and P-selectin secretion. Moreover, CLL2-1 prevented Rap1 activation caused by thrombin, the Ca(2+) ionophore A23187, and the diacylglycerol mimetic phorbol 12-myristate 13-acetate, while only slightly inhibited thrombin-induced increases in [Ca(2+)]i and did not inhibit protein kinase C activation. Western blots after reducing SDS-PAGE showed that treatment of either platelets or platelet lysates with CLL2-1 led to a decrease of monomeric CalDAG-GEFI and appearance of cross-linked oligomers of CalDAG-GEFI, and these effects were inhibited by pretreatment of platelets or lysates with thiol reducing agents prior to the addition of CLL2-1, indicating thiol modification of CalDAG-GEFI by CLL2-1. Furthermore, the thiol reducing agents also prevented the inhibitory effect of CLL2-1 on Rap1 activation, GPIIb/IIIa activation, and platelet aggregation. In CalDAG-GEFI-overexpressing human embryonic kidney 293T cells, CLL2-1 also inhibited CalDAG-GEFI-mediated Rap1 activation. Taken together, our results suggest that the antiplatelet effect of CLL2-1 is due to, at least in part, inhibition of CalDAG-GEFI-mediated Rap1 activation, and provide the basis for development of novel antiplatelet drugs. Copyright © 2014 Elsevier Inc. All rights reserved.
Chapter
Antithrombotic drugs (also known as antiplatelet drugs or antiplatelet agents) are used for the prevention of platelet-associated thrombotic complications including myocardial infarction and stroke. Clinical complications associated with the use of currently available antithrombotic drugs warrant the development of safer and more effective therapies. A better understanding of the molecular signaling mechanisms involved in platelet activation and a better appreciation of the pharmacologic and pathophysiologic variables that may alter the anticipated clinical outcomes in given patients are essential. This article provides an overview of the signaling mechanisms involved in platelet activation and discusses the available antiplatelet agents with a focus on their mechanisms of action, complications associated with, and limitations on their use. In addition, the platelet function tests and the experimental animal models of thrombosis that may be used for testing the antiplatelet agents are described.
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Cerebral malaria (CM), defined as the presence of P. falciparum asexual stages on peripheral blood smear in a person with coma and no other cause for encephalopathy, is estimated to affect more than 800,000 people a year and has a 15–20 % mortality rate. CM predominantly affects children RANTES; release of free heme during hemolysis; endothelial activation leading to blood–brain barrier breakdown; CNS nitric oxide production; and genetic polymorphisms (e.g., sickle cell trait) that alter these responses or protect in other ways from severe disease. Murine models of cerebral malaria have provided new insights into the disease, but the difference in the parasite species and the host response has limited translation of findings from murine models into human CM studies. Nonhuman primate models are closer to human disease, but are limited by cost and ethical concerns. Therapies currently being studied for adjunctive therapy in CM include arginine (a donor of nitric oxide), inhaled nitric oxide, and recombinant erythropoietin. The potential benefits and harm of each therapy require close study, as many areas of CM pathogenesis remain unclear. Further studies are required, particularly in human disease, to better understand pathogenesis so that effective adjunctive therapy for this illness can be developed.
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Nutrition plays an important role in the prevention and management of disease. Whole grain cereals contain a host of nutrients and bioactive substances that have health promoting effects. Epidemiological evidence shows a consistent inverse association between whole grain intake and the risk of chronic disease. Despite a concerted effort by scientists, educators, and policy makers, to promote the consumption of whole grains, it remains dismally short of the recommended intakes. Pulses (dried beans and peas) differ from whole grains in their structural and physicochemical properties, and have varying amounts of fiber, resistant starch, vitamins, minerals, and other bioactive components; nevertheless, these foods groups complement each other. Observational as well as intervention trials show that pulse consumption has beneficial effects on the prevention and management of chronic disease. The nutritional and phytochemical components of pulses coupled with that of whole grains suggest a potential synergistic effect that could provide unprecedented health benefits.
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Being mainly known for their role in the antimicrobial defense and collateral damage they cause in tissues as agents of oxidative stress, reactive oxygen species were considered “the bad guys” for decades. However, in the last years it was shown that the absence of reactive oxygen species can lead to the development of immune-mediated inflammatory diseases. Animal models of lupus, arthritis and psoriasis revealed reactive oxygen species-deficiency as a potent driver of pathogenesis. On the contrary, in chronic stages oxidative stress can still contribute to progression of inflammation. It seems that a neatly adjusted redox balance is necessary to sustain an immune state that both prevents the development of overt autoimmunity and attenuates chronic stages of disease.
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Kupffer cells are liver resident macrophages that are responsible for screening and clearing blood of pathogens and foreign particles. It has recently been shown that Kupffer cells interact with platelets, through an adhesion based mechanism, to aid in pathogen clearance and then these platelets re-enter the general systemic circulation. Thus, a mechanism has been identified that relates liver inflammation to possible changes in the systemic circulation. However, the role that Kupffer cells play in cardiovascular disease initiation/progression has not been elucidated. Thus, our objective was to determine whether or not Kupffer cells are responsive to a classical cardiovascular risk factor and if these changes can be transmitted into the general systemic circulation. If Kupffer cells initiate inflammatory responses after exposure to classical cardiovascular risk factors, then this provides a potential alternative/synergistic pathway for cardiovascular disease initiation. We aimed to elucidate the prevalence of this potential pathway. We hypothesized that Kupffer cells would initiate a robust inflammatory response after exposure to tobacco cigarette or e-cigarette products and that the inflammatory response would have the potential to antagonize other salient cells for cardiovascular disease progression. To test this, Kupffer cells were incubated with tobacco smoke extracts, e-cigarette vapor extracts or pure nicotine. Complement deposition onto Kupffer cells, Kupffer cell complement receptor expression, oxidative stress production, cytokine release and viability and density were assessed after the exposure. We observed a robust inflammatory response, oxidative stress production and cytokine release after Kupffer cells were exposed to tobacco or e-cigarette extracts. We also observed a marginal decrease in cell viability coupled with a significant decrease in cell density. In general, this was not a function of the extract formulation (e.g. tobacco vs. e-cigarette products or the formulation of the cigarette product). These results indicate that Kupffer cells are responsive to classical cardiovascular risk factors and that an inflammatory response is initiated that may pass into the general systemic circulation. Copyright © 2015 Elsevier Ltd. All rights reserved.
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Studies have suggested a pivotal role for free sulfhydryls in platelet integrin function, and enzyme-mediated reduction of disulfide bonds on platelets has been implicated. The platelet fibrinogen receptor αIIbβ3 is the best-studied platelet integrin and serves as a model system for studying the structure-function relation in this family of adhesion receptors. The demonstration of free sulfhydryls on the exofacial domain of purified αIIbβ3, specifically in its activated conformation, prompted us to explore the potential for activation-dependent, enzymatically catalyzed thiol expression on intact platelets and the possible role of surface-associated protein disulfide isomerase (PDI) in αIIbβ3 ligation. Using the membrane-impermeant sulfhydryl blocker para-chloromercuriphenyl sulfonate, the inhibitor of disulfide exchange bacitracin, and the monoclonal anti-PDI antibody RL90, we examined fibrinogen binding to αIIbβ3 as well as ligation-induced allosteric changes in the conformation of αIIbβ3. We sought to distinguish the possible involvement of disulfide exchange in agonist-induced platelet stimulation from its role in integrin ligation. Analysis of the role of free thiols in platelet aggregation suggested a thiol-independent initial ligation followed by a thiol-dependent stabilization of binding. Flow cytometric analysis showed that sustained binding of fibrinogen, as well as expression of ligand-induced binding site epitopes and ligand-bound conformation, depended on free thiols and disulfide exchange. Expression of P-selectin was minimally affected, even with complete inhibition of αIIbβ3function. These data indicate that although agonist-induced platelet stimulation is independent of ecto-sulfhydryls, engagement of integrin αIIbβ3 on the intact platelet depends totally on their enzymatically catalyzed surface expression.
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Diabetes mellitus is associated with aggravated development of vascular complications. Yet, it has not been established whether platelet hyperreactivity contributes as a pathogenetic factor. In order to study the role of activated platelets in diabetes mellitus, we investigated the expression of the membrane activation markers CD63 (GP53) and CD62 (GMP-140) as direct indicators of in vivo activation. The CD63-positive fraction was significantly higher in patients (6.1% X 3.7 +/- 1) than in controls (2.7% X 3 +/- 1). In parallel, the CD62-positive fraction was significantly elevated in patients to 5% X 2.5 +/- 1 in comparison to controls (3% X 2 +/- 1). Patients with angiopathy had a mean increase of 304% in CD63-positive and of 223% in CD62-positive platelets. Patients without clinically detectable angiopathy showed a trend to an increased fraction in CD63-/CD62-positive platelets. There was no correlation of the activation markers with fasting blood glucose, HbA1 or platelet count. CD63 platelet bound fluorescence significantly increased with platelet size in the patient group. We conclude that in diabetes mellitus an increased number of large platelets circulate in an activated state predominantly in patients with angiopathy. This could imply that platelets become activated by vascular lesions. The trend in patients without vascular disease, however, suggests that activated platelets may also basically contribute to the prethrombotic state in diabetes mellitus.
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Ticagrelor is an oral, reversible, direct-acting inhibitor of the adenosine diphosphate receptor P2Y12 that has a more rapid onset and more pronounced platelet inhibition than clopidogrel. In this multicenter, double-blind, randomized trial, we compared ticagrelor (180-mg loading dose, 90 mg twice daily thereafter) and clopidogrel (300-to-600-mg loading dose, 75 mg daily thereafter) for the prevention of cardiovascular events in 18,624 patients admitted to the hospital with an acute coronary syndrome, with or without ST-segment elevation. At 12 months, the primary end point--a composite of death from vascular causes, myocardial infarction, or stroke--had occurred in 9.8% of patients receiving ticagrelor as compared with 11.7% of those receiving clopidogrel (hazard ratio, 0.84; 95% confidence interval [CI], 0.77 to 0.92; P<0.001). Predefined hierarchical testing of secondary end points showed significant differences in the rates of other composite end points, as well as myocardial infarction alone (5.8% in the ticagrelor group vs. 6.9% in the clopidogrel group, P=0.005) and death from vascular causes (4.0% vs. 5.1%, P=0.001) but not stroke alone (1.5% vs. 1.3%, P=0.22). The rate of death from any cause was also reduced with ticagrelor (4.5%, vs. 5.9% with clopidogrel; P<0.001). No significant difference in the rates of major bleeding was found between the ticagrelor and clopidogrel groups (11.6% and 11.2%, respectively; P=0.43), but ticagrelor was associated with a higher rate of major bleeding not related to coronary-artery bypass grafting (4.5% vs. 3.8%, P=0.03), including more instances of fatal intracranial bleeding and fewer of fatal bleeding of other types. In patients who have an acute coronary syndrome with or without ST-segment elevation, treatment with ticagrelor as compared with clopidogrel significantly reduced the rate of death from vascular causes, myocardial infarction, or stroke without an increase in the rate of overall major bleeding but with an increase in the rate of non-procedure-related bleeding. (ClinicalTrials.gov number, NCT00391872.)
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Aggregation of human washed platelets with collagen is accompanied by a concentration-dependent increase in cyclic GMP but not cyclic AMP. NG-Monomethyl-L-arginine (L-MeArg), a selective inhibitor of nitric oxide (NO) synthesis from L-arginine, reduces this increase and enhances aggregation. L-Arginine, which has no effect on the basal levels of cyclic GMP, augments the increase in this nucleotide induced by collagen and also inhibits aggregation. Both of these effects of L-arginine are attenuated by L-MeArg. The anti-aggregatory action of L-arginine is potentiated by prostacyclin and by M&B22948, a selective inhibitor of the cyclic GMP phosphodiesterase, but not by HL725, a selective inhibitor of the cyclic AMP phosphodiesterase. L-Arginine also inhibits platelet aggregation in whole blood in a similar manner, although the concentrations required are considerably higher. L-Arginine stimulates the soluble guanylate cyclase and increases cyclic GMP in platelet cytosol. This stimulation is dependent on NADPH and Ca2+ and is associated with the formation of NO. Both the formation of NO and the stimulation of the soluble guanylate cyclase induced by L-arginine are enantiomer specific and abolished by L-MeArg. Thus, human platelets contain an NO synthase which is activated when platelets are stimulated. The consequent generation of NO modulates platelet reactivity by increasing cyclic GMP. Changes in the activity of this pathway in platelets may have physiological, pathophysiological, and therapeutic significance.
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Dyslipidemia is associated with a prothrombotic phenotype; however, the mechanisms responsible for enhanced platelet reactivity remain unclear. Proatherosclerotic lipid abnormalities are associated with both enhanced oxidant stress and the generation of biologically active oxidized lipids, including potential ligands for the scavenger receptor CD36, a major platelet glycoprotein. Using multiple mouse in vivo thrombosis models, we now demonstrate that genetic deletion of Cd36 protects mice from hyperlipidemia-associated enhanced platelet reactivity and the accompanying prothrombotic phenotype. Structurally defined oxidized choline glycerophospholipids that serve as high-affinity ligands for CD36 were at markedly increased levels in the plasma of hyperlipidemic mice and in the plasma of humans with low HDL levels, were able to bind platelets via CD36 and, at pathophysiological levels, promoted platelet activation via CD36. Thus, interactions of platelet CD36 with specific endogenous oxidized lipids play a crucial role in the well-known clinical associations between dyslipidemia, oxidant stress and a prothrombotic phenotype.
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Tyrosyl radicals have been detected during turnover of prostaglandin endoperoxide H synthase (PGHS), and they are speculated to participate in cyclooxygenase catalysis. Spectroscopic approaches to elucidate the identity of the radicals have not been definitive, so we have attempted to trap the radical(s) with nitric oxide (NO). NO quenched the EPR signal generated by reaction of purified ram seminal vesicle PGHS with arachidonic acid, suggesting that NO coupled with a tyrosyl radical to form inter alianitrosocyclohexadienone. Subsequent formation of nitrotyrosine was detected by Western blotting of PGHS incubated with NO and arachidonic acid or organic hydroperoxides using an antibody against nitrotyrosine. Both arachidonic acid and NO were required to form nitrotyrosine, and tyrosine nitration was blocked by the PGHS inhibitor indomethacin. The presence of superoxide dismutase had no effect on nitration, indicating that peroxynitrite was not the nitrating agent. To identify which tyrosines were nitrated, PGHS was digested with trypsin, and the resulting peptides were separated by high pressure liquid chromatography and monitored with a diode array detector. A single peptide was detected that exhibited a spectrum consistent with the presence of nitrotyrosine. Consistent with Western blotting results, both NO and arachidonic acid were required to observe nitration of this peptide, and its formation was blocked by the PGHS inhibitor indomethacin. Peptide sequencing indicated that the modified residue was tyrosine 385, the source of the putative catalytically active tyrosyl radical.
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C.Brakebusch and W.Bergmeier contributed equally to this work Platelet adhesion on and activation by components of the extracellular matrix are crucial to arrest post-traumatic bleeding, but can also harm tissue by occluding diseased vessels. Integrin a2b1 is thought to be essential for platelet adhesion to subendothelial collagens, facilitating subsequent interactions with the activating platelet collagen receptor, glycoprotein VI (GPVI). Here we show that Cre/loxP-mediated loss of b1 integrin on platelets has no signi®cant effect on the bleeding time in mice. Aggregation of b1-null platelets to native ®brillar collagen is delayed, but not reduced, whereas aggregation to enzymatically digested soluble collagen is abolished. Furthermore, b1-null platelets adhere to ®brillar, but not soluble collagen under static as well as low (150 s ±1) and high (1000 s ±1) shear ¯ow conditions, probably through binding of aIIbb3 to von Willebrand factor. On the other hand, we show that platelets lacking GPVI can not activate integrins and consequently fail to adhere to and aggregate on ®brillar as well as soluble collagen. These data show that GPVI plays the central role in platelet±collagen interactions by activating different adhesive receptors, including a2b1 integrin, which strengthens adhesion without being essential.
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Platelet activation and blood coagulation are complementary, mutually dependent processes in haemostasis and thrombosis. Platelets interact with several coagulation factors, while the coagulation product thrombin is a potent platelet-activating agonist. Activated platelets come in a procoagulant state after a prolonged elevation in cytosolic [Ca2+](i). Such platelets, e. g. when adhering to collagen via glycoprotein VI, expose phosphatidylserine (PS) at their outer surface and produce (PS-exposing) membrane blebs and microvesicles. Inhibition of aminophospholipid translocase and activation of phospholipid scramblase mediate the exposure of PS, whereas calpain-mediated protein cleavage leads to membrane blebbing and vesiculation. Surface-exposed PS strongly propagates the coagulation process by facilitating the assembly and activation of tenase and prothrombinase complexes. Factor IXa and platelet-bound factor Va support these activities. In addition, platelets can support the initiation phase of coagulation by providing binding sites for prothrombin and factor XI. They thereby take over the initiating role of tissue factor and factor VIIa in coagulation activation.
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Platelet integrins alpha2beta1 and alphaIIbbeta3 play critical roles in platelet adhesion and thrombus formation after vascular injury. On resting platelets, both integrins are in a low-affinity state. However, agonist stimulation results in conformational changes that enable ligand binding that can be detected with conformation dependent monoclonal antibodies (mAbs). By using such conformation-dependent mAbs, we could demonstrate that activation of integrin alphaIIbbeta3 is not only sufficient, but also a prerequisite for alpha2beta1 activation. Compared with platelets in plasma, stimulation of washed platelets resulted in only a minor activation of alpha2beta1, as detected with the activation-sensitive mAb IAC-1. Addition of fibrinogen to stimulated washed platelets greatly potentiated activation of this integrin. Also, treatment of alphaIIbbeta3 with the ligand-mimetic peptide RGDS, resulting in outside-in signaling, led to a powerful alpha2beta1 activation, even in the absence of overall platelet activation, involving tyrosine kinase activity but no protein kinase C activation. The absolute necessity of alphaIIbbeta3 for proper alpha2beta1 activation on platelets was demonstrated by using the alphaIIbbeta3 antagonist aggrastat, which was able to completely abolish alpha2beta1 activation, both under static and flow conditions. In addition, analogous experiments with Glanzmann platelets lacking alphaIIbbeta3 confirmed the indispensability of alphaIIbbeta3 for alpha2beta1 activation.
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Background: TRA-PCI was a multicenter, randomized, double-blind, placebo-controlled study demonstrating the safety of SCH 530348, a potent thrombin receptor antagonist (TRA), in patients undergoing non-urgent percutaneous coronary intervention (PCI). A secondary endpoint in a sub-study within the primary evaluable patient cohort was inhibition of platelet aggregation (IPA) induced by appropriate agonists relative to baseline. We hypothesized that TRA therapy would selectively inhibit platelet aggregation induced by the thrombin receptor agonist peptide (TRAP) and that sustained inhibition would be observed. Pharmacokinetic (PK) studies were also carried out. Methods: Platelet aggregation responses to TRAP, adenosine diphosphate (ADP), collagen and arachidonic acid (AA) were measured using light transmission aggregometry (LTA) at baseline, 30, 60, 90, 120 min following a loading dose (10, 20 or 40 mg vs placebo) and after a maintenance dose (0.5, 1.0 or 2.5 mg/day) at 30 and 60 days. PK was assessed at 30 and 60 min and 2 hr after loading dose administration. Results: TRA was active in inhibiting 15 μM TRAP-induced platelet aggregation with onset of inhibition directly related to dose. Loading doses of 10, 20 or 40 mg achieved >90% inhibition of platelet aggregation (IPA) 60 –120 min post dose with the 40 mg dose achieving >90% inhibition between 60 and 90 min. Patients receiving maintenance doses of 0.5, 1.0 or 2.5 mg exhibited >90% IPA at 30 and 60 days. Placebo treated patients had on average ≥10% IPA to TRAP. TRA had no significant effects on ADP, collagen or AA -induced platelet aggregation compared with placebo controls. TRA PK was characterized by fast distribution and slow elimination (t1/2 = 165–311 hr) with clear evidence of hysteresis. Conclusion: Among PCI patients treated with TRA, there was a specific, dose-related, sustained inhibition of TRAP-induced platelet aggregation, without an impact on other aggregation related pathways. These data suggest that TRA is a potent, selective inhibitor of PAR-1 activity induced by thrombin activation of platelets, and, in view of this, is a promising therapeutic utility for treatment of thrombosis.
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Platelets bound to thrombogenic surfaces have been shown to support activation-dependent firm adhesion of neutrophils in flow following selectin-mediated tethering and rolling. The specific receptor(s) responsible for mediating adhesion-strengthening interactions between neutrophils and platelets has not previously been identified. Furthermore, the ability of adherent platelets to support the migration of bound neutrophils has not been tested. We studied neutrophil interactions with activated, surface-adherent platelets as a model for leukocyte binding in vascular shear flow and emigration at thrombogenic sites. Our results demonstrate that the beta 2-integrin Mac-1 (CD11b/CD18) is required for both firm attachment to and transmigration of neutrophils across surface-adherent platelets. In flow assays, neutrophils from patients with leukocyte adhesion deficiency-1 (LAD-I), which lack beta 2-integrin receptors, formed P-selectin-mediated rolling interactions, but were unable to develop firm adhesion to activated platelets, in contrast to healthy neutrophils, which developed firm adhesion within 5 to 30 seconds after initiation of rolling. Furthermore, the adhesion-strengthening interaction observed for healthy neutrophils could be specifically inhibited by monoclonal antibodies (mAbs) to Mac-1, but not to lymphocyte function-associated antigen-1 (LFA-1; CD11a/CD18) or intercellular adhesion molecule-2 (ICAM-2; CD102). Further evidence for a beta 2-integrin-dependent neutrophil/platelet interaction is demonstrated by the complete inhibition of interleukin (IL)-8-induced neutrophil transmigration across platelets bound to fibronectin-coated polycarbonate filters by mAbs to Mac-1. Thus, Mac-1 is required for firm adhesion of neutrophils to activated, adherent platelets and may play an important role in promoting neutrophil accumulation on and migration across platelets deposited at sites of vascular injury.
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The capacity of platelets to generate thromboxane A2, reflected by measurement of serum thromboxane B2 (TxB2), greatly exceeds the systemic production of thromboxane in vivo. Thus, it is possible that substantial but incomplete inhibition of thromboxane formation ex vivo would still allow marked augmentation of thromboxane production in vivo. To address this hypothesis, we administered aspirin 120 mg, a selective inhibitor of thromboxane synthase (TxSl), 3-(1H-imidazol-1-yl- methyl)-2-methyl-1H-indole-1-propanoic acid (UK-38, 485) 200 mg, and a combination of both drugs to 12 healthy volunteers and measured the effects on serum TxB2 and urinary 2,3-dinor-thromboxane B2 (Tx-M), an index of endogenous thromboxane biosynthesis. Although serum TxB2 was maximally inhibited by 94 +/- 1% after aspirin and 96 +/- 2% after the TxSl, maximal depression of Tx-M was only 28 +/- 8% and 37 +/- 9%, respectively. Combination of aspirin with the TxSl resulted in a small but significant increase in inhibition of thromboxane generation ex vivo (98 +/- 1% v 94 +/- 1%; P less than 0.05), but a disproportionately greater fall in thromboxane synthesis in vivo (58 +/- 7%; P less than 0.01). Consistent with further inhibition of platelet thromboxane synthesis, addition of the TxSl abolished the transient decline in prostacyclin formation after aspirin alone. Administration of a lower dose of aspirin (20 mg) to 6 healthy subjects caused a small reduction in Tx-M (12 +/- 4%; P less than 0.05) and inhibited serum TxB2 by 48 +/- 2%. The relationship between inhibition of platelet capacity to form thromboxane ex vivo (serum TxB2) and synthesis in vivo (Tx-M) departed markedly from the line of identity. When total blockade of the capacity of platelets to generate thromboxane is approached, minor decrements in capacity result in a disproportionate depression of actual thromboxane biosynthesis. These results imply that pharmacologic inhibition of serum TxB2 must be virtually complete before thromboxane- dependent platelet activation is influenced in vivo.
Article
The adhesion of platelets to purified laminin under flow conditions was investigated. Adhesion to laminin was strongly dependent on the presence of divalent cations. In the absence of cations platelet adhesion (8% coverage in 5 minutes) was maximal at a shear rate of 100/s and no adhesion could be detected at shear rates above 800/s. In the presence of 0.8 mmol/L Mg2+ and 2 mmol/L Ca2+ platelet adhesion reached its maximum (30% coverage) around 800/s. At 1,800/s platelets still adhered to purified laminin (coverage of 6%). Antibodies against the E8 domain of laminin and antibodies against the alpha 6 and beta 1 chains of platelet membrane glycoprotein very late activation antigen-6 (VLA-6), completely inhibited adhesion. No inhibition was found with antibodies against glycoprotein IIb:IIIa, against the alpha 2 chain of VLA-2, and against the alpha 5 chain of VLA-5. Fibronectin and von Willebrand factor were not involved in laminin-dependent adhesion. Anti- VLA-6 partly inhibited platelet adhesion to the extracellular matrix of endothelial cells at shear rates below 800/s. Preincubation of the matrices with antilaminin E8 antibodies did not influence the adhesion. These results show that purified laminin supports platelet adhesion and that the presence of VLA-6 is important for platelet adhesion under flow conditions. The protein in the matrix with which VLA-6 interacts is currently unknown.
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Collagens are important platelet activators in the vascular subendothelium and vessel wall. Since the regulation of platelet activation is a key step in distinguishing normal haemostasis from pathological thrombosis, collagen interactions with platelets are important targets for pharmacological control. Platelets have two major receptors for collagens, the integrin α2 β1, with a major role in adhesion and platelet anchoring and the Ig superfamily member, GPVI, principally responsible for signalling and platelet activation. In addition, GPIb-V-IX, can be considered as an indirect collagen receptor acting via von Willebrand factor as bridging molecule and is essential for platelet interactions with collagen at high shear rates. There is some evidence for additional receptors, which may regulate the response to individual collagen types. This review discusses how these receptors work separately with specific agonists and proposes possible mechanisms for how they work together to regulate platelet activation by collagen, which remains controversial and poorly understood.
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Platelet function testing is essential for the diagnosis of congenital/acquired bleeding disorders and may be useful for the prediction of surgical bleeding. Nowadays there is also much interest in monitoring the efficacy of anti-platelet therapy and measuring platelet hyper-function. However, this often presents clinical laboratories with significant challenges as platelet function tests are complex, poorly standardized, time consuming and quality assurance is not straightforward. There are also few comprehensive modern guidelines available and many recent published surveys have revealed poor standardization between laboratories. Up until the late 1980’s the traditional clinical platelet function tests that were available were the bleeding time (BT), light transmission (LTA) and whole blood aggregometry (WBA) and various biochemical assays. These were also usually performed within specialized research and clinical laboratories. Since the last BCSH guidelines were published in 1988 a variety of new platelet function tests have become available. These include flow cytometry and an ever increasing choice of new commercial instruments. Although the potential clinical utility of the new assays is emerging some have not yet entered into routine clinical practice. It is encouraging that a number of standardization committees (e. g. CLSI, BCSH and ISTH Platelet Physiology SSC) are now beginning to produce new platelet function testing guidelines and this will hopefully improve clinical practice, quality assurance and result in less variability between different laboratories.
Article
Background-Diabetes mellitus (DM) is associated with enhanced lipid peroxidation and persistent platelet activation. We tested the hypothesis that the in vivo formation of the F-2-isoprostane 8-iso-prostaglandin (PG)F-2 alpha, a bioactive product of arachidonic acid peroxidation, is enhanced in DM and contributes to platelet activation. Methods and Results-Urine samples were obtained from 85 diabetic patients and 85 age- and sex-matched healthy subjects for measurement of immunoreactive 8-iso-PGF(2 alpha) and 11-dehydro-thromboxane B-2 (TXM), an in vivo index of platelet activation. Sixty-two had non-insulin-dependent (NID)DM, and 23 had insulin-dependent (ID) DM, Vitamin E supplementation, metabolic control, and cyclooxygenase inhibitors were used to investigate the mechanisms of formation of 8-iso-PGF(2 alpha) in this setting. Urinary 8-iso-PGF(2 alpha) excretion was significantly higher (P=0.0001) in NIDDM patients (419 +/- 208 pg/mg creatinine; range 160 to 1014) than in age-matched control subjects (208 +/- 92; 41 to 433), Urinary 8-iso-PGF(2 alpha) was linearly correlated with blood glucose and urinary TXM. 8-iso-PGF(2 alpha) excretion was also significantly (P=0.0001) higher in IDDM patients (400 +/- 146; 183 to 702) than in control subjects (197 +/- 69; 95 to 353), Vitamin E supplementation (600 mg/d for 14 days) was associated with a statistically significant reduction in both urinary 8-iso-PGF(2 alpha) (by 37%) and TXM (by 43%) in 10 NIDDM patients. Improved metabolic control was associated with a significant (P=0.0001) reduction in 8-iso-PGF(2 alpha) and TXM excretion by 32% and 41%, respectively, in 21 NIDDM patients. 8-iso-PGF(2 alpha) was unchanged after 2-week dosing with aspirin and indobufen despite profound suppression of TXM excretion. Conclusions-We conclude that DM is associated with increased formation of F-2-isoprostanes, as a correlate of impaired glycemic control and enhanced lipid peroxidation. This may provide an important biochemical link between impaired glycemic control and persistent platelet activation. These results provide a rationale for dos-finding studies of antioxidant treatment in diabetes.
Article
A latex agglutination test was compared with the micro-titration haemolysin inhibition method for the detection of anti-streptolysin O (ASO) antibodies in 428 serum samples. After slight modification of the latex method to produce maximal agglutination good agreement was shown between the results obtained by the two methods. The latex test had a sensitivity of 83·6%, a specificity of 93·3%, a predictive positive value of 86·5% and a predictive negative value of 91·6%. It was convenient, required less labour than the haemolysin test, and permitted economic testing of small numbers of sera.
Article
Background Low-dose aspirin is of definite and substantial net benefit for many people who already have occlusive vascular disease. We have assessed the benefits and risks in primary prevention. Methods We undertook meta-analyses of serious vascular events (myocardial infarction, stroke, or vascular death) and major bleeds in six primary prevention trials (95000 individuals at low average risk, 660000 person-years, 3554 serious vascular events) and 16 secondary prevention trials (17000 individuals at high average risk, 43 000 person-years, 3306 serious vascular events) that compared long-term aspirin versus control. We report intention-to-treat analyses of first events during the scheduled treatment period. Findings in the primary prevention trials, aspirin allocation yielded a 12% proportional reduction in serious vascular events (0.51% aspirin vs 0.57% control per year, p=0.0001), due mainly to a reduction of about a fifth in non-fatal myocardial infarction (0.18% vs 0.23% per year, p<0.0001). The net effect on stroke was not significant (0.20% vs 0.21% per year, p=0.4: haernorrhagic stroke 0.04% vs 0.03%, p=0.05; other stroke 0.16% vs 0.18% per year, p=0.08). Vascular mortality did not differ significantly (0.19% vs 0.19% per year, p=0.7). Aspirin allocation increased major gastrointestinal and extracranial bleeds (0.10% vs 0.07% per year, p<0.0001), and the main risk factors for coronary disease were also risk factors for bleeding. In the secondary prevention trials, aspirin allocation yielded a greater absolute reduction in serious vascular events (6.7% vs 8.2% per year, p<0.0001), with a non-significant increase in haernorrhagic stroke but reductions of about a fifth in total stroke (2.08% vs 2.54% per year, p=0.002) and in coronary events (4.3% vs 5.3% per year, p<0.0001). In both primary and secondary prevention trials, the proportional reductions in the aggregate of all serious vascular events seemed similar for men and women. Interpretation In primary prevention without previous disease, aspirin is of uncertain net value as the reduction in occlusive events needs to be weighed against any increase in major bleeds. Further trials are in progress. Funding UK Medical Research Council, British Heart Foundation, Cancer Research UK, and the European Community Biomed Programme.