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Satya P. Kunapuli - One of the best experts on this subject based on the ideXlab platform.
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fucoidan is a novel Platelet Agonist for the c type lectin like receptor 2 clec 2
Journal of Biological Chemistry, 2013Co-Authors: Bhanukanth Manne, Todd M Getz, Craig E Hughes, Carol T Dangelmaier, Steve P Watson, Osama Alshehri, Ulhas P Naik, Satya P. KunapuliAbstract:Fucoidan, a sulfated polysaccharide from Fucus vesiculosus, decreases bleeding time and clotting time in hemophilia, possibly through inhibition of tissue factor pathway inhibitor. However, its effect on Platelets and the receptor by which fucoidan induces cellular processes has not been elucidated. In this study, we demonstrate that fucoidan induces Platelet activation in a concentration-dependent manner. Fucoidan-induced Platelet activation was completely abolished by the pan-Src family kinase (SFK) inhibitor, PP2, or when Syk is inhibited. PP2 abolished phosphorylations of Syk and Phospholipase C-γ2. Fucoidan-induced Platelet activation had a lag phase, which is reminiscent of Platelet activation by collagen and CLEC-2 receptor Agonists. Platelet activation by fucoidan was only slightly inhibited in FcRγ-chain null mice, indicating that fucoidan was not acting primarily through GPVI receptor. On the other hand, fucoidan-induced Platelet activation was inhibited in Platelet-specific CLEC-2 knock-out murine Platelets revealing CLEC-2 as a physiological target of fucoidan. Thus, our data show fucoidan as a novel CLEC-2 receptor Agonist that activates Platelets through a SFK-dependent signaling pathway. Furthermore, the efficacy of fucoidan in hemophilia raises the possibility that decreased bleeding times could be achieved through activation of Platelets.
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fucoidan is a novel Platelet Agonist for clec 2 receptor
Blood, 2012Co-Authors: Bhanukanth Manne, Todd M Getz, Craig E Hughes, Carol T Dangelmaier, Steve P Watson, Satya P. KunapuliAbstract:Abstract Abstract 94 Fucoidan, a sulphated polysaccharide from fucus vesiculosus, decreases bleeding time and clotting time in hemophilia, possibly through inhibition of tissue factor pathway inhibitor (TFPI) (Prasad et al., Blood 111:672, 2008). The decrease in bleeding times in the hemophilia animal models by in vivo administration of fucoidan suggests the beneficial effect of fucoidan as a novel treatment. Furthermore, in vitro studies using Platelet poor plasma from hemophilia animal models and human patients has shown that fucoidan inhibits TFPI thereby contributing to an increase in the extrinsic coagulation pathway activity. The effect of fucoidan on Platelets however has not been studied. As it is known that the Platelet count remains unaffected in hemophilia A patients and bleeding times are primarily measured to assess normal Platelet function, we hypothesize that the decrease in bleeding times in the hemophilia animal models may be due to Platelet activation by fucoidan. In this study, we demonstrate for the first time that fucoidan induces Platelet activation in a concentration dependent manner. Fucoidan-induced Platelet activation is completely abolished by the pan-Src family kinase (SFK) inhibitor, PP2, and in the absence of Syk and PLC-g2. Furthermore, fucoidan-induced Platelet activation has a lag phase, which is reminiscent of Platelet activation by collagen and by CLEC-2 receptor Agonists. Platelet activation by fucoidan however was only slightly inhibited in FcRg-chain null mice indicating that fucoidan is not acting primarily through GPVI receptor. On the other hand, fucoidan-induced Platelet activation was inhibited in CLEC-2 deficient mouse Platelets revealing CLEC-2 as a physiological target of fucoidan. Thus, our data shows fucoidan as a novel CLEC-2 receptor Agonist that activates Platelets through an SFK-dependent signaling pathway. Further, the efficacy of fucoidan in hemophilia raises the possibility that decreased bleeding times could be achieved through activation of Platelets. Download : Download high-res image (57KB) Download : Download full-size image Disclosures: No relevant conflicts of interest to declare.
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adenosine diphosphate adp induced thromboxane a2generation in human Platelets requires coordinated signaling through integrin αiibβ3 and adp receptors
Blood, 2002Co-Authors: Jianguo Jin, Todd M. Quinton, Jin Zhang, Susan E Rittenhouse, Satya P. KunapuliAbstract:Adenosine diphosphate (ADP) is a Platelet Agonist that causes Platelet shape change and aggregation as well as generation of thromboxane A2, another Platelet Agonist, through its effects on P2Y1, P2Y12, and P2X1 receptors. It is now reported that both 2-propylthio-D-βγ-dichloromethylene adenosine 5′-triphosphate (AR-C67085), a P2Y12 receptor–selective antAgonist, and adenosine-2′-phosphate-5′-phosphate (A2P5P), a P2Y1 receptor–selective antAgonist, inhibited ADP-induced thromboxane A2 generation in a concentration-dependent manner, indicating that coactivation of the P2Y12 and P2Y1 receptors is essential for this event. SC49992, a fibrinogen receptor antAgonist, blocked ADP-induced Platelet aggregation and thromboxane A2 production in a concentration-dependent manner. Similarly, P2 receptor antAgonists or SC49992 blocked ADP-induced arachidonic acid liberation. Whereas SC49992 blocked arachidonic acid–induced Platelet aggregation, it failed to inhibit thromboxane A2 generation induced by arachidonic acid. Thus, ADP-induced arachidonic acid liberation, but not subsequent conversion to thromboxane A2, requires outside-in signaling through the fibrinogen receptor. The Fab fragment of ligand-induced binding site–6 (LIBS6) antibody, which induces a fibrinogen-binding site on the integrin αIIbβ3, caused both Platelet aggregation and thromboxane A2 generation. Inhibitors of phosphoinositide 3-kinase, Syk, Src kinases, or protein tyrosine phosphatases inhibited Platelet aggregation but not thromboxane A2 generation, indicating that these signaling molecules have no significant role in phospholipase A2 activation. In the presence of P2 receptor antAgonists A2P5P or AR-C67085, LIBS6 failed to generate thromboxane A2, suggesting that inside-out signaling through ADP receptors is necessary for this event. It was concluded that both outside-in signaling from the fibrinogen receptor and inside-out signaling from the P2Y1 and P2Y12 receptors are necessary for phospholipase A2 activation, resulting in arachidonic acid liberation and thromboxane A2 generation.
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adenosine diphosphate adp induced thromboxane a2generation in human Platelets requires coordinated signaling through integrin αiibβ3 and adp receptors
Blood, 2002Co-Authors: Jianguo Jin, Todd M. Quinton, Jin Zhang, Susan E Rittenhouse, Satya P. KunapuliAbstract:Adenosine diphosphate (ADP) is a Platelet Agonist that causes Platelet shape change and aggregation as well as generation of thromboxane A(2), another Platelet Agonist, through its effects on P2Y1, P2Y12, and P2X1 receptors. It is now reported that both 2-propylthio-D-beta gamma-dichloromethylene adenosine 5'-triphosphate (AR-C67085), a P2Y12 receptor-selective antAgonist, and adenosine-2'-phosphate-5'-phosphate (A2P5P), a P2Y1 receptor-selective antAgonist, inhibited ADP-induced thromboxane A(2) generation in a concentration-dependent manner, indicating that coactivation of the P2Y12 and P2Y1 receptors is essential for this event. SC49992, a fibrinogen receptor antAgonist, blocked ADP-induced Platelet aggregation and thromboxane A(2) production in a concentration-dependent manner. Similarly, P2 receptor antAgonists or SC49992 blocked ADP-induced arachidonic acid liberation. Whereas SC49992 blocked arachidonic acid-induced Platelet aggregation, it failed to inhibit thromboxane A(2) generation induced by arachidonic acid. Thus, ADP-induced arachidonic acid liberation, but not subsequent conversion to thromboxane A(2), requires outside-in signaling through the fibrinogen receptor. The Fab fragment of ligand-induced binding site-6 (LIBS6) antibody, which induces a fibrinogen-binding site on the integrin alpha(IIb)beta(3), caused both Platelet aggregation and thromboxane A(2) generation. Inhibitors of phosphoinositide 3-kinase, Syk, Src kinases, or protein tyrosine phosphatases inhibited Platelet aggregation but not thromboxane A(2) generation, indicating that these signaling molecules have no significant role in phospholipase A(2) activation. In the presence of P2 receptor antAgonists A2P5P or AR-C67085, LIBS6 failed to generate thromboxane A(2), suggesting that inside-out signaling through ADP receptors is necessary for this event. It was concluded that both outside-in signaling from the fibrinogen receptor and inside-out signaling from the P2Y1 and P2Y12 receptors are necessary for phospholipase A(2) activation, resulting in arachidonic acid liberation and thromboxane A(2) generation.
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coactivation of two different g protein coupled receptors is essential for adp induced Platelet aggregation
Proceedings of the National Academy of Sciences of the United States of America, 1998Co-Authors: Satya P. KunapuliAbstract:ADP is an important Platelet Agonist causing shape change and aggregation required for physiological hemostasis. We recently demonstrated the existence of two distinct G protein-coupled ADP receptors on Platelets, one coupled to phospholipase C, P2Y1, and the other to inhibition of adenylyl cyclase, P2TAC. In this study, using specific antAgonists for these two receptors, we demonstrated that concomitant intracellular signaling from both the P2TAC and P2Y1 receptors is essential for ADP-induced Platelet aggregation. Inhibition of signaling through either receptor, by specific antAgonists, is sufficient to block ADP-induced Platelet aggregation. Furthermore, signaling through the P2TAC receptor could be replaced by activation of α2A-adrenergic receptors. On the other hand, activation of serotonin receptors supplements signaling through the P2Y1 receptor. Moreover, this mechanism of ADP-induced Platelet aggregation could be mimicked by coactivation of two non-ADP receptors coupled to Gi and Gq, neither of which can cause Platelet aggregation by itself. We propose that Platelet aggregation results from concomitant signaling from both the Gi and Gq, a mechanism by which G protein-coupled receptors elicit a physiological response.
Yun Zhang - One of the best experts on this subject based on the ideXlab platform.
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bm tff2 a toad trefoil factor promotes cell migration survival and wound healing
Biochemical and Biophysical Research Communications, 2010Co-Authors: Yong Q Zhang, Guoyu Yu, Yang Xiang, Jianbo Wu, Ping Jiang, Yun ZhangAbstract:Research highlights: {yields} Bm-TFF2 binds to epithelial cells and induces cell migration and wound healing. {yields} Bm-TFF2 suppresses cell apoptosis. {yields} Bm-TFF2 has no effect on cell proliferation. -- Abstract: Toad skin is naked and continually confronted by various injurious factors. Constant skin renewal and repairs occur frequently. However, the mechanisms of the renewal and repair have not clearly elucidated. In our previous work, a trefoil factor (TFF), Bm-TFF2, has been purified from the Bombina maxima skin and characterized as a Platelet Agonist. The mRNA of TFFs in toad skin was up-regulated greatly during the metamorphosis, indicating a pivotal role of TFFs in amphibian skin. Here, we presented the effects of Bm-TFF2 on the cell migration, apoptosis and proliferation. Bm-TFF2 bound to epithelial cells and showed strong cell motility activity. At the concentrations of 1-100 nM, Bm-TFF2-induced migration of human epithelial AGS and HT-29 cells, and rat intestinal epithelial IEC-6 cell lines. The in vitro wound healing assay also verified the activity of Bm-TFF2. Bm-TFF2 could also inhibit cell apoptosis induced by ceramide and sodium butyrate. The cell migration-promoting activity was abolished by MEK1 inhibitors, U0126 and PD98059, suggesting that ERK1/2 activation is crucial for Bm-TFF2 to stimulate cell migration. more » Taken together, Bm-TFF2 promoted wound healing by stimulating cell migration via MAPK pathway and preventing cell apoptosis. The potent biological activity of Bm-TFF2 makes it a useful molecular tool for further studies of structure-function relationship of the related human TFFs. « less
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tmva a snake c type lectin like protein from trimeresurus mucrosquamatus venom activates Platelet via gpib
Toxicon, 2004Co-Authors: Min Su, Jianxin Song, Xinding Zhou, Hongmei Ouyang, Wanyu Wang, Yuliang Xiong, Yun ZhangAbstract:Abstract TMVA is a C-type lectin-like protein with potent Platelet activating activity from Trimeresurus mucrosquamatus venom. In the absence of von Willebrand factor (vWF), TMVA dose-dependently induced aggregation of washed Platelets. Anti-GP Ib monoclonal antibodies (mAbs), HIP1, specifically inhibited TMVA-induced aggregation in a dose-dependent manner. The aggregation was also inhibited by mAb P2 (an anti-GP IIb mAb). Flow cytometric analysis revealed that FITC-TMVA bound to human formalin-fixed Platelets in a saturable manner, and its binding was specifically blocked by HIP1 in a dose-dependent manner. Flow cytometric analysis showed that TMVA did not bind to Platelet GPIX, GPIIb, GPIIIa, GPIa, GPIIa and GPIV. Moreover, the Platelet aggregation induced by TMVA was partially inhibited when Platelet was pretreated with mocarhagin, a snake venom protease that specifically cleaves human GPIb. These results suggest that TMVA is a strong Platelet Agonist via GPIb and it might have multiple functional binding-sites on GPIb molecule or on other unknown receptor.
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stejnulxin a novel snake c type lectin like protein from trimeresurus stejnegeri venom is a potent Platelet Agonist acting specifically via gpvi
Thrombosis and Haemostasis, 2003Co-Authors: Wenhui Lee, Kenneth J Clemetson, Yun ZhangAbstract:Stejnulxin, a novel snake C-type lectin-like protein with potent Platelet activating activity, was purified and characterized from Trimeresurus stejnegeri venom. Under non-reducing conditions, it migrated on a SDS-polyacrylamide gel with an apparent molecular mass of 120 kDa. On reduction, it separated into three polypeptide subunits with apparent molecular masses of 16 kDa (alpha), 20 kDa (beta1) and 22 kDa (beta2), respectively. The complete amino acid sequences of its subunits were deduced from cloned cDNAs. The N-terminal sequencing and cDNA cloning indicated that beta1 and beta2 subunits of stejnulxin have identical amino acid sequences and each contains two N-glycosylation sites. Accordingly, the molecular mass difference between beta1 and beta2 is caused by glycosylation heterogenity. The subunit amino acid sequences of stejnulxin are similar to those of convulxin, with sequence identities of 52.6% and 66.4% for the alpha and beta, respectively. Stejnulxin induced human Platelet aggregation in a dose-dependent manner. Antibodies against alphaIIbbeta3 inhibited the aggregation response to stejnulxin, indicating that activation of alphaIIbbeta3 and binding of fibrinogen are involved in stejnulxin-induced Platelet aggregation. Antibodies against GPIbalpha or alpha2beta1 as well as echicetin or rhodocetin had no significant effect on stejnulxin-induced Platelet aggregation. However, Platelet activation induced by stejnulxin was blocked by anti-GPVI antibodies. In addition, stejnulxin induced a tyrosine phosphorylation profile in Platelets that resembled that produced by convulxin. Biotinylated stejnulxin bound specifically to Platelet membrane GPVI.
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stejnulxin a novel snake c type lectin like protein from trimeresurus stejnegeri venom is a potent Platelet Agonist acting specifically via gpvi
Thrombosis and Haemostasis, 2003Co-Authors: Xiaoyan Du, Kenneth J Clemetson, Qiumin Lu, Yun ZhangAbstract:Stejnulxin, a novel snake C-type lectin-like protein with potent Platelet activating activity, was purified and characterized from Trimeresurus stejnegeri venom. Under non-reducing conditions, it migrated on a SDS-polyacrylamide gel with an apparent molecular mass of 120 kDa. On reduction, it separated into three polypeptide subunits with apparent molecular masses of 16 kDa, 20 kDa and 22 kDa, respectively.The com-plete amino acid sequences of its subunits were deduced from cloned cDNAs.The N-terminal sequencing and cDNA cloning indicated that β1 and β2 subunits of stejnulxin have identical amino acid sequences and each contains two N-glycosylation sites. Accordingly, the molecular mass difference between β1 and s 2 is caused by glycosylation heterogenity. The subunit amino acid sequences of stejnulxin are similar to those of convulxin, with sequence identities of 52.6% and 66.4% for the a and β , respectively. Stejnulxin induced human Platelet aggrega-tion in a dose-dependent manner. Antibodies against αIibβ3 inhibited the aggregation response to stejnulxin, indicating that activation of αIibβ3 and binding of fibrinogen are involved in stejnulxin-induced Platelet aggregation. Antibodies against GPIbα or a α2β1 as well as echicetin or rhodocetin had no significant effect on stejnulxin-induced Platelet aggregation. However, Platelet activation induced by stejnulxin was blocked by anti-GPVI antibodies. In addition, stejnulxin induced a tyrosine phosphorylation profile in Platelets that resembled that produced by convulxin. Biotinylated stejnulxin bound specifically to Platelet membrane GPVI.
Kenneth J Clemetson - One of the best experts on this subject based on the ideXlab platform.
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Platelet activation induces metalloproteinase dependent gp vi cleavage to down regulate Platelet reactivity to collagen
Blood, 2005Co-Authors: Gillian Stephens, Martine Jandrotperrus, Yibing Yan, Jeanluc Villeval, Kenneth J Clemetson, David R PhillipsAbstract:Glycoprotein (GP) VI, the primary collagen receptor on Platelets, has been shown to have variable expression, possibly as a consequence of immune modulation. The present study was designed to determine the mechanism by which GP VI clearance occurs. We found that direct activation of GP VI both by a GP VI-specific antibody and by GP VI ligands (collagen and convulxin) reduced binding of biotinylated convulxin to the stimulated Platelets. Analysis of immunoblots of Platelets and supernatants showed that the stimulated Platelets contained less GP VI, while the soluble fraction contained a 57-kDa cleavage product. Stimulation of Platelets with PAR-1 Agonists (TRAP peptide and thrombin) also caused GP VI cleavage, although the amount of GP VI loss was less than that observed with direct GP VI ligands. The metalloproteinase (MMP) inhibitors GM6001 and TAPI prevented both the clearance of GP VI from the Platelet surface and the appearance of the soluble cleavage product. Induction of GP VI cleavage caused specific down-regulation of collagen-induced Platelet aggregation, providing a mechanism for the modulation of Platelet responsiveness to this important Platelet Agonist.
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stejnulxin a novel snake c type lectin like protein from trimeresurus stejnegeri venom is a potent Platelet Agonist acting specifically via gpvi
Thrombosis and Haemostasis, 2003Co-Authors: Wenhui Lee, Kenneth J Clemetson, Yun ZhangAbstract:Stejnulxin, a novel snake C-type lectin-like protein with potent Platelet activating activity, was purified and characterized from Trimeresurus stejnegeri venom. Under non-reducing conditions, it migrated on a SDS-polyacrylamide gel with an apparent molecular mass of 120 kDa. On reduction, it separated into three polypeptide subunits with apparent molecular masses of 16 kDa (alpha), 20 kDa (beta1) and 22 kDa (beta2), respectively. The complete amino acid sequences of its subunits were deduced from cloned cDNAs. The N-terminal sequencing and cDNA cloning indicated that beta1 and beta2 subunits of stejnulxin have identical amino acid sequences and each contains two N-glycosylation sites. Accordingly, the molecular mass difference between beta1 and beta2 is caused by glycosylation heterogenity. The subunit amino acid sequences of stejnulxin are similar to those of convulxin, with sequence identities of 52.6% and 66.4% for the alpha and beta, respectively. Stejnulxin induced human Platelet aggregation in a dose-dependent manner. Antibodies against alphaIIbbeta3 inhibited the aggregation response to stejnulxin, indicating that activation of alphaIIbbeta3 and binding of fibrinogen are involved in stejnulxin-induced Platelet aggregation. Antibodies against GPIbalpha or alpha2beta1 as well as echicetin or rhodocetin had no significant effect on stejnulxin-induced Platelet aggregation. However, Platelet activation induced by stejnulxin was blocked by anti-GPVI antibodies. In addition, stejnulxin induced a tyrosine phosphorylation profile in Platelets that resembled that produced by convulxin. Biotinylated stejnulxin bound specifically to Platelet membrane GPVI.
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stejnulxin a novel snake c type lectin like protein from trimeresurus stejnegeri venom is a potent Platelet Agonist acting specifically via gpvi
Thrombosis and Haemostasis, 2003Co-Authors: Xiaoyan Du, Kenneth J Clemetson, Qiumin Lu, Yun ZhangAbstract:Stejnulxin, a novel snake C-type lectin-like protein with potent Platelet activating activity, was purified and characterized from Trimeresurus stejnegeri venom. Under non-reducing conditions, it migrated on a SDS-polyacrylamide gel with an apparent molecular mass of 120 kDa. On reduction, it separated into three polypeptide subunits with apparent molecular masses of 16 kDa, 20 kDa and 22 kDa, respectively.The com-plete amino acid sequences of its subunits were deduced from cloned cDNAs.The N-terminal sequencing and cDNA cloning indicated that β1 and β2 subunits of stejnulxin have identical amino acid sequences and each contains two N-glycosylation sites. Accordingly, the molecular mass difference between β1 and s 2 is caused by glycosylation heterogenity. The subunit amino acid sequences of stejnulxin are similar to those of convulxin, with sequence identities of 52.6% and 66.4% for the a and β , respectively. Stejnulxin induced human Platelet aggrega-tion in a dose-dependent manner. Antibodies against αIibβ3 inhibited the aggregation response to stejnulxin, indicating that activation of αIibβ3 and binding of fibrinogen are involved in stejnulxin-induced Platelet aggregation. Antibodies against GPIbα or a α2β1 as well as echicetin or rhodocetin had no significant effect on stejnulxin-induced Platelet aggregation. However, Platelet activation induced by stejnulxin was blocked by anti-GPVI antibodies. In addition, stejnulxin induced a tyrosine phosphorylation profile in Platelets that resembled that produced by convulxin. Biotinylated stejnulxin bound specifically to Platelet membrane GPVI.
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alboluxin a snake c type lectin from trimeresurus albolabris venom is a potent Platelet Agonist acting via gpib and gpvi
Thrombosis and Haemostasis, 2002Co-Authors: Xiaoyan Du, Edith Magnenat, Timothy N C Wells, Kenneth J ClemetsonAbstract:Alboluxin, a potent Platelet activator, was purified from Trimere-surus albolabris venom with a mass of 120 kDa non-reduced and, after reduction, subunits of 17 and 24 kDa. Alboluxin induced a tyro-sine phosphorylation profile in Platelets that resembles those produced by collagen and convulxin, involving the time dependent tyrosine phos-phorylation of Fc receptor chain (Fc), phospholipase C2 (PLCγ2), LAT and p72 SYK . Antibodies against both GPIb and GPVI inhibited Platelet aggregation induced by alboluxin, whereas antibodies against α2 Β1 had no effect. Inhibition of αIIbβ3 reduced the aggregation respon-se to alboluxin, as well as tyrosine phosphorylation of Platelet proteins, showing that activation of αIIbβ3 and binding of fibrinogen are involv-ed in alboluxin-induced Platelet aggregation and it is not simply ag-glutination. N-terminal sequence data from the -subunit of alboluxin indicates that it belongs to the snake C-type lectin family. The C-type lectin subunits are larger than usual possibly due to post-translational modifications such as glycosylation. Alboluxin is a hexameric (αβ) 3 snake C-type lectin which activates Platelets via both GPIb and GPVI.
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alboluxin a snake c type lectin from trimeresurus albolabris venom is a potent Platelet Agonist acting via gpib and gpvi
Thrombosis and Haemostasis, 2002Co-Authors: Edith Magnenat, Timothy N C Wells, Kenneth J ClemetsonAbstract:Alboluxin, a potent Platelet activator, was purified from Trimeresurus albolabris venom with a mass of 120 kDa non-reduced and, after reduction, subunits of 17 and 24 kDa. Alboluxin induced a tyrosine phosphorylation profile in Platelets that resembles those produced by collagen and convulxin, involving the time dependent tyrosine phosphorylation of Fc receptor gamma chain (Fc gamma), phospholipase Cgamma2 (PLCgamma2), LAT and p72SYK. Antibodies against both GPIb and GPVI inhibited Platelet aggregation induced by alboluxin, whereas antibodies against alpha2beta1 had no effect. Inhibition of alphaIIb beta3 reduced the aggregation response to alboluxin, as well as tyrosine phosphorylation of Platelet proteins, showing that activation of alphaIIb beta3 and binding of fibrinogen are involved in alboluxin-induced Platelet aggregation and it is not simply agglutination. N-terminal sequence data from the beta-subunit of alboluxin indicates that it belongs to the snake C-type lectin family. The C-type lectin subunits are larger than usual possibly due to post-translational modifications such as glycosylation. Alboluxin is a hexameric (alphabeta)3 snake C-type lectin which activates Platelets via both GPIb and GPVI.
Xiaoyan Du - One of the best experts on this subject based on the ideXlab platform.
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stejnulxin a novel snake c type lectin like protein from trimeresurus stejnegeri venom is a potent Platelet Agonist acting specifically via gpvi
Thrombosis and Haemostasis, 2003Co-Authors: Xiaoyan Du, Kenneth J Clemetson, Qiumin Lu, Yun ZhangAbstract:Stejnulxin, a novel snake C-type lectin-like protein with potent Platelet activating activity, was purified and characterized from Trimeresurus stejnegeri venom. Under non-reducing conditions, it migrated on a SDS-polyacrylamide gel with an apparent molecular mass of 120 kDa. On reduction, it separated into three polypeptide subunits with apparent molecular masses of 16 kDa, 20 kDa and 22 kDa, respectively.The com-plete amino acid sequences of its subunits were deduced from cloned cDNAs.The N-terminal sequencing and cDNA cloning indicated that β1 and β2 subunits of stejnulxin have identical amino acid sequences and each contains two N-glycosylation sites. Accordingly, the molecular mass difference between β1 and s 2 is caused by glycosylation heterogenity. The subunit amino acid sequences of stejnulxin are similar to those of convulxin, with sequence identities of 52.6% and 66.4% for the a and β , respectively. Stejnulxin induced human Platelet aggrega-tion in a dose-dependent manner. Antibodies against αIibβ3 inhibited the aggregation response to stejnulxin, indicating that activation of αIibβ3 and binding of fibrinogen are involved in stejnulxin-induced Platelet aggregation. Antibodies against GPIbα or a α2β1 as well as echicetin or rhodocetin had no significant effect on stejnulxin-induced Platelet aggregation. However, Platelet activation induced by stejnulxin was blocked by anti-GPVI antibodies. In addition, stejnulxin induced a tyrosine phosphorylation profile in Platelets that resembled that produced by convulxin. Biotinylated stejnulxin bound specifically to Platelet membrane GPVI.
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alboluxin a snake c type lectin from trimeresurus albolabris venom is a potent Platelet Agonist acting via gpib and gpvi
Thrombosis and Haemostasis, 2002Co-Authors: Xiaoyan Du, Edith Magnenat, Timothy N C Wells, Kenneth J ClemetsonAbstract:Alboluxin, a potent Platelet activator, was purified from Trimere-surus albolabris venom with a mass of 120 kDa non-reduced and, after reduction, subunits of 17 and 24 kDa. Alboluxin induced a tyro-sine phosphorylation profile in Platelets that resembles those produced by collagen and convulxin, involving the time dependent tyrosine phos-phorylation of Fc receptor chain (Fc), phospholipase C2 (PLCγ2), LAT and p72 SYK . Antibodies against both GPIb and GPVI inhibited Platelet aggregation induced by alboluxin, whereas antibodies against α2 Β1 had no effect. Inhibition of αIIbβ3 reduced the aggregation respon-se to alboluxin, as well as tyrosine phosphorylation of Platelet proteins, showing that activation of αIIbβ3 and binding of fibrinogen are involv-ed in alboluxin-induced Platelet aggregation and it is not simply ag-glutination. N-terminal sequence data from the -subunit of alboluxin indicates that it belongs to the snake C-type lectin family. The C-type lectin subunits are larger than usual possibly due to post-translational modifications such as glycosylation. Alboluxin is a hexameric (αβ) 3 snake C-type lectin which activates Platelets via both GPIb and GPVI.
Jianguo Jin - One of the best experts on this subject based on the ideXlab platform.
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adenosine diphosphate adp induced thromboxane a2generation in human Platelets requires coordinated signaling through integrin αiibβ3 and adp receptors
Blood, 2002Co-Authors: Jianguo Jin, Todd M. Quinton, Jin Zhang, Susan E Rittenhouse, Satya P. KunapuliAbstract:Adenosine diphosphate (ADP) is a Platelet Agonist that causes Platelet shape change and aggregation as well as generation of thromboxane A2, another Platelet Agonist, through its effects on P2Y1, P2Y12, and P2X1 receptors. It is now reported that both 2-propylthio-D-βγ-dichloromethylene adenosine 5′-triphosphate (AR-C67085), a P2Y12 receptor–selective antAgonist, and adenosine-2′-phosphate-5′-phosphate (A2P5P), a P2Y1 receptor–selective antAgonist, inhibited ADP-induced thromboxane A2 generation in a concentration-dependent manner, indicating that coactivation of the P2Y12 and P2Y1 receptors is essential for this event. SC49992, a fibrinogen receptor antAgonist, blocked ADP-induced Platelet aggregation and thromboxane A2 production in a concentration-dependent manner. Similarly, P2 receptor antAgonists or SC49992 blocked ADP-induced arachidonic acid liberation. Whereas SC49992 blocked arachidonic acid–induced Platelet aggregation, it failed to inhibit thromboxane A2 generation induced by arachidonic acid. Thus, ADP-induced arachidonic acid liberation, but not subsequent conversion to thromboxane A2, requires outside-in signaling through the fibrinogen receptor. The Fab fragment of ligand-induced binding site–6 (LIBS6) antibody, which induces a fibrinogen-binding site on the integrin αIIbβ3, caused both Platelet aggregation and thromboxane A2 generation. Inhibitors of phosphoinositide 3-kinase, Syk, Src kinases, or protein tyrosine phosphatases inhibited Platelet aggregation but not thromboxane A2 generation, indicating that these signaling molecules have no significant role in phospholipase A2 activation. In the presence of P2 receptor antAgonists A2P5P or AR-C67085, LIBS6 failed to generate thromboxane A2, suggesting that inside-out signaling through ADP receptors is necessary for this event. It was concluded that both outside-in signaling from the fibrinogen receptor and inside-out signaling from the P2Y1 and P2Y12 receptors are necessary for phospholipase A2 activation, resulting in arachidonic acid liberation and thromboxane A2 generation.
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adenosine diphosphate adp induced thromboxane a2generation in human Platelets requires coordinated signaling through integrin αiibβ3 and adp receptors
Blood, 2002Co-Authors: Jianguo Jin, Todd M. Quinton, Jin Zhang, Susan E Rittenhouse, Satya P. KunapuliAbstract:Adenosine diphosphate (ADP) is a Platelet Agonist that causes Platelet shape change and aggregation as well as generation of thromboxane A(2), another Platelet Agonist, through its effects on P2Y1, P2Y12, and P2X1 receptors. It is now reported that both 2-propylthio-D-beta gamma-dichloromethylene adenosine 5'-triphosphate (AR-C67085), a P2Y12 receptor-selective antAgonist, and adenosine-2'-phosphate-5'-phosphate (A2P5P), a P2Y1 receptor-selective antAgonist, inhibited ADP-induced thromboxane A(2) generation in a concentration-dependent manner, indicating that coactivation of the P2Y12 and P2Y1 receptors is essential for this event. SC49992, a fibrinogen receptor antAgonist, blocked ADP-induced Platelet aggregation and thromboxane A(2) production in a concentration-dependent manner. Similarly, P2 receptor antAgonists or SC49992 blocked ADP-induced arachidonic acid liberation. Whereas SC49992 blocked arachidonic acid-induced Platelet aggregation, it failed to inhibit thromboxane A(2) generation induced by arachidonic acid. Thus, ADP-induced arachidonic acid liberation, but not subsequent conversion to thromboxane A(2), requires outside-in signaling through the fibrinogen receptor. The Fab fragment of ligand-induced binding site-6 (LIBS6) antibody, which induces a fibrinogen-binding site on the integrin alpha(IIb)beta(3), caused both Platelet aggregation and thromboxane A(2) generation. Inhibitors of phosphoinositide 3-kinase, Syk, Src kinases, or protein tyrosine phosphatases inhibited Platelet aggregation but not thromboxane A(2) generation, indicating that these signaling molecules have no significant role in phospholipase A(2) activation. In the presence of P2 receptor antAgonists A2P5P or AR-C67085, LIBS6 failed to generate thromboxane A(2), suggesting that inside-out signaling through ADP receptors is necessary for this event. It was concluded that both outside-in signaling from the fibrinogen receptor and inside-out signaling from the P2Y1 and P2Y12 receptors are necessary for phospholipase A(2) activation, resulting in arachidonic acid liberation and thromboxane A(2) generation.
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molecular basis for adp induced Platelet activation ii the p2y1 receptor mediates adp induced intracellular calcium mobilization and shape change in Platelets
Journal of Biological Chemistry, 1998Co-Authors: Jianguo Jin, James L Daniel, Satya P. KunapuliAbstract:Abstract ADP is an important Platelet Agonist causing shape change from smooth discoid shape to spiculated spheres and Platelet aggregation. However, the molecular mechanisms involved in ADP-induced Platelet activation have not been elucidated. We demonstrated earlier the existence of two distinct ADP receptors on Platelets, one coupled to phospholipase C, P2TPLC, and the other to inhibition of adenylyl cyclase, P2TAC (Daniel, J. L., Dangelmaier, C., Jin, J., Ashby, B., Smith, J. B., and Kunapuli, S. P. (1998)J. Biol. Chem. 273, 2024–2029), in addition to the previously described P2X1 receptor. Here we report the cloning of a cDNA clone encoding the P2Y1 receptor from a human Platelet cDNA library by homology screening with radiolabeled P2Y1-P2Y6 receptor cDNAs. ADP or 2-methyl(thio)-ADP-induced intracellular calcium increases were inhibited by the P2Y1 receptor-specific antAgonists, adenosine 3′-phosphate 5′-phosphosulfate (A3P5PS), adenosine 3′-phosphate 5′-phosphate (A3P5P), and adenosine 2′-phosphate 5′-phosphate (A2P5P), in a concentration-dependent manner, but not by ARL 66096 or α,β-MeATP. A3P5PS, A3P5P, and A2P5P also inhibited the shape change of aspirinated Platelets induced by 10 μm ADP or 3 μm 2-methyl(thio)-ADP in a concentration-dependent manner, with complete inhibition occurring at 300 μm. On the other hand ARL 66096 (100 nm), a potent P2TAC antAgonist and α,β-methylene-ATP (40 μm), a P2X1 receptor Agonist, had no effect on ADP-induced Platelet shape change. On the contrary, ADP-induced inhibition of adenylyl cyclase was blocked by ARL 66096, but not by α,β-MeATP or the P2Y1 receptor-specific antAgonists, A3P5PS, A3P5P, or A2P5P. These results demonstrate the role of the P2Y1 receptor in ADP-induced Platelet shape change and calcium mobilization and support the idea that several P2 receptors are involved in the regulation of different aspects of Platelet stimulus-response coupling.