Natural IgM Against Oxidation-Specific Epitopes Inhibit Microvesicle-Driven Coagulation

Increasing evidence demonstrates an important role for immune functions in coagulation and thrombosis, and both microbial and sterile triggers have been implicated in these responses. Oxidation-specific epitopes (OSE) are products of lipid peroxidation that are present on oxidized low-density lipoproteins and dying cells, and represent important sterile triggers of inflammation in cardiovascular diseases. We have previously shown that a large part of naturally occurring IgM antibodies have specificity for OSEs and that 50% of circulating procoagulant microvesicles (MV) carry OSEs, such as malondialdehyde-adducts (MDA), on their surface. Because OSE-specific natural IgM bind MVs and increased levels of circulating MVs have been implicated in an increased thrombotic risk, we studied the capacity of these antibodies in directly modulating the procoagulant potential of MVs in vitro and in vivo. We first showed that increasing concentrations of three different monoclonal IgM antibodies with specificity for MDA (LR04, NA17, E014) significantly delayed MV-induced fibrin formation in plasma, while an MDA-specific IgG antibody or F(ab')₂ fragments of LR04 had no effect. Moreover, peak thrombin generation of normal, factor VII- or factor XII- deficient microvesicle poor plasma (MVPP) substituted with tissue factor positive MVs was significantly reduced in the presence of LR04 but not a control IgM antibody, indicating that it exerts its effect on the common pathway of the coagulation cascade. A similar anticoagulatory effect of LR04 was observed in freshly drawn whole blood analyzed by rotational thrombelastometry. Addition of LR04, compared to a control IgM antibody, increased clotting time, clot formation time, and the time to reach maximum clot firmness, and decreased maximum clot firmness. Finally, in a murine pulmonary embolism model, we could show that 30-minute survival of mice that received intravenous injections of epinephrine together with pro-coagulant MVs derived from the pancreatic cancer cell line HPAFII was significantly increased in mice co-injected with LR04 compared to controls. Our study identifies an inhibitory role of MDA-specific IgM antibodies in MV-induced coagulation. These effects are exerted on the common pathway of the coagulation cascade and require the intact structure of IgM antibodies. The findings may offer novel diagnostic and therapeutic approaches in the prevention and treatment of thrombosis.