thromboplastin and defibrotide

The coagulation-inflammation cycle has been implicated as a critical component in malaria pathogenesis. Defibrotide (DF), a mixture of DNA aptamers, displays anticoagulant, anti-inflammatory, and endothelial cell (EC)-protective activities and has been successfully used to treat comatose children with veno-occlusive disease. DF was investigated here as a drug to treat cerebral malaria.. DF blocks tissue factor expression by ECs incubated with parasitized red blood cells and attenuates prothrombinase activity, platelet aggregation, and complement activation. In contrast, it does not affect nitric oxide bioavailability. We also demonstrated that Plasmodium falciparum glycosylphosphatidylinositol (Pf-GPI) induces tissue factor expression in ECs and cytokine production by dendritic cells. Notably, dendritic cells, known to modulate coagulation and inflammation systemically, were identified as a novel target for DF. Accordingly, DF inhibits Toll-like receptor ligand-dependent dendritic cells activation by a mechanism that is blocked by adenosine receptor antagonist (8-p-sulfophenyltheophylline) but not reproduced by synthetic poly-A, -C, -T, and -G. These results imply that aptameric sequences and adenosine receptor mediate dendritic cells responses to the drug. DF also prevents rosetting formation, red blood cells invasion by P. falciparum and abolishes oocysts development in Anopheles gambiae. In a murine model of cerebral malaria, DF affected parasitemia, decreased IFN-γ levels, and ameliorated clinical score (day 5) with a trend for increased survival.. Therapeutic use of DF in malaria is proposed.

Topics: Animals; Anti-Inflammatory Agents; Anticoagulants; Antimalarials; Blood Coagulation; Cells, Cultured; Complement Activation; Cytokines; Dendritic Cells; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelial Cells; Female; Glycosylphosphatidylinositols; Hemoglobins; Humans; Inflammation Mediators; Malaria, Cerebral; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Nitric Oxide; Plasmodium berghei; Plasmodium falciparum; Platelet Aggregation; Polydeoxyribonucleotides; Receptors, Purinergic P1; Severity of Illness Index; Thromboplastin; Time Factors

Defibrotide (DF), a polydeoxyribonucleotide with antithrombotic properties, has recently proven effective in patients with severe hepatic veno-occlusive disease (VOD), a life-threatening complication of high-dose chemo/radiotherapy regimens for stem cell transplantation. To understand the mechanism of its beneficial effect, we studied the impact of DF on the expression of tissue factor (TF) and fibrinolytic proteins (PAI-1 and t-PA) on endothelial cells. The in vitro response to DF of two types of human endothelial cells (ECs) of different origins, that is from macrovascular (HUVEC) and microvascular (HMEC-1 cell line) beds, was evaluated in the presence or absence of a proinflammatory stimulus (ie bacterial endotoxin, LPS). The results show that DF was able to significantly reduce the LPS-induced TF expression by HMEC-1, and less prominently by HUVEC. In addition, DF importantly influenced the fibrinolytic properties of both HMEC-1 and HUVEC. Specifically, it dose-dependently counteracted the LPS-induced increase in PAI-1 levels and decrease in t-PA activity expression. It also significantly incremented t-PA antigen in resting EC. Decreasing the procoagulant activity and increasing the fibrinolytic potential of EC favors an anticoagulant phenotype of the endothelium, which may protect from fibrin deposition and vascular occlusion.

Topics: Blood Coagulation Factors; Cell Line; Dose-Response Relationship, Drug; Endothelium, Vascular; Fibrinolytic Agents; Humans; Lipopolysaccharides; Plasminogen Activator Inhibitor 1; Polydeoxyribonucleotides; Thromboplastin; Tissue Plasminogen Activator; Umbilical Veins

We evaluated the release of tissue factor pathway inhibitor (TFPI) induced by defibrotide (DF), a single-stranded, negatively charged polydeoxyribonucleotide extracted from mammalian organ. Ten normal volunteers were injected with an intravenous bolus of 400 mg DF and 2,000 IU unfractionated heparin (UFH). In addition, three volunteers were also injected with an intravenous bolus of 2,000 anti-Xa U of two low-molecular-weight heparins (LMWHs), enoxaparin and nadroparin. UFH caused a 4-fold increase in plasma TFPI at 5 minutes, with a decrease that was parallel to the heparin level measured by the anti-Xa assay. However, at 80 minutes, although the plasma anti-Xa activity of UFH was almost undetectable, the level of TFPI remained 2-fold baseline. DF induced an increase of TFPI that was 2-fold higher than the baseline level, with a steady state achieved between 5 and 20 minutes. At 40 minutes, the TFPI levels returned to basal level. This pattern was not coincident with the clearance of DF and at 40 minutes, the concentration of DF was still one third of the levels at 5 minutes (25.4 +/- 4.04 microg/mL). Both of the LMWHs induced a similar TFPI peak level at 5 minutes (1.5-fold increase) and at 40 minutes the TFPI levels returned to the initial levels. At 5 minutes, both LMWHs showed a higher plasma anti-Xa activity than UFH, which was detectable even at 80 minutes. The current study demonstrated that one of the mechanisms of the antithrombotic activity of DF is mediated via TFPI. Furthermore, the release of TFPI by heparin is mediated by non-antithrombin III binding fragments. Thus, polyanionic electrolytes are capable of releasing TFPI irrespective of their antithrombin III effect.

Topics: Adult; Anticoagulants; Endothelium, Vascular; Enoxaparin; Factor Xa Inhibitors; Female; Fibrinolytic Agents; Heparin; Humans; Lipoproteins; Male; Middle Aged; Nadroparin; Polydeoxyribonucleotides; Thrombomodulin; Thromboplastin