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Mouse Platelets (mouse + platelet)
Selected AbstractsDistribution and dynamic changes of sphingolipids in blood in response to platelet activationJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 12 2006F. DAHM Summary.,Background:,Sphingolipids are signaling molecules in a range of biological processes. While sphingosine-1-phosphate (S1P) is thought to be abundantly stored in platelets and released upon stimulation, knowledge about the distribution and function of other sphingolipids in blood is lacking. Objectives:,To analyze the sphingolipid content of blood components with special emphasis on dynamic changes in platelets. Methods:,Blood components from mice and humans were prepared by gradient centrifugation and analyzed by liquid chromatography,mass spectrometry. Additionally, murine platelets were activated in vitro and in vivo. Results:,Isolated non-activated platelets of mice were devoid of S1P, but instead contained dihydrosphingosine-1-phosphate (dhS1P), along with a high concentration of ceramide. Activation of platelets in vitro led to a loss of dhS1P and an increase in sphingosine, accompanied by a reduction of ceramide content. Platelet activation in vivo led to an immediate and continuous rise of dhS1P in plasma, while S1P remained stable. The sphingolipid distribution of human blood was markedly different from mice. Human platelets contained dhS1P in addition to S1P. Conclusions:,Mouse platelets contain dhS1P instead of S1P. Platelet activation causes loss of dhS1P and breakdown of ceramide, implying ceramidase activation. Release of dhS1P from activated platelets might be a novel signaling pathway. Finally, the sphingolipid composition of mouse and human blood shows large differences, which must be considered when studying sphingolipid biology. [source] Molecular basis of platelet activation by an ,IIb,3-CHAMPS peptideJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 2 2009B. GRYGIELSKA Summary.,Background:,A novel method, known as computed helical anti-membrane protein (CHAMP), for the design of peptides that bind with high affinity and selectivity to transmembrane helices was recently described and illustrated using peptides that bind ,IIb- and ,v-integrin subunits, which induce selective activation of integrins ,IIb,3 and ,v,3, respectively [1]. Objectives:,In the present study, we have investigated the ability of an ,IIb-CHAMPS peptide (termed integrin-activatory-peptide or IAP) to stimulate protein tyrosine phosphorylation and aggregation in human and mouse platelets. Methods:,The ability of IAP to stimulate platelet aggregation and dense granule secretion was measured in washed preparations of human and mouse platelets. Samples were taken for measurement of tyrosine phosphorylation. Results:,IAP stimulates robust tyrosine phosphorylation of the tyrosine kinase Syk and the FcR ,-chain, but only weak phosphorylation of PLC,2. Aggregation to low but not high concentrations of IAP is reduced in the presence of the Src kinase inhibitor, PP1, or by inhibitors of the two feedback agonists, ADP and thromboxane A2 (TxA2) suggesting that activation is reinforced by Src kinase-driven release of ADP and TxA2. Unexpectedly, aggregation by IAP is only partially inhibited in human and mouse platelets deficient in integrin ,IIb,3. Further, IAP induces partial aggregation of formaldehyde-fixed platelets. Conclusions:,The present study demonstrates that the ,IIb-CHAMPS peptide induces platelet activation through integrin ,IIb,3-dependent and independent pathways with the former mediating tyrosine phosphorylation of FcR ,-chain and Syk. The use of the ,IIb-CHAMPS peptide to study integrin ,IIb,3 function is compromised by non-integrin-mediated effects. [source] The P2Y1 receptor plays an essential role in the platelet shape change induced by collagen when TxA2 formation is preventedJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 6 2004P. Mangin Summary., ADP and TxA2 are secondary agonists which play an important role as cofactors when platelets are activated by agonists such as collagen or thrombin. The aim of the present study was to characterize the role of the ADP receptor P2Y1 in collagen-induced activation of washed platelets. Inhibition of P2Y1 alone with the selective antagonist MRS2179 prolonged the lag phase preceding aggregation in response to low or high concentrations of fibrillar collagen, without affecting the maximum amplitude of aggregation or secretion. A combination of MRS2179 and aspirin resulted in complete inhibition of platelet shape change at low and high collagen concentrations, together with a profound decrease in aggregation and secretion. Scanning electron microscopy showed that these platelets had conserved the discoid morphology typical of the resting state. A lack of shape change was also observed in aspirin-treated P2Y1 - and G,q -deficient mouse platelets and in ,-storage pool-deficient platelets from Fawn Hooded rats. In contrast, when the second ADP receptor P2Y12 was inhibited with AR-C69931MX, aspirin-treated platelets were still able to change shape and displayed only a moderate decrease in aggregation and secretion. In conclusion, this study provides evidence that collagen requires not only the TxA2 receptor Tp,, but also P2Y1, to induce platelet shape change. [source] Fps/Fes and Fer non-receptor protein-tyrosine kinases regulate collagen- and ADP-induced platelet aggregationJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 5 2003Y. A. Senis Summary., Fps/Fes and Fer proto-oncoproteins are structurally related non-receptor protein-tyrosine kinases implicated in signaling downstream from cytokines, growth factors and immune receptors. We show that Fps/Fes and Fer are expressed in human and mouse platelets, and are activated following stimulation with collagen and collagen-related peptide (CRP), suggesting a role in GPVI receptor signaling. Fer was also activated following stimulation with thrombin and a protease-activated receptor4 (PAR4)-activating peptide, suggesting a role in signaling downstream from the G protein-coupled PAR4. There were no detectable perturbations in CRP-induced activation of Syk, PLC,2, cortactin, Erk, Jnk, Akt or p38 in platelets from mice lacking Fps/Fes, Fer, or both kinases. Platelets lacking Fps/Fes, from a targeted fps/fes null strain of mice, showed increased rates and amplitudes of collagen-induced aggregation, relative to wild-type platelets. P-Selectin expression was also elevated on the surface of Fps/Fes-null platelets in response to CRP. Fer-deficient platelets, from mice targeted with a kinase-inactivating mutation, disaggregated more rapidly than wild-type platelets in response to ADP. This report provides the first evidence that Fps/Fes and Fer are expressed in platelets and become activated downstream from the GPVI collagen receptor, and that Fer is activated downstream from a G-protein coupled receptor. Furthermore, using targeted mouse models we show that deficiency in Fps/Fes or Fer resulted in disregulated platelet aggregation and disaggregation, demonstrating a role for these kinases in regulating platelet functions. [source] Lineage-specific overexpression of the P2Y1 receptor induces platelet hyper-reactivity in transgenic miceJOURNAL OF THROMBOSIS AND HAEMOSTASIS, Issue 1 2003B. Hechler Summary., In order to investigate the role of the platelet P2Y1 receptor in several aspects of platelet activation and thrombosis, transgenic (TG) mice overexpressing this receptor specifically in the megakaryocytic/platelet lineage were generated using the promoter of the tissue-specific platelet factor 4 gene. Studies of the saturation binding of [33P]2MeSADP in the presence or absence of the selective P2Y1 antagonist MRS2179 indicated that wild-type (WT) mouse platelets bore 150 ± 31 P2Y1 receptors and TG platelets 276 ± 34, representing an 84% increase in P2Y1 receptor density. This led to a well defined phenotype of platelet hyper-reactivity in vitro, as shown by increased aggregations in response to adenosine 5,-diphosphate (ADP) and low concentration of collagen in TG as compared with WT platelets. Moreover, overexpression of the P2Y1 receptor enabled ADP to induce granule secretion, unlike in WT platelets, which suggests that the level of P2Y1 expression is critical for this event. Our results further suggest that the weak responses of normal platelets to ADP are due to a limited number of P2Y1 receptors rather than to activation of a specific transduction pathway. TG mice displayed a shortened bleeding time and an increased sensitivity to in vivo platelet aggregation induced by infusion of a mixture of collagen and epinephrine. Overall, these findings emphasize the importance of the P2Y1 receptor in hemostasis and thrombosis and suggest that variable expression levels of this receptor on platelets might play a role in thrombotic states in human, which remains to be assessed. [source] Ultrastructure of activated mouse platelets: A qualitative scanning electron microscopy studyMICROSCOPY RESEARCH AND TECHNIQUE, Issue 6 2008E. Pretorius Abstract Platelets form an integral part of the coagulation process, and their ultrastructure can provide valuable information regarding diseases associated with hemostasis. During coagulation, platelets aggregate; this aggregation can be achieved in vitro, by adding thrombin to platelet-rich plasma. Previous research showed that human thrombin could be used successfully to activate mouse platelets. When conservative changes are included, the amino acid similarity between human and mouse thrombin is ,75%. In this qualitative study, we compare the ultrastructure of mouse platelet aggregates activated by human thrombin as well as two concentrations of mouse thrombin, using the scanning electron microscope. Results show that both human and mouse thrombin activate platelets to form aggregates with typical pseudopodia formation. Magnification up to 250,000× showed membrane morphology with the open canalicular system pores visible in both the mouse- and human-activated platelets. It is therefore concluded that mouse platelets can be successfully aggregated using either mouse or human thrombin. Microsc. Res. Tech. 2008. © 2008 Wiley-Liss, Inc. [source] | ||||||||||