Aspirin Inhibits Platelet-Derived Sphingosine-1-Phosphate Induced Endothelial Cell Migration

Pharmacology ◽  
2017 ◽  
Vol 101 (1-2) ◽  
pp. 72-75 ◽  
Author(s):  
Amin Polzin ◽  
Betül Knoop ◽  
Andreas Böhm ◽  
Lisa Dannenberg ◽  
Mark Zurek ◽  
...  
Keyword(s):  
Cell Migration ◽  
Endothelial Cell Migration ◽  
Boyden Chamber ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Dependent Manner ◽  
S1p1 Receptor ◽  
Activated Platelets ◽  
Sphingosine 1 Phosphate ◽  
S1p Receptor

Background: Aspirin plays a crucial role in the prevention of cardiovascular diseases. We previously described that aspirin has effects beyond inhibition of platelet aggregation, as it inhibited thrombin-mediated release of sphingosine-1-phosphate (S1P) from human platelets. S1P is a bioactive lipid with important functions on inflammation and apoptosis. In endothelial cells (EC), S1P is a key regulator of cell migration. In this study, we aimed to analyze the effects of aspirin on platelet-induced EC migration. Methods: Human umbilical EC migration was measured by Boyden chamber assay. EC migration was induced by platelet supernatants of thrombin receptor-activating peptide-1 (AP1) stimulated platelets. To investigate the S1P receptor subtype that promotes EC migration, specific inhibitors of S1P receptor subtypes were applied. Results: S1P induced EC migration in a concentration-dependent manner. EC migration induced by AP1-stimulated platelet supernatants was reduced by aspirin. S1P1 receptor inhibition almost completely abolished EC migration induced by activated platelets. The inhibition of S1P2 or S1P3 receptor had no effect. Conclusion: Aspirin inhibits EC migration induced by activated platelets that is in part due to S1P and mediated by the endothelial S1P1 receptor. The clinical significance of this novel mechanism of aspirin action has to be investigated in future studies.

scholarly journals Sphingosine-1-phosphate receptor subtype-specific positive and negative regulation of Rac and haematogenous metastasis of melanoma cells

2003 ◽  
Vol 374 (3) ◽  
pp. 715-722 ◽  
Author(s):  
Hironori YAMAGUCHI ◽  
Joji KITAYAMA ◽  
Noriko TAKUWA ◽  
Kayo ARIKAWA ◽  
Isao INOKI ◽  
...  
Keyword(s):  
Cell Migration ◽  
G Protein ◽  
Lung Metastasis ◽  
Melanoma Cells ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Sphingosine 1 Phosphate ◽  
S1p Receptor ◽  
G Protein Coupled ◽  
Stimulation Of

We have recently reported that S1P (sphingosine-1-phosphate) differentially regulates cellular Rac activity and cell migration in either a positive or a negative direction via distinct G-protein-coupled receptor subtypes, i.e. S1P1/Edg1 (endothelial differentiation gene) and S1P2/Edg5 respectively, when each of the S1P receptor subtypes is expressed in CHO (Chinese-hamster ovary) cells. In B16F10 mouse melanoma cells, in which S1P2, but not the other S1P receptor subtypes, is endogenously expressed, S1P inhibited cell migration with concomitant inhibition of Rac and stimulation of RhoA in dose-dependent manners. Overexpression of S1P2 in the melanoma cells resulted in potentiation of S1P inhibition of both Rac and cell migration. In contrast, overexpression of S1P1 led to stimulation of cell migration, particularly at the lower S1P concentrations. Treatment of B16F10 cells with S1P inhibited lung metastasis 3 weeks after injection into mouse tail veins. Intriguingly, overexpression of S1P2 greatly potentiated the inhibition of metastasis by S1P, whereas that of S1P1 resulted in aggravation of metastasis. Suppression of cellular Rac activity by adenovirus-transduced expression of N17Rac, but not N19RhoA, strongly inhibited cell migration in vitro and lung metastasis in vivo. These results provide the first evidence that G-protein-coupled receptors could participate in the regulation of metastasis, in which ligand-dependent, subtype-specific regulation of the cellular Rac activity is probably critically involved as a mechanism.

ADAMTS13 Promotes Angiogenesis and Modulates VEGF-Mediated Angiogenic Activities

Blood ◽  
2011 ◽  
Vol 118 (21) ◽  
pp. 1145-1145
Author(s):  
Manfai Lee ◽  
Jonathan Baza ◽  
George M. Rodgers
Keyword(s):  
Cell Proliferation ◽  
Endothelial Cells ◽  
Cell Migration ◽  
Endothelial Cell ◽  
Endothelial Cell Migration ◽  
Molar Ratio ◽  
Assay Method ◽  
Boyden Chamber ◽  
Dependent Manner

Abstract Abstract 1145 Severe plasma ADAMTS13 deficiency results in the clinical disorder thrombotic thrombocytopenic purpura. However, other potential pathophysiological roles of ADAMTS13 in endothelial cell biology remain unexplored. To assess the possible role of ADAMTS13 in angiogenesis, cell proliferation and migration of human umbilical vein endothelial cells (HUVEC) were studied in vitro. ADAMTS13 was found to be a highly potent chemoattractant, and additionally was capable of neutralizing VEGF activity in two angiogenesis assays-cell proliferation and cell migration. In the Boyden chamber cell migration assay, treatment of endothelial cells with exogenous recombinant ADAMTS13 promoted cell migration in a dose-dependent manner, with 1 ng/mL increasing cell migration across a gelatinized polycarbonate membrane by 14-fold. In the same model, 5 ng/mL VEGF165 (molar ratio of ADAMTS13:VEGF165 = 1/19) only increased cell migration by 7 fold. A steady decrease in endothelial cell migration was observed when the concentration of ADAMTS13 exceeded 1 ng/mL (Figure 1). Coincubation of 30 ng/mL ADAMTS13 with 6.16 ng/mL VEGF165 (molar ratio of ADAMTS13/VEGF165 = 1.3/1) inhibited endothelial cell migration by 45% compared to VEGF alone (Figure 2). A second model using an in vitro scratch-wound assay confirmed the Boyden chamber data. Substitution of ADAMTS13 with ADAM17, an analog of ADAMTS13 without the thrombospondin domain reversed the inhibition of VEGF-mediated cell migration, suggesting that the thrombospondin domain of ADAMTS13 is responsible for the inhibitory interaction with VEGF165. This finding was in agreement with our previously published co-immunoprecipitation assay data (Blood 2010, 116, 4307). Similar patterns of inhibition were observed with VEGF121 and VEGF189, indicating that other isoforms of VEGF may interact with the TSP domain of ADAMTS13. Using a manual proliferation assay method, HUVEC treated with 30 ng/mL ADAMTS13 and 6.16 ng/mL VEGF165 proliferated 40% slower than the control treated with VEGF alone. Combined with our findings on the inhibition of endothelial cell-tube formation in a Matrigel assay with ADAMTS13 and VEGF165 previously reported, our cumulative data suggest that 1) ADAMTS13 promotes angiogenesis by increasing cell migration and 2) ADAMTS13 can modulate VEGF-mediated angiogenic activities. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Modulation of endothelial cell migration by extracellular nucleotides

2005 ◽  
Vol 93 (04) ◽  
pp. 735-742 ◽  
Author(s):  
Laurie Erb ◽  
Katarzyna Koziak ◽  
Robert Jarzyna ◽  
Marcia Wink ◽  
Olaf Guckelberger ◽  
...  
Keyword(s):  
Cell Migration ◽  
Endothelial Cell ◽  
P2 Receptors ◽  
Cellular Adhesion ◽  
Extracellular Nucleotides ◽  
Endothelial Cell Migration ◽  
Free Calcium ◽  
Receptor Subtypes ◽  
Dependent Manner ◽  
Cell Functions

SummaryExtracellular nucleotides bind to type-2 purinergic/pyrimidinergic (P2) receptors that mediate various responses, such as cell activation, proliferation and apoptosis, implicated in inflammatory processes. The role of P2 receptors and their associated signal transduction pathways in endothelial cell responses has not been fully investigated. Here, it is shown that stimulation of human umbilical vein endothelial cells (HUVEC) with extracellular ATP or UTP increased intracellular free calcium ion concentrations ([Ca2+]i), induced phosphorylation of focal adhesion kinase (FAK), p130cas and paxillin, and caused cytoskeletal rearrangements with consequent cell migration. Furthermore, UTP increased migration of HUVEC in a phosphatidylinositol 3-kinase (PI3-K)-dependent manner. BAPTA or thapsigargin inhibited the extracellular nucleotide-induced increase in [Ca2+]i, a response crucial for both FAK phosphorylation and cell migration. Furthermore, long-term exposure of HUVEC to ATP and UTP, agonists of the G protein-coupled P2Y2 and P2Y4 receptor subtypes, caused upregulation of αv integrin expression, a cell adhesion molecule known to directly interact with P2Y2 receptors. Our results suggest that extracellular nucleotides modulate signaling pathways in HUVEC influencing cell functions, such as cytoskeletal changes, cellular adhesion and motility, typically associated with integrin-activation and the action of growth factors. We propose that P2Y2 and possibly P2Y4 receptors mediate those responses that are important in vascular inflammation, atherosclerosis and angiogenesis.

scholarly journals Sphingosine 1-Phosphate (S1P) Receptor Subtypes S1P1and S1P3, Respectively, Regulate Lymphocyte Recirculation and Heart Rate

2004 ◽  
Vol 279 (14) ◽  
pp. 13839-13848 ◽  
Author(s):  
M. Germana Sanna ◽  
Jiayu Liao ◽  
Euijung Jo ◽  
Christopher Alfonso ◽  
Min-Young Ahn ◽  
...  
Keyword(s):  
Heart Rate ◽  
Receptor Subtypes ◽  
Sphingosine 1 Phosphate ◽  
S1p Receptor ◽  
Lymphocyte Recirculation

Urokinase-induced migration of human vascular smooth muscle cells requires coupling of the small GTPases RhoA and Rac1 to the Tyk2/PI3-K signalling pathway

2003 ◽  
Vol 89 (05) ◽  
pp. 904-914 ◽  
Author(s):  
Natalia Tkachuk ◽  
Hermann Haller ◽  
Inna Dumler ◽  
Ioulia Kiian
Keyword(s):  
Smooth Muscle ◽  
Cell Migration ◽  
Vascular Smooth Muscle ◽  
Kinase Inhibitor ◽  
Rho Kinase ◽  
Small Gtpases ◽  
Janus Kinase ◽  
Immunocytochemical Staining ◽  
Boyden Chamber ◽  
Dependent Manner

SummaryUrokinase-type plasminogen activator (uPA) facilitates cell migration by localizing proteolisys on the cell surface and by inducing intracellular signalling pathways. In human vascular smooth muscle cell (VSMC) uPA stimulates migration via the uPA receptor (uPAR) signalling complex containing the Janus kinase Tyk2 and phosphatidylinositol 3-kinase (PI3-K). We report that active GTP-bound forms of small GTPases RhoA and Rac1, but not Cdc42, are directly associated with Tyk2 and PI3-K in an uPA/uPAR-dependent fashion. Endogenous RhoA, but not Rac1 or Cdc42, was significantly activated in response to uPA. RhoA activation was abolished by cell treatment with two unrelated, structurally distinct, specific inhibitors of PI3-K, wortmannin, and LY294002. Downstream of RhoA, phosphorylation of myosin light chain (MLC) was dramatically upregulated by uPA in a Rho kinase- and PI3-K-dependent manner. Thus, selective Rho kinase inhibitor Y27632 and PI3-K inhibitors wortmannin and LY294002 prevented the uPA-induced stimulation of MLC phosphorylation. Rho kinase inhibition also decreased uPA-stimulated VSMC migration as observed in a Boyden chamber. VSMC immunocytochemical staining demonstrated redistribution of RhoA and Rac1 active forms to the newly formed leading edge of migrating cell. VSMC microinjection with antibodies to either Rho or Rac1 decreased uPA-stimulated cell migration, indicating the involvement of both GTPases in the migration process. Our results provide evidence that the small GTPases RhoA and Rac1, together with Rho kinase, are necessary to mediate the uPA/uPAR-directed migration via the Tyk2/PI3-K signalling complex in human VSMC.

Characterization of angiotensin receptors mediating prostaglandin synthesis in C6 glioma cells

1991 ◽  
Vol 260 (5) ◽  
pp. R1000-R1006 ◽  
Author(s):  
N. Jaiswal ◽  
D. I. Diz ◽  
E. A. Tallant ◽  
M. C. Khosla ◽  
C. M. Ferrario
Keyword(s):  
Glioma Cells ◽  
C6 Glioma ◽  
C6 Glioma Cells ◽  
Angiotensin Receptors ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Dependent Manner ◽  
Ang Ii ◽  
Selective Antagonist ◽  
Pge2 Release

The heptapeptide angiotensin (ANG)-(1-7) mimics some but not all the central actions of ANG II, suggesting that receptor subtypes may exist. The effects of ANG-(1-7), ANG II, and ANG I on prostaglandin (PG) E2 and prostacyclin (PGI2) synthesis were investigated in neurally derived rat C6 glioma cells. All three ANG peptides stimulated PG release in a dose-dependent manner with the order of potency ANG-(1-7) greater than ANG I greater than ANG II. PGE2 release induced by ANG-(1-7) (10(-7) M) was partially blocked by [Sar1,Ile8]ANG II (10(-6) M), [Sar1,Thr8]ANG II (10(-6) M), or the subtype 1 selective antagonist Du Pont 753 (10(-5) M) but not by the subtype 2 selective antagonist CGP 42112A (10(-7)-10(-5) M). PGI2 release was inhibited only by [Sar1,Thr8]ANG II. ANG II-induced PGE2 release was blocked by [Sar1,Thr8]ANG II (10(-6) M), [Sar1,Ile8]ANG II (10(-6) M), or Du Pont 753 (10(-7) M) but not by CGP 42112A (10(-7)-10(-5) M). In contrast, ANG II-induced PGI2 release was blocked by Du Pont 753 (10(-7) M) as well as [Sar1,Ile8]ANG II (10(-6) M) but not by [Sar1,Thr8]ANG II or CGP 42112A. Thus ANG II-stimulated PGE2 and PGI2 syntheses in C6 glioma cells are mediated via receptor subtype 1. ANG-(1-7)-induced PGE2 synthesis is also mediated via subtype 1 receptors; however, PGI2 release was blocked by [Sar1,Thr8]ANG II only.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Sphingosine Kinase as a Regulator of Calcium Entry through Autocrine Sphingosine 1-Phosphate Signaling in Thyroid FRTL-5 Cells

Endocrinology ◽  
2009 ◽  
Vol 150 (11) ◽  
pp. 5125-5134 ◽  
Author(s):  
Dan Gratschev ◽  
Christoffer Löf ◽  
Jari Heikkilä ◽  
Anders Björkbom ◽  
Pramod Sukumaran ◽  
...  
Keyword(s):  
Intracellular Signaling ◽  
Sphingosine Kinase ◽  
Calcium Entry ◽  
Dominant Negative ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Entry Pathway ◽  
Sphingosine 1 Phosphate ◽  
S1p Receptor ◽  
In Cells

Calcium entry is one of the main regulators of intracellular signaling. Here, we have described the importance of sphingosine, sphingosine kinase 1 (SK1), and sphingosine 1-phosphate (S1P) in regulating calcium entry in thyroid FRTL-5 cells. In cells incubated with the phosphatase inhibitor calyculin A, which evokes calcium entry without mobilizing sequestered intracellular calcium, sphingosine inhibited calcium entry in a concentration-dependent manner. Furthermore, inhibiting SK1 or the ATP-binding cassette ABCC1 multidrug transporter attenuated calcium entry. The addition of exogenous S1P restored calcium entry. Neither sphingosine nor inhibition of SK1 attenuated thapsigargin-evoked calcium entry. Blocking S1P receptor 2 or phospholipase C attenuated calcium entry, whereas blocking S1P receptor 3 did not. Overexpression of wild-type SK1, but not SK2, enhanced calyculin-evoked calcium entry compared with mock-transfected cells, whereas calcium entry was decreased in cells transfected with the dominant-negative G82D SK1 mutant. Exogenous S1P restored calcium entry in G82D cells. Our results suggest that the calcium entry pathway is blocked by sphingosine and that activation of SK1 and the production of S1P, through an autocrine mechanism, facilitate calcium entry through activation of S1P receptor 2. This is a novel mechanism by which the sphingosine-S1P rheostat regulates cellular calcium homeostasis.

Secreted phospholipase A2 induces vascular endothelial cell migration

Blood ◽  
2000 ◽  
Vol 96 (12) ◽  
pp. 3809-3815 ◽  
Author(s):  
Maria Teresa Rizzo ◽  
Elisabeth Nguyen ◽  
Marlene Aldo-Benson ◽  
Gerard Lambeau
Keyword(s):  
Cell Migration ◽  
Endothelial Cell ◽  
Synovial Fluid ◽  
Phospholipase A2 ◽  
Lipolytic Activity ◽  
Vascular Endothelial Cell ◽  
Endothelial Cell Migration ◽  
Boyden Chamber ◽  
Secreted Phospholipase A2 ◽  
Human Synovial Fluid

Abstract Secreted phospholipase A2 (sPLA2) regulates a variety of cellular functions. The present investigation was undertaken to elucidate the potential role of sPLA2 in endothelial cell (EC) migration. Bovine aortic endothelial cells (BAECs) exposed to sPLA2 placed in the lower compartment of a modified Boyden chamber displayed increased migration compared to cells exposed to vehicle. The effect of sPLA2 on EC migration was time and dose dependent. Migration of BAECs was observed at 30 minutes, increased over 1 to 2 hours, and declined thereafter. At 2 hours of stimulation, sPLA2 (0.01-2 μmol/L) induced 1.2- to 3-fold increased cell migration compared with media alone. Among the different sPLA2s tested, bee venom, Naja naja, and porcine and human pancreatic PLA2s all evoked a migratory response in ECs. Moreover, human synovial fluid, obtained from patients with arthritis and containing sPLA2 activity, induced EC migration. Migration of ECs was significantly reduced after exposure to a catalytic site mutant of pancreatic sPLA2with decreased lipolytic activity as compared to wild-type sPLA2. Similarly, pretreatment of human synovial fluid withp-bromophenacyl bromide, an irreversible inhibitor of sPLA2, markedly decreased the ability of human synovial fluid to stimulate EC migration. Moreover, migration of ECs was stimulated on exposure to hydrolytic products of sPLA2activity including arachidonic acid, lysophosphatidic acid, and lysophosphatidylcholine. These findings suggest that sPLA2plays a physiologic role in induction of EC migration. Moreover, the effects of sPLA2 on EC migration are mediated, at least in part, by its catalytic activity.

Secreted phospholipase A2 induces vascular endothelial cell migration

Blood ◽  
2000 ◽  
Vol 96 (12) ◽  
pp. 3809-3815
Author(s):  
Maria Teresa Rizzo ◽  
Elisabeth Nguyen ◽  
Marlene Aldo-Benson ◽  
Gerard Lambeau
Keyword(s):  
Cell Migration ◽  
Endothelial Cell ◽  
Synovial Fluid ◽  
Phospholipase A2 ◽  
Lipolytic Activity ◽  
Vascular Endothelial Cell ◽  
Endothelial Cell Migration ◽  
Boyden Chamber ◽  
Secreted Phospholipase A2 ◽  
Human Synovial Fluid

Secreted phospholipase A2 (sPLA2) regulates a variety of cellular functions. The present investigation was undertaken to elucidate the potential role of sPLA2 in endothelial cell (EC) migration. Bovine aortic endothelial cells (BAECs) exposed to sPLA2 placed in the lower compartment of a modified Boyden chamber displayed increased migration compared to cells exposed to vehicle. The effect of sPLA2 on EC migration was time and dose dependent. Migration of BAECs was observed at 30 minutes, increased over 1 to 2 hours, and declined thereafter. At 2 hours of stimulation, sPLA2 (0.01-2 μmol/L) induced 1.2- to 3-fold increased cell migration compared with media alone. Among the different sPLA2s tested, bee venom, Naja naja, and porcine and human pancreatic PLA2s all evoked a migratory response in ECs. Moreover, human synovial fluid, obtained from patients with arthritis and containing sPLA2 activity, induced EC migration. Migration of ECs was significantly reduced after exposure to a catalytic site mutant of pancreatic sPLA2with decreased lipolytic activity as compared to wild-type sPLA2. Similarly, pretreatment of human synovial fluid withp-bromophenacyl bromide, an irreversible inhibitor of sPLA2, markedly decreased the ability of human synovial fluid to stimulate EC migration. Moreover, migration of ECs was stimulated on exposure to hydrolytic products of sPLA2activity including arachidonic acid, lysophosphatidic acid, and lysophosphatidylcholine. These findings suggest that sPLA2plays a physiologic role in induction of EC migration. Moreover, the effects of sPLA2 on EC migration are mediated, at least in part, by its catalytic activity.

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