Monocyte-bound PF4 in the pathogenesis of heparin-induced thrombocytopenia

Abstract

Heparin-induced thrombocytopenia (HIT)is a life-and limb-threatening thrombotic disorder that develops after exposure to heparin, often in the setting of inflammation. We have shown previously that HIT is associated with antibodies to complexes that form between platelet factor 4 and glycosaminoglycan (GAG) side chains on the surface of platelets. However, thrombosis can occur in the absence of thrombocytopenia. We now show that platelet factor 4 binds to monocytes and forms antigenic complexes with their surface GAG side chains more efficiently than on platelets likely due to differences in GAG composition. Binding to monocytes is enhanced when the cells are activated by endotoxin. Monocyte accumulation within developing arteriolar thrombi was visualized by situ microscopy. Monocyte depletion or inactivation in vivo attenuates thrombus formation induced by photochemical injury of the carotid artery in a modified murine model of HIT while paradoxically exacerbating thrombocytopenia. These studies demonstrate a previously unappreciated role for monocytes in the pathogenesis of arterial thrombosis in HIT and suggest that therapies targeting these cells might provide an alternative approach to help limit thrombosis in this and possibly other thrombotic disorders that occur in the setting of inflammation. IntroductionPlatelet factor 4 (PF4) is a cationic chemokine with high affinity for unfractionated heparin (UFH) and other large, negatively charged molecules. 1 PF4 is stored in platelet ␣-granules, released upon activation, when it then binds rapidly to glycosaminoglycan (GAG) side chains expressed on the surface of platelets 2 and other vascular cells, with little remaining free in the circulation. 3 Heparin-induced thrombocytopenia (HIT) is an iatrogenic complication of heparin therapy caused by antibodies that recognize complexes of human (h) PF4 with heparin or other GAGs. 4,5 In solution, formation of antigenic complexes between PF4 and heparin is critically dependent on their molar ratio, with loss of antibody binding when the optimal ratio is disrupted by an excess of either component. 6,7 Antigen formation on the platelet surface also follows a bell-shaped curve as PF4 concentration is increased, with maximal binding of antibody seen at an exogenous PF4 concentration of 50 g/mL. 8 Chondroitin sulfates (CSs) are the predominant GAG side chains expressed on platelets. 9,10 We have shown that the binding of the HIT-like monoclonal antibody KKO 11 to platelets is abrogated by chondroitinase ABC, 8 indicating that HIT antibodies bind to PF4/CS complexes on this cell type. Therapeutic concentrations of UFH disrupt antibody binding in part by eluting PF4 from the platelet surface, which reduces formation of antigenic complexes and the potential for platelet activation 8 through platelet Fc␥RIIA. 12 Thus, variation in the expression of platelet-derived PF4 (or CS) might help to explain why only a small percentage of patients who generate antibodies to PF4/UFH develop HIT. 13 Although it has been generally accepte...

Małopłytkowość indukowana protaminą — nowy problem czy inny typ małopłytkowości poheparynowej?

“…Patogeneza zjawiska HIT nie jest w pełni wyjaśniona. HIT abs oprócz aktywacji trombocytów wpły-wają także na monocyty i komórki śródbłonka, indukując ekspresję czynnika tkankowego oraz trombiny [14,16,17]. Podwyższone stężenie trombiny, a nie sama trombocytopenia, jest główną przyczyną ryzyka zakrzepicy [14].…”

“…Krótszy okres trwania PIT dodatkowo determinują inne właściwości przeciwciał. PIT abs nie reagują krzyżowo z innymi kompleksami, jak ma to miejsce w przypadku HIT abs a PF4 związanym z glikozaminoglikanami na powierzchni płytek krwi [16]. Dodatkowo heparynowy efekt z odbicia jest dowodem na ograniczony czas przebywania kompleksów protaminy i heparyny w ustroju [23].…”

Antigen and substrate withdrawal in the management of autoimmune thrombotic disorders

“…17 The in vivo relevance of these findings is supported by the relationship between the concentration of exogenous PF4 and binding of anti-PF4/H antibodies to platelets and monocytes (wherein antibody binding increases with addition of PF4 until an optimal concentration is reached above which binding falls) and the expression level of endogenous PF4 and the development of thrombocytopenia and thrombosis in an animal model. 12,14 The finding that activation of platelets and monocytes increases PF4 binding capacity may provide insight into why the risk of thrombosis is greater in settings, such as bypass surgery and trauma, among others, that are characterized by intense PF4 release, inflammation, and vascular injury.…”

“…PF4 also binds extensively to endothelium and other hematopoietic cells that express GAGs with higher affinity for PF4, such as heparan sulfate. 14 How does this help to explain the discrepancy between the incidence of seropositivity and clinical disease? Insight into this issue begins with the finding that PF4 and heparin form complexes that vary in size depending on the molar ratio of reactants.…”

Experts’ opinion or the serotonin release assay as a gold standard for the diagnosis of heparin‐induced thrombocytopenia (HIT)?

“…Third-, anti-PF4/heparin IgM and IgA antibodies are not detected by the SRA test. Although their pathogenic role is still a matter of debate, to rely on HIT diagnosis on SRA alone eliminates a priori the possibility that IgM and IgA antibodies might play a role as well as the potential implication of other cells (such as endothelial cells, monocytes) [12,13], which could lead to an underestimation of the disease. Conversely, the expertsÕ efficiency in diagnosing HIT can be disputed.…”