A noble function of BAY 11-7082: Inhibition of platelet aggregation mediated by an elevated cAMP-induced VASP, and decreased ERK2/JNK1 phosphorylations
ABSTRACT 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.
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ABSTRACT: Platelets are small anucleate blood cells derived from megakaryocytes. In addition to their pivotal roles in hemostasis, platelets are the smallest, yet most abundant, immune cells and regulate inflammation, immunity, and disease progression. Although platelets lack DNA, and thus no functional transcriptional activities, they are nonetheless rich sources of RNAs, possess an intact spliceosome, and are thus capable of synthesizing proteins. Previously, it was thought that platelet RNAs and translational machinery were remnants from the megakaryocyte. We now know that the initial description of platelets as "cellular fragments" is an antiquated notion, as mounting evidence suggests otherwise. Therefore, it is reasonable to hypothesize that platelet transcription factors are not vestigial remnants from megakaryocytes, but have important, if only partly understood functions. Proteins play multiple cellular roles to minimize energy expenditure for maximum cellular function; thus, the same can be expected for transcription factors. In fact, numerous transcription factors have non-genomic roles, both in platelets and in nucleated cells. Our lab and others have discovered the presence and non-genomic roles of transcription factors in platelets, such as the nuclear factor kappa β (NFκB) family of proteins and peroxisome proliferator-activated receptor gamma (PPARγ). In addition to numerous roles in regulating platelet activation, functional transcription factors can be transferred to vascular and immune cells through platelet microparticles. This method of transcellular delivery of key immune molecules may be a vital mechanism by which platelet transcription factors regulate inflammation and immunity. At the very least, platelets are an ideal model cell to dissect out the non-genomic roles of transcription factors in nucleated cells. There is abundant evidence to suggest that transcription factors in platelets play key roles in regulating inflammatory and hemostatic functions.Frontiers in Immunology 02/2015; 6:48. DOI:10.3389/fimmu.2015.00048
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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 · 2.18 Impact Factor
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ABSTRACT: The current study intended to examine the signal transduction pathway of N-(quinolin-2-ylmethyl) butane-1, 4-diamine (QMA) in antiplatelet aggregation. Rats were divided randomly into five groups: control group; QMA-treated groups (0.3, 1, and 3 mg/kg); and r-Hirudin-treated group (0.3 mg/kg). Sample groups intravenously injected the corresponding agents once a day for 5 days; control group took 0.9% NaCl in the same way. Ten minutes after the last injection, blood samples were obtained from the rat abdominal aorta. Aggregation ex vivo was tested after irritating platelets by 1.5 U/ml thrombin for 5 min with a platelet aggregometer. Malondialdehyde production, activity of superoxide dismutase and nitric oxide production were determined by the microplate reader. Measurement of [Ca]i was performed using a fluorescence spectrophotometer. Thromboxane A2, cyclic adenosine monophosphate and cyclic guanosine monophosphate levels, vasodilator-stimulated phosphoprotein, and mitogen-activated protein kinase phosphorylation were measured with ELISA kits. Phospholipase C γ2 and protein kinase C were observed by immunoblotting study. QMA inhibited thrombin-induced platelet aggregation ex vivo. QMA significantly elevated superoxide dismutase activity, levels of cyclic adenosine monophosphate, cyclic guanosine monophosphate, nitric oxide, and subsequently promoted vasodilator-stimulated phosphoprotein phosphorylation. Meanwhile, QMA suppressed phospholipase C γ2, protein kinase C and mitogen-activated protein kinase phosphorylation, as well as malondialdehyde, thromboxane A2 formation and [Ca]i mobilization. QMA has a strong antiplatelet potential via its multitarget mechanism.Blood coagulation & fibrinolysis: an international journal in haemostasis and thrombosis 02/2014; 25(5). DOI:10.1097/MBC.0000000000000091 · 1.38 Impact Factor