Involvement of Calcium and G Proteins in the Acute Release of Tissue-Type Plasminogen Activator and von Willebrand Factor From Cultured Human Endothelial Cells

Eijnden-Schrauwen, Y. van den; Atsma, Douwe E.; Lupu, Florea; Vries, R.E.M. de; Kooistra, Teake; Emeis, J.J.

doi:10.1161/01.atv.17.10.2177

Cited by 54 publications

(50 citation statements)

References 53 publications

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“…Differential pathways of VWF and t-PA release in HUVECs with respect to pertussis toxin sensitivity and levels of intracellular Ca 2ϩ have been described. 37 These functional differences may have a structural explanation, because it has been observed that t-PA is stored in small dense vesicles distinct from WPBs. 38 However, this observation is controversial because other studies have shown that t-PA is stored in WPBs in HUVECs.…”

Section: Selective Release Of Weibel-palade Bodies That Contain Predomentioning

confidence: 99%

Differential regulation of endothelial exocytosis of P-selectin and von Willebrand factor by protease-activated receptors and cAMP

Cleator

Zhu

Vaughan

et al. 2006

Blood

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Thrombin-mediated endothelial-cell release of von Willebrand factor (VWF) and P-selectin functionally links proteaseactivated receptors (PARs) to thrombosis and inflammation. VWF release can be stimulated by both Ca 2؉ and cAMP, and, although both VWF and P-selectin are found in Weibel-Palade bodies (WPBs), we found that their release could be differentially regulated. In these studies, human umbilical vein endothelial cells stimulated with cAMP or PAR2-AP led to a delayed release of VWF and significantly less P-selectin release compared with histamine, thrombin, or PAR1-AP. Doseresponse studies revealed that PAR2-AP was significantly less efficacious in promoting the release of P-selectin compared with VWF. PAR2-AP-induced robust stimulation of intracellular Ca 2؉ coupled with a significantly greater inhibitory effect of calcium chelation on release of VWF compared with cell-surface expression of P-selectin, suggests an additional Ca 2؉ -independent pathway involved in release of P-selectin. PAR2-AP failed to increase global cAMP levels; however, inhibition of protein kinase A

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Section: Selective Release Of Weibel-palade Bodies That Contain Predomentioning

confidence: 99%

Differential regulation of endothelial exocytosis of P-selectin and von Willebrand factor by protease-activated receptors and cAMP

Cleator

Zhu

Vaughan

et al. 2006

Blood

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“…low pH) to avoid preanalytical complex binding with PAI-1 [44]. Regulated release of t-PA from endothelial cells can be induced by a number of substances with the common denominator that they activate G-protein-coupled cell surface receptors [45]. Several products formed during thrombus formation, i.e.…”

Section: Regulation Of Intravascular Fibrinolysis In Vivomentioning

confidence: 99%

Inhibition of carboxypeptidase U (TAFIa) activity improves rt-PA induced thrombolysis in a dog model of coronary artery thrombosis

Björkman

Abrahamsson

Nerme

et al. 2005

Thrombosis Research

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The physiologically most important activator of intravascular fibrinolysis is tissue-type plasminogen activator (t-PA) released from endothelial cells. In man, sympathomimetic drugs increase the systemic concentration of t-PA. It is therefore of interest to investigate whether cardiac sympathetic activation can induce a local t-PA release, which could counteract intra-coronary clot formation.Thrombolytic therapy with recombinant t-PA (rt-PA) is effective in acute myocardial infarction, but the treatment is limited by a fairly slow reperfusion rate and frequent early reocclusions. A potential mechanism behind early reocclusions might be that active thrombin is released from the thrombus during thrombolytic therapy. Thrombin has recently been shown to activate procarboxypeptidase U, which in its active form (CPU) down-regulates endogenous fibrinolysis. Therefore, one way of improving thrombolytic efficacy may be to combine rt-PA with a lowmolecular weight direct thrombin inhibitor, which theoretically could have a pro-fibrinolytic effect, either by inhibition of fibrin-bound thrombin and/or by inhibition of CPU activation. An alternative way may be direct inhibition of CPU.In a porcine model, experimental activation of cardiac sympathetic nerves by electrical stimulation at 1 and 8 Hz induced 5-and 20-fold increase in the release of both total and active t-PA together with frequency-dependent increases in heart rate, blood pressure, and coronary blood flow. The t-PA release was independent of the heart rate and coronary flow response, but local infusion of isoprenaline suggested that part of the t-PA response was mediated by stimulation of β-adrenergic receptors. Next, we studied the combined effect of rt-PA and thrombin inhibitors (melagatran, hirudin and heparin) in a canine model of copper coil-induced coronary thrombosis. The profibrinolytic effect of rt-PA, either measured as patency rate or time-to-patency, was significantly enhanced with the low-molecular weight direct thrombin inhibitor melagatran, but to a lesser degree by hirudin and heparin. In the same model it was shown that active CPU is produced locally in the coronary vascular bed during both thrombus formation and clot lysis. Inhibition of thrombin attenuated CPU formation and improved patency. A similar effect was obtained with a direct inhibitor of CPU.In conclusion, the coronary t-PA response to sympathetic stimulation may constitute a thromboprotective defence mechanism to counteract its prothrombotic effects on the systemic level. Furthermore, direct thrombin and/or CPU inhibition may be potential targets for prevention of thrombus formation via facilitation of the endogenous fibrinolytic system.

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“…15,16 Inhibition studies have shown that intracellular Ca 2ϩ exerts its effect on regulated secretion of vWF via calmodulin. 13,15 …”

mentioning

confidence: 99%

“…10 -12 The mechanism of thrombin-induced exocytosis of Weibel-Palade bodies has been only partially elucidated. [13][14][15] Thrombin induces the elevation of intracellular Ca 2ϩ levels, which appears crucial for the release of vWF from WeibelPalade bodies. 15,16 Inhibition studies have shown that intracellular Ca 2ϩ exerts its effect on regulated secretion of vWF via calmodulin.…”

mentioning

confidence: 99%

Small GTP-Binding Protein Ral Modulates Regulated Exocytosis of von Willebrand Factor by Endothelial Cells

Leeuw

Fernandez‐Borja

Reits

et al. 2001

ATVB

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Abstract-Weibel-Palade bodies are endothelial cell-specific organelles, which contain von Willebrand factor (vWF), P-selectin, and several other proteins. Recently, we found that the small GTP-binding protein Ral is present in a subcellular fraction containing Weibel-Palade bodies. In the present study, we investigated whether Ral is involved in the regulated exocytosis of Weibel-Palade bodies. Activation of endothelial cells by thrombin resulted in transient cycling of Ral from its inactive GDP-bound to its active GTP-bound state, which coincided with release of vWF. Ral activation and exocytosis of Weibel-Palade bodies were inhibited by incubation with trifluoperazine, an inhibitor of calmodulin, before thrombin stimulation. 3,4 A number of other components have been identified in Weibel-Palade bodies; these include P-selectin, CD63, endothelin, and interleukin-8. [5][6][7][8][9] On stimulation with agonists such as thrombin and histamine, Weibel-Palade bodies release their contents into the blood. 10 -12 The mechanism of thrombin-induced exocytosis of Weibel-Palade bodies has been only partially elucidated. [13][14][15] Thrombin induces the elevation of intracellular Ca 2ϩ levels, which appears crucial for the release of vWF from WeibelPalade bodies. 15,16 Inhibition studies have shown that intracellular Ca 2ϩ exerts its effect on regulated secretion of vWF via calmodulin. 13,15 See coverLittle attention has been directed at involvement of small GTPases in regulated secretion in endothelial cells. Small GTPases cycle between an active GTP-bound and inactive GDP-bound form. Guanine nucleotide exchange factors enhance the conversion from the inactive GDP-bound to the active GTP-bound form, whereas GTPase activating proteins promote the GTP hydrolysis of small GTPases. In many cells, small GTP-binding proteins have been implicated in regulated exocytosis, as exemplified by the pivotal role of Rab3A in the release of synaptic vesicles at the nerve terminal. 17 Therefore, it seems likely that small GTP-binding proteins are also involved in the release of vWF through the regulated pathway in endothelial cells. Recently, we identified the small GTP-binding protein Ral in a subcellular fraction containing Weibel-Palade bodies, suggesting a role for this GTPase in regulated exocytosis of these organelles. 18 Ral is a geranylgeranylated GTPase that is ubiquitously expressed. 19 Activated GTP-bound Ral binds to RLIP76 (or Ral binding protein), an effector molecule that possesses GTPase activity for cdc42 and Rac, suggesting a link between the activation of Ral and rearrangement of the cytoskeleton. 20 Morphological studies have identified Ral on dense granules in platelets and on synaptic vesicles in nerve terminals, suggesting a role for Ral in regulated exocytosis. 21,22 Interestingly, Ral has been proposed to interact with calmodulin in a Ca 2ϩ -dependent manner. 23 Binding to calmodulin enhances GTP binding to Ral 2-to 3-fold. 24 These observations may suggest a regulatory role for Ral in the calmodulin-mediated ...

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Involvement of Calcium and G Proteins in the Acute Release of Tissue-Type Plasminogen Activator and von Willebrand Factor From Cultured Human Endothelial Cells

Cited by 54 publications

References 53 publications

Differential regulation of endothelial exocytosis of P-selectin and von Willebrand factor by protease-activated receptors and cAMP

Differential regulation of endothelial exocytosis of P-selectin and von Willebrand factor by protease-activated receptors and cAMP

Inhibition of carboxypeptidase U (TAFIa) activity improves rt-PA induced thrombolysis in a dog model of coronary artery thrombosis

Small GTP-Binding Protein Ral Modulates Regulated Exocytosis of von Willebrand Factor by Endothelial Cells

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