P146	The JAK-STAT-signaling pathway is involved in platelet activation by leptin.
1C.Dellas, 1K.Schäfer, 2D.J.Loskutoff, 1S.V.Konstantinides
1Georg-August-Universität, Göttingen, DE; 2The Scripps Research Institute, La Jolla, US.

We and others have shown that the adipokine leptin promotes platelet aggregation and enhances the thrombotic response to vascular injury. In the present study, we investigated the binding of leptin to its receptor (ObR) on human platelets, and the pathways mediating the effects of the hormone on platelet activation. Leptin binding to human platelets was investigated using flow cytometry analysis and confocal microsopy. FITC-labelled leptin (300 nM) bound specifically to resting platelets, and binding was inhibited by preincubation with 1 µM soluble ObR. Importantly, activation of platelets with 0.5 U/mL thrombin or other agonists induced a rapid and significant increase in leptin binding (93±2% of activated vs. 41±6% of non-activated platelets were leptin-positive; P<0.01; 6 donors). The expression of ObR on platelets was confirmed at the mRNA and protein level by PCR and western blotting (WB). The detected long isoform of the ObR was highly glycosylated with a MW of 200 kDa, which was reduced to 132 kDa after deglycosylation. To begin to identify intracellular signaling molecules downstream of the receptor, platelets were stimulated by leptin and lysates were subject to immunoprecipitation and WB. Leptin time- and dose-dependently induced tyrosine phosphorylation of several proteins, including the long isoform of ObR, within 10 seconds after stimulation. The highest levels of phosphorylation (3-fold increase compared to non-stimulated control platelets) were observed 2 minutes after preincubation with 10 ng/mL leptin. This effect was specific and could be inhibited by the tyrosine kinase inhibitor genistein. We could further demonstrate leptin-induced phosphorylation of the signaling molecules JAK2 and STAT3 (2.1-fold and 1.9-fold increase, respectively). These latter effects were prevented by preincubation with the JAK2-inhibitor AG490. Thus, leptin specifically binds to its receptor on resting and, particularly, activated platelets. This effect rapidly induces phosphorylation of signaling molecules including those of the JAK-STAT pathway. Dissecting the intracellular pathways involved in leptin signaling may help improve our understanding of the link between the hyperleptinemia and the increased cardiovascular risk in obesity.

Copyright © 2005 S. Karger AG, Basel. Any further use of this abstract requires written permission from the publisher.