Atherosclerosis is characterized by chronic inflammation of the arterial wall due to chemokine-driven mononuclear cell recruitment. Activated platelets can synergize with chemokines to exacerbate atherogenesis; for example, by deposition of the chemokines platelet factor-4 (PF4, also known as CXCL4) and RANTES (CCL5), triggering monocyte arrest on inflamed endothelium. Homo-oligomerization is required for the recruitment functions of CCL5, and chemokine heteromerization has more recently emerged as an additional regulatory mechanism, as evidenced by a mutual modulation of CXCL8 and CXCL4 activities and by enhanced monocyte arrest resulting from CCL5-CXCL4 interactions. The CCL5 antagonist Met-RANTES reduces diet-induced atherosclerosis; however, CCL5 antagonism may not be therapeutically feasible, as suggested by studies using Ccl5-deficient mice which imply that direct CCL5 blockade would severely compromise systemic immune responses, delay macrophage-mediated viral clearance and impair normal T cell functions. Here we determined structural features of CCL5-CXCL4 heteromers and designed stable peptide inhibitors that specifically disrupt proinflammatory CCL5-CXCL4 interactions, thereby attenuating monocyte recruitment and reducing atherosclerosis without the aforementioned side effects. These results establish the in vivo relevance of chemokine heteromers and show the potential of targeting heteromer formation to achieve therapeutic effects.

The aim of this study was to explore further the hypothesis that early stages of normal human hematopoiesis might be coregulated by autocrine/paracrine regulatory loops and by cross-talk among early hematopoi-

Heparin-induced thrombocytopenia (HIT)antibodies recognize complexes between heparin and platelet factor 4 (PF4). Heparin and PF4 bind HIT antibodies only over a narrow molar ratio. We explored the involvement of platelet surface-bound PF4 as an antigen in the pathogenesis of experimental HIT. We show that cellsurface PF4 complexes are also antigenic only over a restricted concentration range of PF4. Heparin is not required for HIT antibody binding but shifts the concentration of PF4 needed for optimal surface antigenicity to higher levels. These data are supported by in vitro studies involving both human and murine platelets with exogenous recombinant human (h) PF4 and either an anti-PF4-heparin monoclonal antibody (KKO) or HIT immunoglobulin. Injection of KKO into transgenic mice expressing different levels of hPF4 demonstrates a correlation between the severity of the thrombocytopenia and platelet hPF4 expression. Therapeutic interventions in this model using high-dose heparin or protamine sulfate support the pathogenic role of surface PF4 antigenic complexes in the etiology of HIT. We believe that this focus on surface PF4 advances our understanding of the pathogenesis of HIT, suggests ways to identify patients at high risk to develop HIT upon heparin exposure, and offers new therapeutic strategies. IntroductionHeparin-induced thrombocytopenia (HIT) is an iatrogenic complication of heparin therapy caused by antibodies that recognize complexes formed between heparin and the endogenous protein platelet factor 4 (PF4). [1][2][3] Approximately half of affected patients develop limb-or life-threatening thrombosis. [4][5][6] Management involves careful monitoring of platelet counts, a high index of clinical suspicion, cessation of heparin exposure, and the introduction of alternative anticoagulants. 7,8 These measures have reduced the incidence of new thromboembolic complications but have had less impact on the incidence of amputations and death. 9,10 Heparin remains an important anticoagulant in widespread use, and studies that help define the pathophysiology of HIT may lead to better identification of patients at risk and to more targeted intervention strategies.The antibody response in HIT is unusual in several respects. First, the major complications of HIT are related to thrombosis in contrast to other drug-induced thrombocytopenias. 11 This high incidence of thrombosis may be related in part to the ability of HIT antibodies to activate platelets via Fc␥RIIA. 12,13 In a murine model of HIT, only mice with platelets that expressed both human (h) PF4 and Fc␥RIIA developed thrombocytopenia and thrombosis when given an anti-PF4-heparin monoclonal antibody (mAb), KKO. 14 A second unusual feature is the surprisingly high incidence of anti-PF4-heparin antibodies in heparinized patients, exceeding a quarter to half of all exposed patients in some settings. [15][16][17] Why only a small portion of these patients develop HIT is not clear and no unequivocal differences between the vast majority of individuals who remai...