Abstract 17878: Kindlin-2 is a Novel Regulator of Hemostasis
Kindlin-2, a widely distributed cytoskeletal protein, has been implicated in integrin activation, and its absence is embryonically lethal in mice. We previously demonstrated that kindlin-2 is a key contributor to angiogenesis since it was defective in kindlin-2+/- mice and zebrafish. In the present study, we tested the hypothesis that kindlin-2 may regulate hemostasis utilizing the kindlin-2+/- mice. Tail bleeding time was significantly prolonged (+ 45%, P=0.02) and blood loss was 4-fold more (P=0.005) in kindlin-2+/- mice as compared to WT animals. In addition, time to carotid artery occlusion after FeCl3 injury was increased by 30% (P=0.02) in the kindlin-2+/- mice. Fibrinogen, vWF, t-PA, TFPI levels or activities were similar in plasma of WT and kindlin-2+/- mice. Platelet count, platelet expression levels of integrin αIIbβ3, P-selectin and GPIbα as well as ex vivo platelet aggregation in response to various agonists were similar in both mouse strains. Bone marrow transplantation from kindlin-2+/- donors to WT mice did not transfer the bleeding prolongation to recipients, indicating the defect was not associated with bone marrow-derived cells in kindlin-2+/- mice. However, mouse aortic endothelial cells (ECs) isolated from the kindlin-2+/- mice showed significantly enhanced capacity to inhibit ADP-induced platelet aggregation as compared to WT ECs (75% platelet aggregation in the presence of WT ECs vs 20% aggregation in the presence of kindlin-2+/- ECs). Although eNOS expression and phosphorylation were similar in both WT and kindlin-2+/- ECs, FACS analyses revealed increased (by 2-3-fold) levels of ATP diphosphohydrolase (CD39) and Ecto-5’-Nucleotidase (CD73) on the surface of the kindlin-2+/- ECs. Accordingly, 2-3 fold more ADP and AMP were hydrolyzed on kindlin-2+/- ECs than on WT ECs, and supernatants collected from ADP-treated kindlin-2+/- ECs did not trigger platelet aggregation as efficiently as those from WT cells. In control experiments, overexpression of kindlin-2 in the kindlin-2+/- EC, decreased CD39 and CD73 levels and blunted the antiplatelet activities of these cells. Taken together, endothelial kindlin-2 regulates ADP levels and ADP-dependent platelet responses and, thereby, plays an important role in thromboregulation.
- © 2011 by American Heart Association, Inc.