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Abstract
Prior studies have shown that PI3Ks play a necessary but incompletely defined role in platelet activation. One potential effector for PI3K is the serine/threonine kinase, Akt, whose contribution to platelet activation was explored here. Two isoforms of Akt were detected in mouse platelets, with expression of Akt2 being greater than Akt1. Deletion of the gene encoding Akt2 impaired platelet aggregation, fibrinogen binding, and granule secretion, especially in response to low concentrations of agonists that activate the Gq-coupled receptors for thrombin and thromboxane A2. Loss of Akt2 also impaired arterial thrombus formation and stability in vivo, despite having little effect on platelet responses to collagen and ADP. In contrast, reducing Akt1 expression had no effect except when Akt2 was also deleted. Activation of Akt by thrombin was abolished by deletion of Gαq but was relatively unaffected by deletion of Gαi2, which abolished Akt activation by ADP. From these results we conclude that Akt2 is a necessary component of PI3K-dependent signaling downstream of Gq-coupled receptors, promoting thrombus growth and stability in part by supporting secretion. The contribution of Akt1 is less evident except in the setting in which Akt2 is absent.
Authors
Donna Woulfe, Hong Jiang, Alicia Morgans, Robert Monks, Morris Birnbaum, Lawrence F. Brass
PLATELET SEROTONIN AND THROMBOSTASIS
Submitter: Fuad Lechin | flechin@telcel.net.ve
Universidad Central de Venezuela
Published April 21, 2004
The recent publication by Woulfe et al. in this journal (113: 441- 450, 2004) report findings that show the effects on secretion, aggregation, and thrombus formation in platelets of mice lacking Akt2. In addition to it, our studies showed that diseases associated with defects on thrombus formation such as idiopathic thrombocytopenic purpura (ITP) reveal very low levels of p-5HT (1). This haematological disorder is worsened by drugs that deplete platelet serotonin content (inhibitors of serotonin uptake and serotonin releasing agents) (2,3). Conversely, we found that tianeptine, a drug that enhances p-5HT, improves thrombostasis and stops bleeding in ITP patients (1). Our findings and those of other authors demonstrate the primary role played by p-5HT in the haemostatic process (4). Further, it has been postulated that serotonin. release from platelets is the most reliable test of platelet activation (5). Furthermore, we demonstrated that platelet serotonin (p-5HT) content rather than platelet count should be considered as the appropriate index for measuring thrombostasis as the former parameter and not the later correlates with bleeding stoppage in ITP patients during relapse periods (1). Similar findings have been reported by other authors measuring these parameters in a experimental model of ITP in mice (6). Our findings are consistent with the demonstrated fact that serotonin is stored in the delta granules of platelets and its release increases during their activation (5). Stimulated platelets use serotonin to enhance their retention of procoagulant proteins on the cell surface (4). Furthermore, the presence of serotonin is covalently linked to fibrinogen bound on the surface of the activated platelet where it increases the retention of procoagulant factors on the cell surface. Serotonin, therefore, plays a central role in haemostatic process and its release is considered the most reliable assay for platelet activation. The finding by Woulfe et al. showing that platelets from Aktl+/-Akt2-/-mice; the release of (3H)5-HT stimulated by AYPGQV was reduced nearly 50% compared with WT control animals, is also consistent with our findings. Furthermore, we found that drugs which increase platelet serotonin content, such as tianeptine (1) are able to stop bleeding in ITP patients. Conversely, we quoted three ITP patients who reported bleeding within the first 3-4 weeks of antidepressant therapy with sertraline, paroxetine and fluoxetine (serotonin uptake inhibitors), respectively, when the p-5HT content reached minimal values: (< 30 ng/ml - normal values = 255 + 25 ng/ml) (3). These patients showed bleeding stoppage within 1-3 days of tianeptine administration (1); their p-5HT increased to 85 ng/ml, 78 ng/ml and 76 ng/ml, respectively. Tianeptine was also able to stop bleeding in five pulmonary hypertension patients who showed low p-5HT + high f-5HT (2,3).
References
1. Lechin, F., van der Dijs, B., Orozco, B., Jahn, E., Rodríguez, S., and Baez, S. 2004 (June). Neuropharmacological treatment of refractory idiopathic thrombocytopenic purpura: Roles of circulating catecholamines and serotonin. Thromb Haemost.. In press.
2. Lechin, F., and van der Dijs, B. 2002. Serotonin and pulmonary vasoconstriction. J. Appl. Physiol. 92:1363-1364.
3. Lechin, F., van der Dijs, B., and Lechin, M.E. 2002. Depression. In: Neurocircuitry and Neuroautonomic Disorders. Reviews and Therapeutic Strategies. F. Lechin, B. van der Dijs and M.E. Lechin, editors. Karger, Basel, Switzerland. 23-27.
4. Dale, G.L., Friese, P., Batar, P., Hamilton, S.F., Reed, G.L., Jackson, K.W., Clemetson, K.J., and Alberio, L. 2002. Stimulated platelets use serotonin to enhance their retention of procoagulant proteins on the cell surface. Science. 415:175–179.
5. Gobbi, G., Mirandola, P., Tazzari, P.L., Ricci, F., Caimi, L., Cacchioli, A., Papa, S., Conte, R., and Vitale, M. 2003. Flow cytometry detection of serotonin content and release in resting and activated platelets. Br. J. Haematol. 121:892-896.
6. Dominguez, V., Govezensky, T., Gevorkian, G., and Larralde, C. 2003. Low platelet counts do not cause bleeding in an experimental model of immune thrombocytopenic purpura in mice. Haematologica. 88:679-687.
Fuad Lechin, MD, PhD, Emeritus Professor;
Bertha van der Dijs, MD;
Department of Physiological Sciences, Sections of Neurochemical, Neurophysiology, Neuroimmunology and Neuropharmacology, Instituto de Medicina Experimental,
Universidad Central de Venezuela, Caracas, Venezuela.
E.Mail: flechin@telcel.net.ve
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ISSN: 0021-9738 (print), 1558-8238 (online)