Antiplatelet activity of carnosol is mediated by the inhibition of TXA2 receptor and cytosolic calcium mobilization

We previously demonstrated that thromboxane A2 and/or prostaglandin H2 (TXA2/PGH2), ADP, and A23187 cause calcium mobilization in intact human platelets. Other studies have also shown that platelet shape change and aggregation induced by a variety of platelet agonists can be reversed by specific antagonists. In the present study, we used the fluorescent calcium probe chlortetracycline to evaluate whether the reversal of platelet activation involves a resequestration of intraplatelet calcium. It was found that the TXA2/PGH2 receptor antagonist 13-azaprostanoic acid (13-APA) reversed calcium mobilization and shape change induced by AA but not that induced by ADP. A similar specificity of action was observed using the specific ADP receptor antagonist, ATP, in that ATP only reversed ADP-induced calcium release and shape change. In contrast, prostacyclin reversed both AA and ADP-induced calcium redistribution and shape change. In the latter experiments, a net calcium sequestration was actually observed on prostacyclin addition. These findings indicate that the resequestration of released calcium leads to platelet deactivation. Furthermore, there appear to be at least two mechanisms by which a reduction in cytosolic calcium can be produced: specific interruption of the agonist-receptor interaction, for example, 13-APA antagonism of TXA2/PGH2; and stimulation of platelet adenosine 3',5'-cyclic monophosphate production by prostacyclin and consequent calcium sequestration.

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Characterization of platelet-activating factor-induced cytosolic calcium mobilization in human eosinophils

Clinical & Experimental Allergy ◽

10.1046/j.1365-2222.2000.00786.x ◽

2000 ◽

Vol 30 (5) ◽

pp. 699-705 ◽

Cited By ~ 14

Author(s):

Oshiro ◽

Kakuta ◽

Shimura ◽

Nara ◽

Shirato

Keyword(s):

Platelet Activating Factor ◽

Cytosolic Calcium ◽

Calcium Mobilization

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Increase in cytosolic calcium upregulates the synthesis of type 1 plasminogen activator inhibitor in the human histiocytic cell line U937

Blood ◽

10.1182/blood.v87.1.162.162 ◽

1996 ◽

Vol 87 (1) ◽

pp. 162-173 ◽

Cited By ~ 20

Author(s):

F Peiretti ◽

C Fossat ◽

F Anfosso ◽

MC Alessi ◽

M Henry ◽

...

Keyword(s):

Mrna Expression ◽

Plasminogen Activator Inhibitor ◽

Cytosolic Calcium ◽

Calcium Mobilization ◽

U937 Cells ◽

Histiocytic Cell ◽

Dose Dependent ◽

Activator Inhibitor ◽

Pai 1

Abstract In the U937 histiocytic cell line, we investigated the effect of calcium-mobilizing agents with or without tumor necrosis factor-alpha (TNF) on the regulation of the synthesis of plasminogen activator inhibitor-type 1 (PAI-1). Cultured U937 cells were stimulated with ionophore A23187 and thapsigargin with or without TNF. The response was analyzed in terms of cytosolic calcium mobilization, PAI-1 accumulation in the medium, and PAI-1 mRNA expression. The study was extended to urokinase (uPA) secretion and surface expression of its receptor (uPAR). Using Fluo-3 as a calcium-indicator dye to measure cytosolic calcium mobilization, we showed by flow cytometry that both agents mobilized calcium in a dose-dependent manner. TNF provoked a slight calcium mobilization that was also observed by digital imaging microscopy. Association of TNF with the calcium-mobilizing agents potentiated the calcium mobilization. Both calcium-mobilizing agents induced at 18 hours a dose-dependent accumulation of PAI-1 in culture medium, whereas uPA was not affected. TNF alone induced a more marked accumulation of PAI-1 than of uPA. Association of TNF with the agents induced a PAI-1 response that was more than additive of the two, whereas the secretion of uPA was not enhanced. Membrane expression of uPAR, measured by flow cytometry, tended to be slightly augmented by the calcium-mobilizing agents only. All the treatments resulted in a significant increase in PAI-1 mRNA level at 3 hours after the stimulation, which was very marked when calcium-mobilizing agents were present. Incubation of U937 cells in a calcium-free medium totally prevented both the mRNA expression and accumulation of PAI-1 induced by calcium-mobilizing agents and, to lesser extent, that induced by TNF. The increase in PAI-1 mRNA expression did not require de novo protein synthesis, as cycloheximide did not suppress the increase in PAI-1 mRNA induced by calcium-mobilizing agents. It is concluded that, in U937 cells, calcium triggers a pathway that upregulates PAI-1 synthesis and positively interacts with the TNF-induced pathway that stimulates PAI-1 synthesis. As uPA and uPAR were differently affected, it is suggested that an increase in cytosolic calcium leads to a reduced pericellular proteolysis.

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Autophagy inhibited Ehrlich ascitic tumor cells apoptosis induced by the nitrostyrene derivative compounds: Relationship with cytosolic calcium mobilization

European Journal of Pharmacology ◽

10.1016/j.ejphar.2011.12.031 ◽

2012 ◽

Vol 678 (1-3) ◽

pp. 6-14 ◽

Cited By ~ 14

Author(s):

Andrana K. Calgarotto ◽

Gustavo J. da Silva Pereira ◽

Alexandre Bechara ◽

Edgar J. Paredes-Gamero ◽

Christiano M.V. Barbosa ◽

...

Keyword(s):

Tumor Cells ◽

Cytosolic Calcium ◽

Calcium Mobilization

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Calcium mobilization and influx during the biphasic cytosolic calcium and secretory responses in agonist-stimulated pituitary gonadotrophs

Archives of Biochemistry and Biophysics ◽

10.1016/0003-9861(89)90388-3 ◽

1989 ◽

Vol 275 (2) ◽

pp. 410-428 ◽

Cited By ~ 24

Author(s):

Shun-Ichiro Izumi ◽

Stanko S. Stojilković ◽

Kevin J. Catt

Keyword(s):

Cytosolic Calcium ◽

Calcium Mobilization

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Increase in cytosolic calcium upregulates the synthesis of type 1 plasminogen activator inhibitor in the human histiocytic cell line U937

Blood ◽

10.1182/blood.v87.1.162.bloodjournal871162 ◽

1996 ◽

Vol 87 (1) ◽

pp. 162-173

Author(s):

F Peiretti ◽

C Fossat ◽

F Anfosso ◽

MC Alessi ◽

M Henry ◽

...

Keyword(s):

Mrna Expression ◽

Plasminogen Activator Inhibitor ◽

Cytosolic Calcium ◽

Calcium Mobilization ◽

U937 Cells ◽

Histiocytic Cell ◽

Dose Dependent ◽

Activator Inhibitor ◽

Pai 1

In the U937 histiocytic cell line, we investigated the effect of calcium-mobilizing agents with or without tumor necrosis factor-alpha (TNF) on the regulation of the synthesis of plasminogen activator inhibitor-type 1 (PAI-1). Cultured U937 cells were stimulated with ionophore A23187 and thapsigargin with or without TNF. The response was analyzed in terms of cytosolic calcium mobilization, PAI-1 accumulation in the medium, and PAI-1 mRNA expression. The study was extended to urokinase (uPA) secretion and surface expression of its receptor (uPAR). Using Fluo-3 as a calcium-indicator dye to measure cytosolic calcium mobilization, we showed by flow cytometry that both agents mobilized calcium in a dose-dependent manner. TNF provoked a slight calcium mobilization that was also observed by digital imaging microscopy. Association of TNF with the calcium-mobilizing agents potentiated the calcium mobilization. Both calcium-mobilizing agents induced at 18 hours a dose-dependent accumulation of PAI-1 in culture medium, whereas uPA was not affected. TNF alone induced a more marked accumulation of PAI-1 than of uPA. Association of TNF with the agents induced a PAI-1 response that was more than additive of the two, whereas the secretion of uPA was not enhanced. Membrane expression of uPAR, measured by flow cytometry, tended to be slightly augmented by the calcium-mobilizing agents only. All the treatments resulted in a significant increase in PAI-1 mRNA level at 3 hours after the stimulation, which was very marked when calcium-mobilizing agents were present. Incubation of U937 cells in a calcium-free medium totally prevented both the mRNA expression and accumulation of PAI-1 induced by calcium-mobilizing agents and, to lesser extent, that induced by TNF. The increase in PAI-1 mRNA expression did not require de novo protein synthesis, as cycloheximide did not suppress the increase in PAI-1 mRNA induced by calcium-mobilizing agents. It is concluded that, in U937 cells, calcium triggers a pathway that upregulates PAI-1 synthesis and positively interacts with the TNF-induced pathway that stimulates PAI-1 synthesis. As uPA and uPAR were differently affected, it is suggested that an increase in cytosolic calcium leads to a reduced pericellular proteolysis.

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The Kinase Inhibitor Sorafenib Induces Cell Death through a Process Involving Induction of Endoplasmic Reticulum Stress

Molecular and Cellular Biology ◽

10.1128/mcb.01080-06 ◽

2007 ◽

Vol 27 (15) ◽

pp. 5499-5513 ◽

Cited By ~ 147

Author(s):

Mohamed Rahmani ◽

Eric Maynard Davis ◽

Timothy Ryan Crabtree ◽

Joseph Reza Habibi ◽

Tri K. Nguyen ◽

...

Keyword(s):

Endoplasmic Reticulum ◽

Cell Death ◽

Er Stress ◽

Kinase Inhibitor ◽

Cytosolic Calcium ◽

Calcium Mobilization ◽

Initiation Factor ◽

Central Component ◽

Human Leukemia ◽

Eif2α Phosphorylation

ABSTRACT Sorafenib is a multikinase inhibitor that induces apoptosis in human leukemia and other malignant cells. Recently, we demonstrated that sorafenib diminishes Mcl-1 protein expression by inhibiting translation through a MEK1/2-ERK1/2 signaling-independent mechanism and that this phenomenon plays a key functional role in sorafenib-mediated lethality. Here, we report that inducible expression of constitutively active MEK1 fails to protect cells from sorafenib-mediated lethality, indicating that sorafenib-induced cell death is unrelated to MEK1/2-ERK1/2 pathway inactivation. Notably, treatment with sorafenib induced endoplasmic reticulum (ER) stress in human leukemia cells (U937) manifested by immediate cytosolic-calcium mobilization, GADD153 and GADD34 protein induction, PKR-like ER kinase (PERK) and eukaryotic initiation factor 2α (eIF2α) phosphorylation, XBP1 splicing, and a general reduction in protein synthesis as assessed by [35S]methionine incorporation. These events were accompanied by pronounced generation of reactive oxygen species through a mechanism dependent upon cytosolic-calcium mobilization and a significant decline in GRP78/Bip protein levels. Interestingly, enforced expression of IRE1α markedly reduced sorafenib-mediated apoptosis, whereas knockdown of IRE1α or XBP1, disruption of PERK activity, or inhibition of eIF2α phosphorylation enhanced sorafenib-mediated lethality. Finally, downregulation of caspase-2 or caspase-4 by small interfering RNA significantly diminished apoptosis induced by sorafenib. Together, these findings demonstrate that ER stress represents a central component of a MEK1/2-ERK1/2-independent cell death program triggered by sorafenib.

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Fibrinogen Binding Is Independent of an Increase in Intracellular Calcium Concentration in Thrombin Degranulated Platelets

Thrombosis and Haemostasis ◽

10.1055/s-0038-1653769 ◽

1995 ◽

Vol 73 (02) ◽

pp. 304-308 ◽

Cited By ~ 4

Author(s):

Fabio M Pulcinelli ◽

James L Daniel ◽

Silvia Riondino ◽

Pier Paolo Gazzaniga ◽

Leon Salganicoff

Keyword(s):

Platelet Aggregation ◽

Intracellular Calcium ◽

Binding Sites ◽

Calcium Concentration ◽

Calcium Release ◽

Shape Change ◽

Cytosolic Calcium ◽

Calcium Mobilization ◽

Induce Platelet Aggregation ◽

Fibrinogen Binding

SummaryIn a suspension of thrombin degranulated platelets (TDP), ADP and epinephrine can induce platelet aggregation, whereas the synthetic agonist of the thromboxane/endoperoxide receptor U46619 causes only shape change. However, U46619 can enhance platelet aggregation induced by ADP and epinephrine. In this paper, we have measured fibrinogen binding in relation to phospholipase C(PLC) activation and calcium mobilization in TDP activated by ADP, epinephrine and U46619.ADP caused fibrinogen binding in TDP but neither activated PLC nor caused a calcium mobilization. The requirement for ADP in inducing exposure of fibrinogen binding sites was not absolute since the combination of epinephrine and U46619 produced an increase in fibrinogen binding. U46619 caused significant PLC activation and cytosolic calcium release but not fibrinogen binding. These results suggest that in TDP the exposure of fibrinogen binding sites, after agonist activation, is independent of both PLC activation and calcium mobilization.

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