Jianguo Jin - Academia.edu (original) (raw)
Papers by Jianguo Jin
Thrombosis and Haemostasis, 2010
SummaryThough antiplatelet drugs are proven beneficial to patients with coronary heart disease an... more SummaryThough antiplatelet drugs are proven beneficial to patients with coronary heart disease and stroke, more effective and safer antiplatelet drugs are still needed. In this study we report the antiplatelet effects and mechanism of BF0801, a novel adenine derivative. BF0801 dramatically inhibited platelet aggregation and ATP release induced by ADP, 2MeSADP, AYPGKF, SFLLRN or convulxin without affecting shape change in vitro. It also potentiated the inhibitory effects of adenosine-based P2Y12 antagonist AR-C69931MX or phosphodiesterase (PDE) inhibitor IBMX on platelet aggregation. The cAMP levels in both resting and forskolin-stimulated platelets were increased by BF0801 suggesting its PDE inhibitor activity, which is further confirmed by the concentration-dependent suppression of BF0801 on the native and recombinant PDE. Similar to AR-C69931MX, BF0801 drastically inhibited 2MeSADP-induced adenylyl cyclase inhibition in platelets indicating its P2Y12 antagonism activity, which is ...
Blood, 2009
770 Protease-activated receptors (PARs) are G-protein coupled receptors that are activated by pro... more 770 Protease-activated receptors (PARs) are G-protein coupled receptors that are activated by proteases. Thrombin is the major agonist for PAR1 and PAR4, whereas tryptase and coagulation factor Xa are the agonists for PAR2. In addition to these major agonists, PARs can be activated by other coagulation proteases. The physiological agonist of PAR3 has not been identified to date; as a result, the molecular pharmacology and physiology of PAR3 remain poorly understood. The purpose of this study is to identify a physiological agonist to PAR3. We used PAR4 null murine platelets, which are known to express only PAR3. In this study, we tested the effect of several coagulation proteases and found that only coagulation factor XIIa (FXIIa) activated PAR4-/- murine platelets, in a concentration-dependent manner. FXIIa caused murine platelet shape change, aggregation, secretion and thromboxane A2 generation and this activation was abolished by C1 esterase inhibitor, a FXIIa inhibitor. FXIIa-ind...
Synthetic Metals, 1989
A new polyconjugated polymer, poly(l,4-phenylene-l,3,5-hexatrienylene-l,4phenylenevinylene) was s... more A new polyconjugated polymer, poly(l,4-phenylene-l,3,5-hexatrienylene-l,4phenylenevinylene) was synthesized via the sulfonium salt process. The polymer obtained in powder form could be doped with 12 and exhibited a maximum conductivity of about ixl0-3 Scm-I. Compared with poly(l,4-phenylenevinylene), the presence of the longer hexatrienylene unit along the backbone improves dopability with 12 and, therefore, the electrical conductivity. The same polymer, when doped with H2SO4, exhibited a maximum conductivity of about 2x10-I Scm-I .
Journal of Neuroimmunology, 1998
Circulation Research, 2020
Objective: Heart failure (HF) with preserved ejection fraction (HFpEF) is characterized by a pres... more Objective: Heart failure (HF) with preserved ejection fraction (HFpEF) is characterized by a preserved cardiac EF, the presence of HF symptoms and diastolic dysfunction. There is a lack of animal models for exploring the mechanisms and treatments of HFpEF. This study aimed to test if cardiomyocyte (CMs) specific, inducible Cavβ2a transgenic (Cavβ2a TG) mice, having hypercontractile CMs, could be a model for HFpEF. Methods: High (HE) and low (LE) expression Cavβ2a TG mice were studied since transgene expression at the age of 2m till the age of 8m monthly to evaluate the systolic and diastolic function. At 8m, animals were euthanized for intra-left ventricular hemodynamic measurement, myocyte function and histological analyses, and Western blotting measurements. Results: LE and HE TG ventricular myocytes (VMs) had greater Ca2+ currents at the age of 2m (LE increased by 95.5%; HE increased by 171.9% ) and maintained at a similar level at the age of 8m. VM contraction and calcium transi...
Ł € Signal Transduction (1930 articles)Ł € Hemostasis, Thrombosis, and Vascular Biology (2497 art... more Ł € Signal Transduction (1930 articles)Ł € Hemostasis, Thrombosis, and Vascular Biology (2497 articles)Ł € Cell Adhesion and Motility (790 articles)Articles on similar topics can be found in the following Blood collectionshttp://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#repub\_requests Information about reproducing this article in parts or in its entirety may be found online at:http://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#reprints Information about ordering reprints may be found online at:http://bloodjournal.hematologylibrary.org/site/subscriptions/index.xhtml Information about subscriptions and ASH membership may be found online at:Copyright 2011 by The American Society of Hematology; all rights reserved. Washington DC 20036.by the American Society of Hematology, 2021 L St, NW, Suite 900, Blood (print ISSN 0006-4971, online ISSN 1528-0020), is published weekly€€€€From bloodjournal.hematologylibrary.org by guest on January 7, 2012. For personal use...
Persistent activation of the sympathoadrenergic system in CHF can cause adverse cardiac remodelin... more Persistent activation of the sympathoadrenergic system in CHF can cause adverse cardiac remodeling, cardiac myocyte death, and fibrosis replacement. Reducing myocyte death has been proposed as one of the mechanisms responsible for the beneficial effects of β-blockers in heart failure patients. The mechanisms of β-adrenergic-mediated myocyte death still are not clearly defined and are the focus of this study. Some studies suggest that protein kinase A (PKA) is the mediator of β-adrenergic-induced myocyte apoptosis by altering Ca regulation, whereas others have suggested that Ca/camodulin-dependent kinase II (CaMKII) can mediate β-adrenergic-induced myocyte death through a PKAindependent process. A cAMP sensor, exchange protein directly activated by cAMP (EPAC), is expressed in the heart and has been suggested to activate CaMKII independent of PKA. The hypothesis of this study is that β-adrenergic-mediated myocyte death requires PKA activation and subsequently enhanced Ca signaling, b...
BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition... more BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition and P2Y12 antagonism Si Zhang1; Liang Hu1; Hongguang Du2; Yan Guo3; Yan Zhang1; Haixia Niu4; Jianguo Jin5; Jian Zhang4; Junling Liu4; Xiaohui Zhang3; Satya P. Kunapuli5; Zhongren Ding1 1Key Laboratory of Molecular Medicine, Ministry of Education, and Department of Biochemistry and Molecular Biology, Fudan University Shanghai Medical College, Shanghai, China; 2College of science, Beijing University of Chemical Technology, Beijing, China; 3Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China; 4Institutes of Medical Sciences, School of Medicine, Shanghai Jiao Tong University, Shanghai, China; 5Department of Physiology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA
Blood
4013 Poster Board III-949 Anti-platelet therapy is a widely accepted treatment regimen for the ma... more 4013 Poster Board III-949 Anti-platelet therapy is a widely accepted treatment regimen for the management of patients with cardiovascular risks. In clinics, many patients with cardiovascular diseases are managed on a combination therapy with Aspirin and Clopidogrel. Aspirin, being a COX-1 inhibitor blocks thromboxane generation and Clopidogrel metabolite antagonizes the P2Y12 receptor (ADP receptor). Thus these drugs block the two most important platelet positive feedback mediators and prevent amplification of platelet responses. The molecular mechanism of P2Y12 receptor antagonism on impairment of serum thromboxane is not understood to-date. Understanding this mechanism will clarify the molecular effects of P2Y12 antagonists when aspirin is clinically contraindicated in patients. Hypothesis We hypothesized that P2Y12 antagonism would lead to a decrease in serum thromboxane generation. We tested our hypothesis by employing both pharmacological and molecular genetic approaches. Metho...
Blood
2020 Positive regulatory role of Protein Kinase C (PKC) isoforms in platelets have been extensive... more 2020 Positive regulatory role of Protein Kinase C (PKC) isoforms in platelets have been extensively studied. However, negative regulatory roles of PKCs in platelets are poorly understood. In this study we investigated the mechanism by which PKCs negatively regulate ADP-induced thromboxane generation and identified PKC isoforms involved in this process. Pan PKC inhibition with GF109203X potentiated ADP-induced cPLA2 phosphorylation suggesting that PKCs negatively regulate thromboxane generation by regulating cPLA2 activation. Inhibition of PKCs potentiated ADP-induced ERK activation and intracellular calcium mobilization, two upstream signaling molecules of cPLA2.These data suggest that PKCs negatively regulate thromboxane by regulating ERK activation and calcium mobilization, which inturn regulate cPLA2 activation. Pan-PKC inhibition potentiated ADP-induced, P2Y1 receptor-mediated calcium mobilization in platelets independent of P2Y12-receptor. Pretreatment of platelets with GF10920...
Blood
Platelets have an integral function in maintaining hemostasis. Signaling pathways are strictly re... more Platelets have an integral function in maintaining hemostasis. Signaling pathways are strictly regulated to ensure the cessation of bleeding without precipitating a thrombotic episode. An important initial physiological step in this process is the interaction of platelets with freshly exposed subendothelial collagen after vascular injury. A major platelet receptor involved is the GPVI/FcRγ-chain complex. Upon activation of this receptor, the immunoreceptor tyrosine-based activation motif (ITAM) present in the FcRγ-chain is phosphorylated by Src family kinases. This causes the tyrosine kinase Syk to bind to the ITAM where it is autophosphorylated, initializing a signaling cascade that activates a number of proteins including PLCγ2, PI-3 kinase and small G proteins. Syk appears to play an early pivotal role in GPVI/FcRγ-chain signal transduction and its regulation is crucial. Recent studies have shown that platelet aggregation, in response to collagen-related peptide (CRP), is potenti...
Blood
Akt is a serine/threonine kinase that is activated by various agonists including thrombin and ADP... more Akt is a serine/threonine kinase that is activated by various agonists including thrombin and ADP in platelets, and activation of Akt in platelets is known to require Gi signaling pathways. Even though thrombin-induced Akt phosphorylation depends on secretion/Gi pathways, thrombin caused much stronger Akt phosphorylation than ADP and epinephrine. In this study, we investigated the contribution of G12/13 pathways to Akt phosphorylation mediated by Gi or Gz pathways. We used selective agonists to activate different G protein pathways. PAR4-activating peptide (AYPGKF) and thrombin failed to induce Akt phosphorylation in Gαq-deficient platelets, but Akt phosphorylation was restored to the levels achieved by AYPGKF and thrombin in wild-type platelets by selective supplement of either Gi or Gz signaling with 2-MeSADP and epinephrine, respectively. This phosphorylation of Akt was dramatically inhibited in the presence of PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazole[3,4-d]pyrimidine)...
Blood
ADP-induced platelet aggregation plays an important role in hemostasis and thrombosis. Human plat... more ADP-induced platelet aggregation plays an important role in hemostasis and thrombosis. Human platelets express two ADP receptors: the P2Y1 and P2Y12 receptors. The Gq-activating P2Y1 receptor plays an important role in ADP-induced platelet shape change, aggregation, and thromboxane A2 generation. In this study, we investigated the role of the carboxyl terminus of the human P2Y1 receptor in Gq activation. Human P2Y1 receptors, either wild type (P2Y1-WT) or a mutant in which the C-terminus was truncated (P2Y1-ΔT330-L373), were stably expressed with an HA-tag at the N-terminus in CHO-K1 cells. Stimulation of P2Y1-WT cells with 2-MeSADP caused inositol phosphate production and mobilization of calcium from intracellular stores. In contrast, P2Y1-ΔT330-L373 completely lost its response to 2-MeSADP, indicating that the C-terminus of the human P2Y1 receptor is essential for the activation of Gq. CHO-K1 cells expressing a chimeric P2Y12 receptor with the P2Y1 carboxyl terminus failed to elic...
Blood
Platelet secretion is an important physiological event in hemostasis. A number of agonists such a... more Platelet secretion is an important physiological event in hemostasis. A number of agonists such as thrombin and thromboxane A2 induce platelet secretion. ADP does not cause dense granule release in aspirin-treated platelets, although ADP induce platelet shape change, aggregation and alpha granule release through Gq and Gi pathways. The protease activated receptors 1 and 4, and the thromboxane receptor activate the G12/13 pathways in addition to the Gq pathways. We postulated that the platelet dense granule release reaction depends on both Gq and G12/13, and in the absence of signaling through either G protein abolishes secretion. In other words, co-stimulation of Gq and G12/13 pathways is a major requirement for dense granule release in platelets. We rationalize that because U46619 and thrombin can activate both Gq and G12/13, they cause platelet dense granule release, whereas ADP fails to cause platelet secretion from dense granules because it does not activate G12/13. As a first s...
Thrombosis and haemostasis, Jan 28, 2016
Phospholipase C (PLC)-β2 (gene PLCB2) is a critical regulator of platelet responses upon activati... more Phospholipase C (PLC)-β2 (gene PLCB2) is a critical regulator of platelet responses upon activation. Mechanisms regulating of PLC-β2 expression in platelets/MKs are unknown. Our studies in a patient with platelet PLC-β2 deficiency revealed the PLCB2 coding sequence to be normal and decreased platelet PLC-β2 mRNA, suggesting a defect in transcriptional regulation. PLCB2 5'- upstream region of the patient revealed a heterozygous 13 bp deletion (-1645/-1633 bp) encompassing a consensus sequence for nuclear factor-κB (NF-κB). This was subsequently detected in three of 50 healthy subjects. To understand the mechanisms regulating PLC-β2, we studied the effect of this variation in the PLCB2. Gel-shift studies using nuclear extracts from human erythroleukaemia (HEL) cells or recombinant p65 showed NF-κB binding to oligonucleotide with NF-κB site; in luciferase reporter studies its deletion reduced PLCB2 promoter activity. PLCB2 expression was decreased by siRNA knockdown of NF-κB p65 su...
Thrombosis and Haemostasis, 2010
SummaryThough antiplatelet drugs are proven beneficial to patients with coronary heart disease an... more SummaryThough antiplatelet drugs are proven beneficial to patients with coronary heart disease and stroke, more effective and safer antiplatelet drugs are still needed. In this study we report the antiplatelet effects and mechanism of BF0801, a novel adenine derivative. BF0801 dramatically inhibited platelet aggregation and ATP release induced by ADP, 2MeSADP, AYPGKF, SFLLRN or convulxin without affecting shape change in vitro. It also potentiated the inhibitory effects of adenosine-based P2Y12 antagonist AR-C69931MX or phosphodiesterase (PDE) inhibitor IBMX on platelet aggregation. The cAMP levels in both resting and forskolin-stimulated platelets were increased by BF0801 suggesting its PDE inhibitor activity, which is further confirmed by the concentration-dependent suppression of BF0801 on the native and recombinant PDE. Similar to AR-C69931MX, BF0801 drastically inhibited 2MeSADP-induced adenylyl cyclase inhibition in platelets indicating its P2Y12 antagonism activity, which is ...
Blood, 2009
770 Protease-activated receptors (PARs) are G-protein coupled receptors that are activated by pro... more 770 Protease-activated receptors (PARs) are G-protein coupled receptors that are activated by proteases. Thrombin is the major agonist for PAR1 and PAR4, whereas tryptase and coagulation factor Xa are the agonists for PAR2. In addition to these major agonists, PARs can be activated by other coagulation proteases. The physiological agonist of PAR3 has not been identified to date; as a result, the molecular pharmacology and physiology of PAR3 remain poorly understood. The purpose of this study is to identify a physiological agonist to PAR3. We used PAR4 null murine platelets, which are known to express only PAR3. In this study, we tested the effect of several coagulation proteases and found that only coagulation factor XIIa (FXIIa) activated PAR4-/- murine platelets, in a concentration-dependent manner. FXIIa caused murine platelet shape change, aggregation, secretion and thromboxane A2 generation and this activation was abolished by C1 esterase inhibitor, a FXIIa inhibitor. FXIIa-ind...
Synthetic Metals, 1989
A new polyconjugated polymer, poly(l,4-phenylene-l,3,5-hexatrienylene-l,4phenylenevinylene) was s... more A new polyconjugated polymer, poly(l,4-phenylene-l,3,5-hexatrienylene-l,4phenylenevinylene) was synthesized via the sulfonium salt process. The polymer obtained in powder form could be doped with 12 and exhibited a maximum conductivity of about ixl0-3 Scm-I. Compared with poly(l,4-phenylenevinylene), the presence of the longer hexatrienylene unit along the backbone improves dopability with 12 and, therefore, the electrical conductivity. The same polymer, when doped with H2SO4, exhibited a maximum conductivity of about 2x10-I Scm-I .
Journal of Neuroimmunology, 1998
Circulation Research, 2020
Objective: Heart failure (HF) with preserved ejection fraction (HFpEF) is characterized by a pres... more Objective: Heart failure (HF) with preserved ejection fraction (HFpEF) is characterized by a preserved cardiac EF, the presence of HF symptoms and diastolic dysfunction. There is a lack of animal models for exploring the mechanisms and treatments of HFpEF. This study aimed to test if cardiomyocyte (CMs) specific, inducible Cavβ2a transgenic (Cavβ2a TG) mice, having hypercontractile CMs, could be a model for HFpEF. Methods: High (HE) and low (LE) expression Cavβ2a TG mice were studied since transgene expression at the age of 2m till the age of 8m monthly to evaluate the systolic and diastolic function. At 8m, animals were euthanized for intra-left ventricular hemodynamic measurement, myocyte function and histological analyses, and Western blotting measurements. Results: LE and HE TG ventricular myocytes (VMs) had greater Ca2+ currents at the age of 2m (LE increased by 95.5%; HE increased by 171.9% ) and maintained at a similar level at the age of 8m. VM contraction and calcium transi...
Ł € Signal Transduction (1930 articles)Ł € Hemostasis, Thrombosis, and Vascular Biology (2497 art... more Ł € Signal Transduction (1930 articles)Ł € Hemostasis, Thrombosis, and Vascular Biology (2497 articles)Ł € Cell Adhesion and Motility (790 articles)Articles on similar topics can be found in the following Blood collectionshttp://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#repub\_requests Information about reproducing this article in parts or in its entirety may be found online at:http://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#reprints Information about ordering reprints may be found online at:http://bloodjournal.hematologylibrary.org/site/subscriptions/index.xhtml Information about subscriptions and ASH membership may be found online at:Copyright 2011 by The American Society of Hematology; all rights reserved. Washington DC 20036.by the American Society of Hematology, 2021 L St, NW, Suite 900, Blood (print ISSN 0006-4971, online ISSN 1528-0020), is published weekly€€€€From bloodjournal.hematologylibrary.org by guest on January 7, 2012. For personal use...
Persistent activation of the sympathoadrenergic system in CHF can cause adverse cardiac remodelin... more Persistent activation of the sympathoadrenergic system in CHF can cause adverse cardiac remodeling, cardiac myocyte death, and fibrosis replacement. Reducing myocyte death has been proposed as one of the mechanisms responsible for the beneficial effects of β-blockers in heart failure patients. The mechanisms of β-adrenergic-mediated myocyte death still are not clearly defined and are the focus of this study. Some studies suggest that protein kinase A (PKA) is the mediator of β-adrenergic-induced myocyte apoptosis by altering Ca regulation, whereas others have suggested that Ca/camodulin-dependent kinase II (CaMKII) can mediate β-adrenergic-induced myocyte death through a PKAindependent process. A cAMP sensor, exchange protein directly activated by cAMP (EPAC), is expressed in the heart and has been suggested to activate CaMKII independent of PKA. The hypothesis of this study is that β-adrenergic-mediated myocyte death requires PKA activation and subsequently enhanced Ca signaling, b...
BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition... more BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition and P2Y12 antagonism Si Zhang1; Liang Hu1; Hongguang Du2; Yan Guo3; Yan Zhang1; Haixia Niu4; Jianguo Jin5; Jian Zhang4; Junling Liu4; Xiaohui Zhang3; Satya P. Kunapuli5; Zhongren Ding1 1Key Laboratory of Molecular Medicine, Ministry of Education, and Department of Biochemistry and Molecular Biology, Fudan University Shanghai Medical College, Shanghai, China; 2College of science, Beijing University of Chemical Technology, Beijing, China; 3Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China; 4Institutes of Medical Sciences, School of Medicine, Shanghai Jiao Tong University, Shanghai, China; 5Department of Physiology, Temple University School of Medicine, Philadelphia, Pennsylvania, USA
Blood
4013 Poster Board III-949 Anti-platelet therapy is a widely accepted treatment regimen for the ma... more 4013 Poster Board III-949 Anti-platelet therapy is a widely accepted treatment regimen for the management of patients with cardiovascular risks. In clinics, many patients with cardiovascular diseases are managed on a combination therapy with Aspirin and Clopidogrel. Aspirin, being a COX-1 inhibitor blocks thromboxane generation and Clopidogrel metabolite antagonizes the P2Y12 receptor (ADP receptor). Thus these drugs block the two most important platelet positive feedback mediators and prevent amplification of platelet responses. The molecular mechanism of P2Y12 receptor antagonism on impairment of serum thromboxane is not understood to-date. Understanding this mechanism will clarify the molecular effects of P2Y12 antagonists when aspirin is clinically contraindicated in patients. Hypothesis We hypothesized that P2Y12 antagonism would lead to a decrease in serum thromboxane generation. We tested our hypothesis by employing both pharmacological and molecular genetic approaches. Metho...
Blood
2020 Positive regulatory role of Protein Kinase C (PKC) isoforms in platelets have been extensive... more 2020 Positive regulatory role of Protein Kinase C (PKC) isoforms in platelets have been extensively studied. However, negative regulatory roles of PKCs in platelets are poorly understood. In this study we investigated the mechanism by which PKCs negatively regulate ADP-induced thromboxane generation and identified PKC isoforms involved in this process. Pan PKC inhibition with GF109203X potentiated ADP-induced cPLA2 phosphorylation suggesting that PKCs negatively regulate thromboxane generation by regulating cPLA2 activation. Inhibition of PKCs potentiated ADP-induced ERK activation and intracellular calcium mobilization, two upstream signaling molecules of cPLA2.These data suggest that PKCs negatively regulate thromboxane by regulating ERK activation and calcium mobilization, which inturn regulate cPLA2 activation. Pan-PKC inhibition potentiated ADP-induced, P2Y1 receptor-mediated calcium mobilization in platelets independent of P2Y12-receptor. Pretreatment of platelets with GF10920...
Blood
Platelets have an integral function in maintaining hemostasis. Signaling pathways are strictly re... more Platelets have an integral function in maintaining hemostasis. Signaling pathways are strictly regulated to ensure the cessation of bleeding without precipitating a thrombotic episode. An important initial physiological step in this process is the interaction of platelets with freshly exposed subendothelial collagen after vascular injury. A major platelet receptor involved is the GPVI/FcRγ-chain complex. Upon activation of this receptor, the immunoreceptor tyrosine-based activation motif (ITAM) present in the FcRγ-chain is phosphorylated by Src family kinases. This causes the tyrosine kinase Syk to bind to the ITAM where it is autophosphorylated, initializing a signaling cascade that activates a number of proteins including PLCγ2, PI-3 kinase and small G proteins. Syk appears to play an early pivotal role in GPVI/FcRγ-chain signal transduction and its regulation is crucial. Recent studies have shown that platelet aggregation, in response to collagen-related peptide (CRP), is potenti...
Blood
Akt is a serine/threonine kinase that is activated by various agonists including thrombin and ADP... more Akt is a serine/threonine kinase that is activated by various agonists including thrombin and ADP in platelets, and activation of Akt in platelets is known to require Gi signaling pathways. Even though thrombin-induced Akt phosphorylation depends on secretion/Gi pathways, thrombin caused much stronger Akt phosphorylation than ADP and epinephrine. In this study, we investigated the contribution of G12/13 pathways to Akt phosphorylation mediated by Gi or Gz pathways. We used selective agonists to activate different G protein pathways. PAR4-activating peptide (AYPGKF) and thrombin failed to induce Akt phosphorylation in Gαq-deficient platelets, but Akt phosphorylation was restored to the levels achieved by AYPGKF and thrombin in wild-type platelets by selective supplement of either Gi or Gz signaling with 2-MeSADP and epinephrine, respectively. This phosphorylation of Akt was dramatically inhibited in the presence of PP2 (4-amino-5-(4-chlorophenyl)-7-(t-butyl)pyrazole[3,4-d]pyrimidine)...
Blood
ADP-induced platelet aggregation plays an important role in hemostasis and thrombosis. Human plat... more ADP-induced platelet aggregation plays an important role in hemostasis and thrombosis. Human platelets express two ADP receptors: the P2Y1 and P2Y12 receptors. The Gq-activating P2Y1 receptor plays an important role in ADP-induced platelet shape change, aggregation, and thromboxane A2 generation. In this study, we investigated the role of the carboxyl terminus of the human P2Y1 receptor in Gq activation. Human P2Y1 receptors, either wild type (P2Y1-WT) or a mutant in which the C-terminus was truncated (P2Y1-ΔT330-L373), were stably expressed with an HA-tag at the N-terminus in CHO-K1 cells. Stimulation of P2Y1-WT cells with 2-MeSADP caused inositol phosphate production and mobilization of calcium from intracellular stores. In contrast, P2Y1-ΔT330-L373 completely lost its response to 2-MeSADP, indicating that the C-terminus of the human P2Y1 receptor is essential for the activation of Gq. CHO-K1 cells expressing a chimeric P2Y12 receptor with the P2Y1 carboxyl terminus failed to elic...
Blood
Platelet secretion is an important physiological event in hemostasis. A number of agonists such a... more Platelet secretion is an important physiological event in hemostasis. A number of agonists such as thrombin and thromboxane A2 induce platelet secretion. ADP does not cause dense granule release in aspirin-treated platelets, although ADP induce platelet shape change, aggregation and alpha granule release through Gq and Gi pathways. The protease activated receptors 1 and 4, and the thromboxane receptor activate the G12/13 pathways in addition to the Gq pathways. We postulated that the platelet dense granule release reaction depends on both Gq and G12/13, and in the absence of signaling through either G protein abolishes secretion. In other words, co-stimulation of Gq and G12/13 pathways is a major requirement for dense granule release in platelets. We rationalize that because U46619 and thrombin can activate both Gq and G12/13, they cause platelet dense granule release, whereas ADP fails to cause platelet secretion from dense granules because it does not activate G12/13. As a first s...
Thrombosis and haemostasis, Jan 28, 2016
Phospholipase C (PLC)-β2 (gene PLCB2) is a critical regulator of platelet responses upon activati... more Phospholipase C (PLC)-β2 (gene PLCB2) is a critical regulator of platelet responses upon activation. Mechanisms regulating of PLC-β2 expression in platelets/MKs are unknown. Our studies in a patient with platelet PLC-β2 deficiency revealed the PLCB2 coding sequence to be normal and decreased platelet PLC-β2 mRNA, suggesting a defect in transcriptional regulation. PLCB2 5'- upstream region of the patient revealed a heterozygous 13 bp deletion (-1645/-1633 bp) encompassing a consensus sequence for nuclear factor-κB (NF-κB). This was subsequently detected in three of 50 healthy subjects. To understand the mechanisms regulating PLC-β2, we studied the effect of this variation in the PLCB2. Gel-shift studies using nuclear extracts from human erythroleukaemia (HEL) cells or recombinant p65 showed NF-κB binding to oligonucleotide with NF-κB site; in luciferase reporter studies its deletion reduced PLCB2 promoter activity. PLCB2 expression was decreased by siRNA knockdown of NF-κB p65 su...