Platelet-induced expression of tissue factor procoagulant activity in freshly isolated human mononuclear cells: implications for experimental use.
Monocytes express tissue factor (TF) as a result of cytokine stimulation or endothelial adherence. We evaluated monocyte-platelet interaction in vitro as another trigger for monocyte TF enhancement in human mononuclear cells isolated by density gradient centrifugation from peripheral blood. Cell TF procoagulant activity (TF-PCA) was quantitated by a one-stage recalcification clotting time assay. Platelets were counted and identified by whole blood flow cytometry as CD61 positive particles, activated platelets were characterized by P-Selectin (CD62) expression, and monocytes by surface CD14 expression. A significant correlation between normalized TF-PCA of isolated mononuclear cells and platelet count was shown (r = 0.43, P < 0.001). Percentage of activated platelets in baseline samples was 4.2 +/- 3.5 while adenosine diphosphate (ADP) increased platelet positivity to 34 +/- 17% (P < 0.001). After isolation, 52 +/- 12% of platelets within suspensions were activated (P < 0.0001). Percentage of CD62-positive monocytes (CD14+ particles) increased from baseline 5% to 13 +/- 6% in ADP-stimulated samples to 53 +/- 17% after isolation (P < 0.001). These findings suggest that density gradient centrifugation activates platelets and that an adhesive interaction between monocytes and platelets may promote TF-PCA expression in isolated mononuclear suspensions. Enhanced monocyte TF expression as a result of an activated platelet-monocyte interaction seems to be an important laboratory effect requiring consideration when utilizing this technique in an experimental setup.
Meisel
S R
SR
Department of Medicine, Division of Cardiology, Atherosclerosis Research Center, Cedar-Sinai Burns & Allen Research Institute, Israel. meisel@netvision.net.il
Shechter
M
M
Xu
X-P
XP
Edgington
T S
TS
Cercek
B
B
Shah
P K
PK
D001792:Blood Platelets / Q000502:physiology*; D002450:Cell Communication; D002499:Centrifugation, Density Gradient; D006801:Humans; D018950:Lipopolysaccharide Receptors / Q000032:analysis; D009000:Monocytes / Q000378:metabolism*; D019007:P-Selectin / Q000032:analysis; D010974:Platelet Aggregation; D013925:Thromboplastin / Q000378:metabolism*
D018950:Lipopolysaccharide Receptors; D019007:P-Selectin; D013925:Thromboplastin