The CD40-CD40L interaction, which was initially shown to have important roles in the T cell-mediated activation of B cells during humoral immune responses, is now known to have roles in activation of endothelial cells, smooth muscle cells, and macrophages within atherosclerotic plaques. Recently, CD40L expression was found in activated platelets in the thrombus in vivo and CD40L was reported to be responsible for the platelet-mediated activation of endothelial cells in vitro. To investigate the activation status of platelets in coronary artery disease patients, we tested expression levels of CD40L, and platelet-endothelial cell adhesion molecule-1 (PECAM-1/CD31) in platelets isolated from peripheral blood, using flow cytometric analysis. Twenty-nine patients with acute coronary syndrome (10 acute myocardial infarction and 19 unstable angina patients) were compared with 14 normal subjects or 14 stable angina patients. In platelets isolated from normal subjects, the expression of CD40L was not detected in all subjects. In the patients with acute coronary syndrome, the average level of CD40L showed a significant increase (p = 0.0028), while stable angina patients did not have any increase when compared to normal subjects. Patients with more complex lesions or vessel occlusion tended to have a high platelet CD40L level compared to patients who do not. The expression levels of CD31 were increased in a small portion of the ACS patients. These data indicate that the rupture of plaque and subsequent formation of thrombus may lead to the activation of CD40L expression in circulating platelets of ACS patients.