In vitro evaluation of blood coagulation activation and microthrombus formation by a microchannel array flow analyzer.
An in vitro assay system for evaluating microthrombus formation in whole blood has not yet been established. In this study, we characterized the relationship between coagulation activation and capillary occlusion in an in vitro assay system using a microchannel array flow analyzer (MC-FAN), which can evaluate microthrombus formation in whole blood as a function of blood flow rate. The flow rate of lipopolysaccharide (LPS)-treated whole blood decreased in a dose- and time-dependent manner. Addition of LPS-pretreated mononuclear cells (PBMCs) to whole blood also decreased the flow rate. Treatment with an anti-tissue factor (TF) antibody blocked the LPS-induced decrease in blood flow rate, suggesting that TF expressed on PBMCs is responsible for decreased blood flow in the microchannels. Known anticoagulants and anti-platelet agents also inhibited the LPS-induced decrease in blood flow rate. Thrombin-antithrombin complexes were increased in LPS-stimulated whole blood, and decreased by anticoagulants but not by anti-platelet agents. These findings suggest that occlusion in microchannels occurs mainly by formation of microthrombi composed of fibrin nets and platelet aggregates, which result from TF expression on activated PBMCs. In addition, the MC-FAN was found to be useful for in vitro evaluation of antithrombotic agents in the capillary.
Kamada
Haruhiko
H
Department of Molecular Pathobiology, Mie University School of Medicine, Edobashi 2-174, Tsu, 514-8507, Japan.
Hattori
Kunihiro
K
Hayashi
Tatsuya
T
Suzuki
Koji
K
D000991:Antithrombins / Q000494:pharmacology; D001780:Blood Coagulation Tests / Q000295:instrumentation* / Q000379:methods; D001783:Blood Flow Velocity / Q000187:drug effects*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004867:Equipment Design; D019544:Equipment Failure Analysis; D005434:Flow Cytometry / Q000295:instrumentation* / Q000379:methods; D017022:Flow Injection Analysis / Q000295:instrumentation / Q000379:methods; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods; D015539:Platelet Activation / Q000187:drug effects*; D013927:Thrombosis / Q000097:blood* / Q000139:chemically induced / Q000503:physiopathology*
D000991:Antithrombins; D008070:Lipopolysaccharides; C433788:lipopolysaccharide, E. coli O26-B6