| Popis: |
Because the lack of thromboresistant vascular biomaterials is in part due to platelet activation, we have attempted, by using fluorescence-activated flow cytometry, to fully characterize the platelet population after in vitro material contact with whole blood. We have used a very simple, near-physiologic system whereby whole blood, anticoagulated with D-phenylalanyl-L-prolyl-arginyl chloromethyl ketone (thrombin inhibitor), contacts materials for 1 hour at 37 degrees C, under low shear. Unlike other tests of platelet compatibility that focus on adherent platelets, this assay evaluates the platelets in the whole blood drained from the tube (1.57 mm internal diameter, 25 cm length) after material contact. We demonstrate for the first time significant materials-induced microparticle formation. One-hour contact with Silastic, polyethylene, and polyvinyl alcohol hydrogel surfaces lead to 30 +/- 1, 33 +/- 4, and 43 +/- 4 x 10(9) microparticles/L, respectively, whereas resting blood samples contained only 10 +/- 1 x 10(9) microparticles/L. In addition, significant increases in activated platelet(s) binding to neutrophils/monocytes after material contact were noted for all surfaces tested. For polyvinyl alcohol hydrogel surfaces a greater than 500% increase in the fluorescent intensity over that of resting whole blood was attained. The addition of monoclonal antibodies to GPIIb/IIIa (A2A9), the tetrapeptide adhesion ligand RGDS (arginine-glycine-aspartate-serine), or the calcium ion chelator ethyleneglycol-bis-(B-aminoethyl-ether)- N,N,N',N'-tetraacetic acid to the whole blood before material contact fully inhibited all platelet reactivity noted for all surfaces--platelet microparticles, platelet P-selectin expression, loss of platelets from bulk, and the formation of platelet/leukocyte aggregates--thereby indicating that material-induced platelet activation is a calcium-dependent process involving GPIIb/IIIa receptors. |
