The generation of reactive oxygen species (ROS) represents a pivotal element of phagocyte defense against microbial invaders. However, oxidative stress also participates in pathophysiological processes of vascular damage leading to cell death of endothelial cells (EC). Currently, ROS-producing cells involved in this process as well as the corresponding extracellular signals required for their activation are ill-defined. In this study, we investigate the impact of the platelet-derived CXC chemokine platelet factor 4 (PF4/CXCL4) on the interaction of human monocytes and EC. We can show for the first time that PF4-activated monocytes become cytotoxic for EC but not epithelial cells. Cytotoxicity was time- and dose-dependent, and earliest effects were seen after 15 h of culture and at a concentration from 0.125 microM PF4 up. By performing transwell experiments and by using specific inhibitory antibodies, we could show that direct cell contact between effector and target cells, mediated by beta(2)integrins as well as their corresponding ligand ICAM-1, is essential for the cytotoxic effect. Investigations of the cellular mechanisms of cytotoxicity revealed that in the presence of EC, PF4-activated monocytes are capable of releasing high amounts of ROS for more than 2 h following stimulation. This causes programmed cell death in EC, as inhibitors of the NADPH oxidase (diphenyleneiodonium and apocynin) effectively blocked PF4-induced monocyte oxidative burst and protected EC from undergoing apoptosis. Taken together, our data suggest a role for platelet-derived PF4 in oxidative stress-mediated vascular disorders, as observed during atherosclerosis or ischemia/reperfusion injury.

Directed migration of cells along a chemotactic gradient is a fundamental cellular process involved e.g. in host defense, tissue development, and wound repair. Surprisingly, although a broad spectrum of different mediators are able to induce a chemotactic response, only very few regulators or inhibitors of this function are known. CXCL4 (platelet factor 4; PF4), a platelet-derived CXCchemokine, modulates long-term immunregulatory functions in T cells but lacks the capacity to induce chemotaxis in these cells. However, in the current study we are able to show for the first time, that CXCL4 acts as a potent inhibitor of T cell chemotaxis induced by CXCR3 ligands CXCL11 and CXCL9, but not CXCL10. CXCL4 did neither interfere with ligand binding to CXCR3 nor with ligandinduced internalization or calcium signaling of CXCR3. By several lines of evidence we could rule out the participation of known CXCL4-receptors, like proteoglycans or CXCR3-B, in CXCL4-mediated inhibition of chemotaxis. We, thus, claim the presence of a further, so far unidentified receptor for this chemokine, which is present on T cells. Intriguingly, CXCL4 also reduces the chemotaxis of T cells and neutrophils induced by the CXCR3-independent ligand CXCL12 (SDF-1alpha). Taken together, our results identify CXCL4 as the first chemokine, which acts as an inhibitor rather than an inducer of chemotaxis on T cells and neutrophils in vitro.