Platelets, though anucleated, possess several transcription factors, including NF-kappaB, that exert non-genomic functions regulating platelet activation. Since platelets have not only been recognized as central players of homeostasis, but also participated in pathological conditions such as thrombosis, atherosclerosis, and inflammation, we examined rat platelet NF-kappaB expression and evaluated the effects of anti-inflammatory drug BAY 11-7082, an inhibitor of NF-kappaB activation, in platelet physiology. Western blotting revealed that rat platelets express NF-kappaB. BAY 11-7082, dose dependently, inhibited collagen- or thrombin-induced-platelet aggregation. ATP release, TXB(2) formation, P-selectin expression, and intercellular Ca(2+) concentration activated by collagen were reduced in BAY 11-7082-treated platelets. BAY 11-7082 elevated intracellular levels of cAMP, but not cGMP, and its co-incubation with cAMP-activating agent (forskolin) or its hydrolyzing enzyme inhibitor (3-isobutyl-1-methylxanthine, IBMX), synergistically inhibited collagen-induced-platelet aggregation. In addition, vasodilator-stimulated-phosphoprotein (VASP) phosphorylation was enhanced in BAY 11-7082-treated platelets, which was partially inhibited by a protein kinase A (PKA) inhibitor, H-89. Moreover, while p38 mitogen-activated protein kinase (MAPK) was not affected, BAY 11-7082 attenuated c-Jun N-terminal kinase 1 (JNK1) and extracellular-signal-regulated protein kinase 2 (ERK2) phosphorylations. In conclusion, BAY 11-7082 inhibits platelet activation, granule secretion, and aggregation, and that this effect is mediated by inhibition of JNK1 and ERK2 phosphorylations, and partially by stimulation of cAMP-dependent PKA VASP phosphorylation. The ability of BAY 11-7082 to inhibit platelet function might be relevant in cases involving aberrant platelet activation where the drug is considered as anti-atherothrombosis, and anti-inflammatory therapy.
"Mammalian platelets devoid of nucleus and basically transcriptional and translational activities are limited. Although platelet transcription factors such as nuclear factor kappa B (NF-κB) are some reported [32, 33], it is largely understood that new functional protein synthesis is limited. Therefore, granule substances take a pivotal role in the platelet activation and aggregation due to its biological and functional significance. "
[Show abstract][Hide abstract] ABSTRACT: Ethnopharmacological Relevance. Morus alba L. leaves (MAE) have been used in fork medicine for the treatment of beriberi, edema, diabetes, hypertension, and atherosclerosis. However, underlying mechanism of MAE on cardiovascular protection remains to be elucidated. Therefore, we investigated whether MAE affect platelet aggregation and thrombosis. Materials and Methods. The anti-platelet activity of MAE was studied using rat platelets. The extent of anti-platelet activity of MAE was assayed in collagen-induced platelet aggregation. ATP and serotonin release was carried out. The activation of integrin α IIb β 3 and phosphorylation of signaling molecules, including MAPK and Akt, were investigated with cytofluorometer and immunoblotting, respectively. The thrombus formation in vivo was also evaluated in arteriovenous shunt model of rats. Results. HPLC chromatographic analysis revealed that MAE contained rutin and isoquercetin. MAE dose-dependently inhibited collagen-induced platelet aggregation. MAE also attenuated serotonin secretion and thromboxane A2 formation. In addition, the extract in vivo activity showed that MAE at 100, 200, and 400 mg/kg significantly and dose-dependently attenuated thrombus formation in rat arterio-venous shunt model by 52.3% (P < 0.001), 28.3% (P < 0.01), and 19.1% (P < 0.05), respectively. Conclusions. MAE inhibit platelet activation, TXB2 formation, serotonin secretion, aggregation, and thrombus formation. The plant extract could be considered as a candidate to anti-platelet and antithrombotic agent.
Evidence-based Complementary and Alternative Medicine 02/2014; 2014:639548. DOI:10.1155/2014/639548 · 1.88 Impact Factor
"The amount of intracellular calcium ([Ca 2+ ]i) was determined with fura-2/AM as described previously . Briefly, the platelet-rich plasma was incubated with 5 μM of fura-2/AM for 60 min at 37 °C. "
[Show abstract][Hide abstract] ABSTRACT: Mushrooms possess untapped source of enormous natural compounds showing anti-inflammatory, antioxidant and anti-platelet activities. Paxillus curtisii, wild mushroom, is a rich source of curtisian E (CE) reported for neuroprotective effects; however, its anti-platelet effect was unknown. Here, therefore, we investigated the anti-platelet activity of CE in rat platelets. Curtisian E (12.5 - 200 μM) attenuated collagen (2.5 μg/ml), thrombin (0.1 U/ml) and ADP (10 μM) induced platelet aggregation in vitro. Likewise, CE diminished intracellular calcium and adenosine triphosphate (ATP) release in collagen activated platelets. Fibrinogen binding and fibronectin adhesion to platelets were also inhibited. While CE downregulated c-jun N-terminal kinase (JNK), extracellular related kinase (ERK), p38, and Akt dose dependently in collagen stimulated platelets, it upregulated intraplatelet cyclic adenosine monophosphate (cAMP) and vasodilator-stimulated-phosphoprotein (VASP) phosphorylation. Protein kinase A inhibitor (H-89) markedly inhibited p-VASP(157) protein expression, suggesting cAMP-PKA-VASP(157) pathway may mediate its anti-platelet effect and thus CE could be considered as a potential anti-thrombotic agent.
"Interestingly, quercetin-mediated attenuation of P-selectin expression and MAP kinase activation in this study suggests that the antiplatelet activity of the compound could be linked to its regulation of hemostatic- and inflammatory responses. Since platelets are involved in inflammation, P-selectin expression on the membrane of activated platelets is the main link between platelets and inflammatory cells [7, 45] and quercetin-mediated suppression of P-selectin expression and MAPKs activation in this paper may be attributed to its anti-inflammatory property. "
[Show abstract][Hide abstract] ABSTRACT: Background. Progressive diseases including cancer, metabolic, and cardiovascular disorders are marked by platelet activation and chronic inflammation. Studies suggest that dietary flavonoids such as quercetin possess antioxidant, anti-inflammatory, and antiplatelet properties, which could prevent various chronic diseases including atherosclerosis and thrombosis. However, the mechanism and the signaling pathway that links quercetin's antiplatelet activity with its anti-inflammatory property is limited and thus further exploration is required. The aim of this paper was to examine the link between antiplatelet and anti-inflammatory roles of quercetin in agonist-induced platelet activation. Methods. Quercetin effects on agonist-activated platelet-aggregation, granule-secretion, [Ca(2+)](i), and glycoprotein-IIb/IIIa activation were examined. Its effects on PI3K/Akt, VASP, and MAPK phosphorylations were also studied on collaged-activated platelets. Results. Quercetin dose dependently suppressed collagen, thrombin, or ADP-induced platelet aggregation. It significantly inhibited collagen-induced ATP release, P-selectin expression, [Ca(2+)](i) mobilization, integrin-α(IIb)β(3) activation, and augmented cAMP and VASP levels. Moreover, quercetin attenuated PI3K, Akt, ERK2, JNK1, and p38 MAPK activations, which were supported by platelet-aggregation inhibition with the respective kinase inhibitors. Conclusion. Quercetin-mediated antiplatelet activity involves PI3K/Akt inactivation, cAMP elevation, and VASP stimulation that, in turn, suppresses MAPK phosphorylations. This result suggests quercetin may have a potential to treat cardiovascular diseases involving aberrant platelet activation and inflammation.
Evidence-based Complementary and Alternative Medicine 12/2012; 2012(5):485262. DOI:10.1155/2012/485262 · 1.88 Impact Factor
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