Jianguo Jin - Profile on Academia.edu (original) (raw)

Papers by Jianguo Jin

British Journal of Pharmacology, Jun 1, 2001

P2Y receptor activation in many cell types leads to phospholipase C activation and accumulation o... more P2Y receptor activation in many cell types leads to phospholipase C activation and accumulation of inositol phosphates, while in blood platelets, C6-2B glioma cells, and in B10 microvascular endothelial cells a P2Y receptor subtype, which couples to inhibition of adenylyl cyclase, historically termed P2Y AC , (P2T AC or P 2T in platelets) has been identi®ed. Recently, this receptor has been cloned and designated P2Y 12 in keeping with current P2 receptor nomenclature. 2 Three selective P 2T receptor antagonists, with a range of anities, inhibited ADP-induced aggregation of washed human or rat platelets, in a concentration-dependent manner, with a rank order of antagonist potency (pIC 50 , human: rat) of AR-C78511 (8.5 : 9.1)4AR-C69581 (6.2 : 6.0)4AR-C70300 (5.4 : 5.1). However, these compounds had no eect on ADP-induced platelet shape change. 3 All three antagonists had no signi®cant eect on the ADP-induced inositol phosphate formation in 1321N1 astrocytoma cells stably expressing the P2Y 1 receptor, when used at concentrations that inhibit platelet aggregation. 4 These antagonists also blocked ADP-induced inhibition of adenylyl cyclase in rat platelets and C6-2B cells with identical rank orders of potency and overlapping concentration ± response curves. 5 RT ± PCR and nucleotide sequence analyses revealed that the C6-2B cells express the P2Y 12 mRNA. 6 These data demonstrate that the P2Y AC receptor in C6-2B cells is pharmacologically identical to the P2T AC receptor in rat platelets.

Abstract 11418: Fibrosis and Calcium Signaling Regulation Are Critical Mechanisms in a Three-Hit HFpEF Mouse Model

Circulation, 2021

Introduction: There is a lack of animal models of heart failure with preserved ejection fraction ... more Introduction: There is a lack of animal models of heart failure with preserved ejection fraction (HFpEF). Hypothesis: A mouse model with increased Ca2+ influx receiving a high-fat diet (HFD) and L-NAME (3 hits) could be a model for HFpEF to help reveal the mechanism. Methods: Cardiomyocyte (CM) specific, inducible Cavβ2a low expression transgenic (Cavβ2a LE TG) and their control littermate mice were fed with either chow diet (LC and CC), HFD+L-NAME (LHLn, CHLn), or HFD+L-NAME+SAHA (LHLnS, CHLnS) since the age of 4months for 6 months. Echocardiography had been done monthly. Hemodynamic analysis, histological analyses, and Western blotting (WB) had been used to figure out the possible mechanisms after euthanization. Results: LHL (3 hits) mice had preserved left ventricular ejection fraction (LVEF) but the highest levels of ANP/BNP and the lowest strain rate. 3-Hit led to obvious diastolic dysfunction indicated by a higher ratio of mitral E wave to mitral annulus e’ velocity (E/e’ rati...

Protease-Activated Receptor 3 Negatively Regulates Plasmin-Mediated Protease-Activated Receptor 4 Activation in Platelets

Blood, 2007

Plasmin, a major extracellular protease, causes intracellular signals to mediate platelet aggrega... more Plasmin, a major extracellular protease, causes intracellular signals to mediate platelet aggregation. Previously, we reported that plasmin-mediated platelet aggregation predominantly occurs through proteolytic cleavage of protease-activated receptor 4 (Quinton et al J. Biol. Chem 2004). We showed that plasmin caused aggregation of mouse platelets more readily than human platelets. In this study, we investigated the mechanism of such a differential sensitivity of mouse platelets to plasmin, using transfected cell lines and platelets. In the platelet system, plasmin caused both human and mouse platelets to shape change and aggregate in a concentration-dependent manner with a different efficiency. Whereas 0.1 U/ml of plasmin causes full aggregation of mouse platelets, that dose only induces shape change in human platelets. In transfected COS7 cells, 1 U/ml plasmin caused a higher intracellular calcium mobilization through mouse PAR4 (mPAR4) than human PAR4 (hPAR4) activation. These re...

Cross talk between serine/threonine and tyrosine kinases regulates ADP-induced thromboxane generation in platelets

Thrombosis and haemostasis, Jan 7, 2015

ADP-induced thromboxane generation depends on Src family kinases (SFKs) and is enhanced with pan-... more ADP-induced thromboxane generation depends on Src family kinases (SFKs) and is enhanced with pan-protein kinase C (PKC) inhibitors, but it is not clear how these two events are linked. The aim of the current study is to investigate the role of Y311 phosphorylated PKCδ in regulating ADP-induced platelet activation. In the current study, we employed various inhibitors and murine platelets from mice deficient in specific molecules to evaluate the role of PKCδ in ADP-induced platelet responses. We show that, upon stimulation of platelets with 2MeSADP, Y311 on PKCδ is phosphorylated in a P2Y1/Gq and Lyn-dependent manner. By using PKCδ and Lyn knockout murine platelets, we also show that tyrosine phosphorylated PKCδ plays a functional role in mediating 2MeSADP-induced thromboxane generation. 2MeSADP-induced PKCδ Y311 phosphorylation and thromboxane generation were potentiated in human platelets pre-treated with either a pan-PKC inhibitor, GF109203X or a PKC α/β inhibitor and in PKC α or β...

Classical PKCs Regulate ADP-Induced Thromboxane Generation by Modulating Tyrosine Phosphorylation On Novel PKC Isoform Delta Through Shptp-1

Blood, Nov 16, 2012

Abstract 1064 Introduction: Adenosine Di-phosphate (ADP) is stored in dense granules of platelets... more Abstract 1064 Introduction: Adenosine Di-phosphate (ADP) is stored in dense granules of platelets and is released upon platelet activation acting as a feedback activator by binding to G-protein coupled P2Y1 and P2Y12 receptors. ADP stimulation causes platelets to change shape, aggregate, release dense and a-granule contents and synthesize thromboxane A2 that can further act as a feedback activator potentiating platelet responses by binding to thromboxane receptor (TP). Protein kinase C is a serine threonine specific kinase that regulates multiple platelet functional responses. Specific PKC isoforms regulating platelet responses downstream of ADP receptors are not completely known. Aim: The aim of the current study is to elucidate the role of PKC isoforms in regulating ADP-induced platelet functional responses in platelets. Methods: We sought to delineate the mechanism of ADP-induced platelet responses by performing platelet aggregation (aggregometry), ATP secretion (luciferin-luciferase reaction) and thromboxane generation (ELISA kit measuring TxB2) in human or murine platelets by pre-incubating the platelets with control (DMSO) or inhibitors wherever mentioned. We also evaluated the role of PKCd to ADP-induced platelet responses by using murine platelets lacking PKCd. Background and Results: Murugappan et al have shown that PKCd was not activated downstream of ADP receptors based on the inability of ADP to cause threonine 507 phosphorylation on PKCd in platelets. However, studies from other labs have shown that PKCd can be activated when it is phosphorylated on its tyrosine residues. In the current study we show that, upon stimulation with 2MeSADP, PKCd is phosphorylated on tyrosine residue 311 in a time-dependent manner in platelets (Fig A). Also, ADP-induced thromboxane generation (Fig B) and ADP-induced thromboxane-mediated dense granule secretion were significantly inhibited in PKCd knockout murine platelets compared to those of wild type platelets. Similarly, thromboxane generation downstream of ADP receptors in human platelets pre-incubated with a PKCd inhibitor is significantly inhibited compared to control indicating a role for PKCd in mediating ADP-induced responses in platelets. Bynagari et al have shown that ADP-induced thromboxane generation is potentiated in the presence of the pan-PKC inhibitor, GF 109203X and the isoform regulating this effect is PKCe. We observed that pre-incubation of PKCe knockout murine platelets with GF 109203X further potentiated ADP-induced thromboxane generation suggesting that there are other PKC isoforms negatively regulating ADP-induced thromboxane generation. We show that this potentiating effect of thromboxane generation with GF 109203X in WT or PKCe KO murine platelets correlate with an increase in the phosphorylation of Y311 on PKCd (Fig C) suggesting that ADP-induced thromboxane generation is regulated through PKCd Y311 phosphorylation. Tyrosine phosphorylation on PKCd is mediated by Src family kinases (SFKs) as the phosphorylation is abolished with PP2, a SFK inhibitor and is only partially inhibited in Fyn knockout murine platelets suggesting that other SFKs also mediate this tyrosine phosphorylation. Surprisingly, pre-incubation of platelets with LY-333531, a classical PKC isoform (a/b) inhibitor potentiated PKCd Y311 phosphorylation (Fig D) as well as thromboxane generation (Fig E) downstream of ADP receptors suggesting a role for classical PKCs. Also, platelets pre-incubated with LY-333531 showed a decrease in the phosphorylation of SHPTP-1 (Fig F), a tyrosine phosphatase, rendering it active. The active SHPTP-1 phosphatase may dephosphorylate and activate SFKs, which can now phosphorylate PKCd on Y311 in platelets. Conclusions: In the current study, we report for the first time that the novel PKC isoform d is tyrosine phosphorylated downstream of ADP receptors through which it mediates ADP-induced thromboxane generation. We also show a novel role for classical PKC isoforms a/b in regulating tyrosine phosphorylation on novel isoform, PKCd possibly through the tyrosine phosphatase SHPTP-1 and Src family kinases in platelets. Disclosures: No relevant conflicts of interest to declare.

and ADP receptors 3 β IIb α human platelets requires coordinated signaling through integrin induced thromboxane A2generation in - Adenosine diphosphate (ADP)

BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition and P2Y12 antagonism

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 ...

receptor-mediated platelet functional responses Role of G protein-gated inwardly-rectifying potassium channels in P2Y12

activation in collagen-related peptide-induced platelet θ Response:Role of PKC

Coagulation Factor XIIa Activates Platelets and Is the Physiological Agonist of Protease-Activated Receptor 3

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

Abstract 383: Cavβ2a Transgenic Mouse as a Hfpef Model With Hypercontractile Cardiomyocytes

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...

interaction with both extracellular cysteine residues, C17 and C270 Inactivation of the human P2Y12 receptor by thiol reagents requires

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)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:

protease-activated receptor-mediated Akt phosphorylation in platelets Relative contribution of G-protein-coupled pathways to

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...

Role of PKCθ in collagen-related peptide-induced platelet activation

Blood, 2009

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...

Tools and DRugs BF 0801 , a novel adenine derivative , inhibits platelet activation via phosphodiesterase inhibition and P 2 Y 12 antagonism

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

Antagonism of P2y12 Receptor Results in Severe Impairment of Serum Thromboxane Levels. Can We Manage Patients without Combination Therapy with Aspirin?

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...

Protein Kinase C Isoform ε Negatively Regulates ADP-Induced Thromboxane Generation by Regulating cPLA2 Activation and Calcium Mobilization In Platelets

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...

British Journal of Pharmacology, Jun 1, 2001

P2Y receptor activation in many cell types leads to phospholipase C activation and accumulation o... more P2Y receptor activation in many cell types leads to phospholipase C activation and accumulation of inositol phosphates, while in blood platelets, C6-2B glioma cells, and in B10 microvascular endothelial cells a P2Y receptor subtype, which couples to inhibition of adenylyl cyclase, historically termed P2Y AC , (P2T AC or P 2T in platelets) has been identi®ed. Recently, this receptor has been cloned and designated P2Y 12 in keeping with current P2 receptor nomenclature. 2 Three selective P 2T receptor antagonists, with a range of anities, inhibited ADP-induced aggregation of washed human or rat platelets, in a concentration-dependent manner, with a rank order of antagonist potency (pIC 50 , human: rat) of AR-C78511 (8.5 : 9.1)4AR-C69581 (6.2 : 6.0)4AR-C70300 (5.4 : 5.1). However, these compounds had no eect on ADP-induced platelet shape change. 3 All three antagonists had no signi®cant eect on the ADP-induced inositol phosphate formation in 1321N1 astrocytoma cells stably expressing the P2Y 1 receptor, when used at concentrations that inhibit platelet aggregation. 4 These antagonists also blocked ADP-induced inhibition of adenylyl cyclase in rat platelets and C6-2B cells with identical rank orders of potency and overlapping concentration ± response curves. 5 RT ± PCR and nucleotide sequence analyses revealed that the C6-2B cells express the P2Y 12 mRNA. 6 These data demonstrate that the P2Y AC receptor in C6-2B cells is pharmacologically identical to the P2T AC receptor in rat platelets.

Abstract 11418: Fibrosis and Calcium Signaling Regulation Are Critical Mechanisms in a Three-Hit HFpEF Mouse Model

Circulation, 2021

Introduction: There is a lack of animal models of heart failure with preserved ejection fraction ... more Introduction: There is a lack of animal models of heart failure with preserved ejection fraction (HFpEF). Hypothesis: A mouse model with increased Ca2+ influx receiving a high-fat diet (HFD) and L-NAME (3 hits) could be a model for HFpEF to help reveal the mechanism. Methods: Cardiomyocyte (CM) specific, inducible Cavβ2a low expression transgenic (Cavβ2a LE TG) and their control littermate mice were fed with either chow diet (LC and CC), HFD+L-NAME (LHLn, CHLn), or HFD+L-NAME+SAHA (LHLnS, CHLnS) since the age of 4months for 6 months. Echocardiography had been done monthly. Hemodynamic analysis, histological analyses, and Western blotting (WB) had been used to figure out the possible mechanisms after euthanization. Results: LHL (3 hits) mice had preserved left ventricular ejection fraction (LVEF) but the highest levels of ANP/BNP and the lowest strain rate. 3-Hit led to obvious diastolic dysfunction indicated by a higher ratio of mitral E wave to mitral annulus e’ velocity (E/e’ rati...

Protease-Activated Receptor 3 Negatively Regulates Plasmin-Mediated Protease-Activated Receptor 4 Activation in Platelets

Blood, 2007

Plasmin, a major extracellular protease, causes intracellular signals to mediate platelet aggrega... more Plasmin, a major extracellular protease, causes intracellular signals to mediate platelet aggregation. Previously, we reported that plasmin-mediated platelet aggregation predominantly occurs through proteolytic cleavage of protease-activated receptor 4 (Quinton et al J. Biol. Chem 2004). We showed that plasmin caused aggregation of mouse platelets more readily than human platelets. In this study, we investigated the mechanism of such a differential sensitivity of mouse platelets to plasmin, using transfected cell lines and platelets. In the platelet system, plasmin caused both human and mouse platelets to shape change and aggregate in a concentration-dependent manner with a different efficiency. Whereas 0.1 U/ml of plasmin causes full aggregation of mouse platelets, that dose only induces shape change in human platelets. In transfected COS7 cells, 1 U/ml plasmin caused a higher intracellular calcium mobilization through mouse PAR4 (mPAR4) than human PAR4 (hPAR4) activation. These re...

Cross talk between serine/threonine and tyrosine kinases regulates ADP-induced thromboxane generation in platelets

Thrombosis and haemostasis, Jan 7, 2015

ADP-induced thromboxane generation depends on Src family kinases (SFKs) and is enhanced with pan-... more ADP-induced thromboxane generation depends on Src family kinases (SFKs) and is enhanced with pan-protein kinase C (PKC) inhibitors, but it is not clear how these two events are linked. The aim of the current study is to investigate the role of Y311 phosphorylated PKCδ in regulating ADP-induced platelet activation. In the current study, we employed various inhibitors and murine platelets from mice deficient in specific molecules to evaluate the role of PKCδ in ADP-induced platelet responses. We show that, upon stimulation of platelets with 2MeSADP, Y311 on PKCδ is phosphorylated in a P2Y1/Gq and Lyn-dependent manner. By using PKCδ and Lyn knockout murine platelets, we also show that tyrosine phosphorylated PKCδ plays a functional role in mediating 2MeSADP-induced thromboxane generation. 2MeSADP-induced PKCδ Y311 phosphorylation and thromboxane generation were potentiated in human platelets pre-treated with either a pan-PKC inhibitor, GF109203X or a PKC α/β inhibitor and in PKC α or β...

Classical PKCs Regulate ADP-Induced Thromboxane Generation by Modulating Tyrosine Phosphorylation On Novel PKC Isoform Delta Through Shptp-1

Blood, Nov 16, 2012

Abstract 1064 Introduction: Adenosine Di-phosphate (ADP) is stored in dense granules of platelets... more Abstract 1064 Introduction: Adenosine Di-phosphate (ADP) is stored in dense granules of platelets and is released upon platelet activation acting as a feedback activator by binding to G-protein coupled P2Y1 and P2Y12 receptors. ADP stimulation causes platelets to change shape, aggregate, release dense and a-granule contents and synthesize thromboxane A2 that can further act as a feedback activator potentiating platelet responses by binding to thromboxane receptor (TP). Protein kinase C is a serine threonine specific kinase that regulates multiple platelet functional responses. Specific PKC isoforms regulating platelet responses downstream of ADP receptors are not completely known. Aim: The aim of the current study is to elucidate the role of PKC isoforms in regulating ADP-induced platelet functional responses in platelets. Methods: We sought to delineate the mechanism of ADP-induced platelet responses by performing platelet aggregation (aggregometry), ATP secretion (luciferin-luciferase reaction) and thromboxane generation (ELISA kit measuring TxB2) in human or murine platelets by pre-incubating the platelets with control (DMSO) or inhibitors wherever mentioned. We also evaluated the role of PKCd to ADP-induced platelet responses by using murine platelets lacking PKCd. Background and Results: Murugappan et al have shown that PKCd was not activated downstream of ADP receptors based on the inability of ADP to cause threonine 507 phosphorylation on PKCd in platelets. However, studies from other labs have shown that PKCd can be activated when it is phosphorylated on its tyrosine residues. In the current study we show that, upon stimulation with 2MeSADP, PKCd is phosphorylated on tyrosine residue 311 in a time-dependent manner in platelets (Fig A). Also, ADP-induced thromboxane generation (Fig B) and ADP-induced thromboxane-mediated dense granule secretion were significantly inhibited in PKCd knockout murine platelets compared to those of wild type platelets. Similarly, thromboxane generation downstream of ADP receptors in human platelets pre-incubated with a PKCd inhibitor is significantly inhibited compared to control indicating a role for PKCd in mediating ADP-induced responses in platelets. Bynagari et al have shown that ADP-induced thromboxane generation is potentiated in the presence of the pan-PKC inhibitor, GF 109203X and the isoform regulating this effect is PKCe. We observed that pre-incubation of PKCe knockout murine platelets with GF 109203X further potentiated ADP-induced thromboxane generation suggesting that there are other PKC isoforms negatively regulating ADP-induced thromboxane generation. We show that this potentiating effect of thromboxane generation with GF 109203X in WT or PKCe KO murine platelets correlate with an increase in the phosphorylation of Y311 on PKCd (Fig C) suggesting that ADP-induced thromboxane generation is regulated through PKCd Y311 phosphorylation. Tyrosine phosphorylation on PKCd is mediated by Src family kinases (SFKs) as the phosphorylation is abolished with PP2, a SFK inhibitor and is only partially inhibited in Fyn knockout murine platelets suggesting that other SFKs also mediate this tyrosine phosphorylation. Surprisingly, pre-incubation of platelets with LY-333531, a classical PKC isoform (a/b) inhibitor potentiated PKCd Y311 phosphorylation (Fig D) as well as thromboxane generation (Fig E) downstream of ADP receptors suggesting a role for classical PKCs. Also, platelets pre-incubated with LY-333531 showed a decrease in the phosphorylation of SHPTP-1 (Fig F), a tyrosine phosphatase, rendering it active. The active SHPTP-1 phosphatase may dephosphorylate and activate SFKs, which can now phosphorylate PKCd on Y311 in platelets. Conclusions: In the current study, we report for the first time that the novel PKC isoform d is tyrosine phosphorylated downstream of ADP receptors through which it mediates ADP-induced thromboxane generation. We also show a novel role for classical PKC isoforms a/b in regulating tyrosine phosphorylation on novel isoform, PKCd possibly through the tyrosine phosphatase SHPTP-1 and Src family kinases in platelets. Disclosures: No relevant conflicts of interest to declare.

and ADP receptors 3 β IIb α human platelets requires coordinated signaling through integrin induced thromboxane A2generation in - Adenosine diphosphate (ADP)

BF0801, a novel adenine derivative, inhibits platelet activation via phosphodiesterase inhibition and P2Y12 antagonism

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 ...

receptor-mediated platelet functional responses Role of G protein-gated inwardly-rectifying potassium channels in P2Y12

activation in collagen-related peptide-induced platelet θ Response:Role of PKC

Coagulation Factor XIIa Activates Platelets and Is the Physiological Agonist of Protease-Activated Receptor 3

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

Abstract 383: Cavβ2a Transgenic Mouse as a Hfpef Model With Hypercontractile Cardiomyocytes

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...

interaction with both extracellular cysteine residues, C17 and C270 Inactivation of the human P2Y12 receptor by thiol reagents requires

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)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:

protease-activated receptor-mediated Akt phosphorylation in platelets Relative contribution of G-protein-coupled pathways to

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...

Role of PKCθ in collagen-related peptide-induced platelet activation

Blood, 2009

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...

Tools and DRugs BF 0801 , a novel adenine derivative , inhibits platelet activation via phosphodiesterase inhibition and P 2 Y 12 antagonism

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

Antagonism of P2y12 Receptor Results in Severe Impairment of Serum Thromboxane Levels. Can We Manage Patients without Combination Therapy with Aspirin?

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...

Protein Kinase C Isoform ε Negatively Regulates ADP-Induced Thromboxane Generation by Regulating cPLA2 Activation and Calcium Mobilization In Platelets

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...