benzyloxycarbonylvalyl-alanyl-aspartyl-fluoromethyl-ketone and Thrombocytopenia

TNF is known to induce a thrombocytopenia, due to a reduced platelet life span. Injection of TNF (10 microg) to mice did markedly increase the number of platelet-derived microparticles in plasma, most pronounced 1h after injection. Injection of TNF induced a transient activation of platelet caspases, -1, -3, -6, -8, -9, as seen by the binding of caspases probes detected by flow cytometry, most pronounced 1h after injection. Activation of caspase-3 was also evidenced by antibodies. Injection of the caspases inhibitor ZVAD-fmk decreased TNF-induced generation of microparticles and thrombocytopenia, indicating a causal role of caspases in platelet fragmentation. Activation of platelet caspases was also evident in platelets exposed to TNF in vitro, indicating that TNF acts on platelets directly. Comparison of platelets from +/+, TNFR1 -/- and TNFR2 -/- mice showed that caspases are activated mainly by the TNFR1. These observations indicate that TNF activates platelet caspases via the TNFR1, which results in platelet fragmentation and thrombocytopenia.

Topics: Amino Acid Chloromethyl Ketones; Animals; B-Cell Lymphoma 3 Protein; Blood Platelets; Blotting, Western; Caspase 3; Caspases; Cell Separation; Enzyme Activation; Flow Cytometry; Fluorescent Dyes; Kinetics; Mice; Mice, Inbred C57BL; Microscopy, Fluorescence; Peptides; Proto-Oncogene Proteins; Recombinant Proteins; Thrombocytopenia; Time Factors; Transcription Factors; Tumor Necrosis Factor-alpha

Infection of mice with Plasmodium Berghei Anka (PbA) leads to a thrombocytopenia, due to a reduced platelet life span, eventually associated with a syndrome of severe or cerebral malaria (CM). Thrombocytopenia was associated with an increase in the number of microparticles (mcp) in plasma. More than >60% of these mcp were of platelet origin, as seen by staining with an anti-platelet antibody. The thrombocytopenia and the amount of mcp were decreased in mice treated with anti CD40L mAb, suggesting that CD40L is the main effector of the thrombocytopenia. Caspase-1, -3, -6, -8, -9 were activated in platelets from infected mice, as seen by the binding of labeled probes or the amount of pro-caspase-3. Treatment of infected mice with the caspases inhibitor ZVAD-fmk decreased the number of mcp and the thrombocytopenia, showing that platelet caspases are responsible for platelet fragmentation. In addition, the caspase inhibitor also caused a decrease in the mortality associated with CM, indicating a critical role of caspases in the expression of CM.

Topics: Amino Acid Chloromethyl Ketones; Animals; Blood Platelets; Blotting, Western; Caspase 1; Caspase 3; Caspase 6; Caspase 8; Caspase 9; Caspases; CD40 Ligand; Cricetinae; Cysteine Proteinase Inhibitors; Disease Models, Animal; Enzyme Activation; Flow Cytometry; Malaria; Megakaryocytes; Mice; Mice, Inbred C57BL; Microscopy, Fluorescence; Peptides; Plasmodium berghei; Protein Binding; Thrombocytopenia; Time Factors; Tumor Necrosis Factor-alpha