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Activation of factor X and its regulation by tissue factor pathway
inhibitor in small-diameter capillaries lined with human endothelial cells
T Lindhout, R Blezer, P Schoen, O Nordfang, C Reutelingsperger and HC Hemker
Department of Biochemistry, Cardiovascular Research Institute Maastricht,
University of Limburg, The Netherlands.
The activation of factor X at the surface of endothelial cells was
investigated under controlled flow conditions. A method is described for
preparing polyethylene capillaries whose inner walls are covered with a
confluent layer of human umbilical vein endothelial cells. To obtain a
stable and unperturbed layer of endothelial cells it was essential to
pre-perfuse the endothelialized capillaries with medium for about 18 hours.
At this stage no tissue factor activity could be detected, but when the
seeded cells were perfused with medium containing tumor necrosis factor
(TNF) a maximum steady-state rate of factor Xa production (16 fmol factor
Xa/min/cm2) was observed within 8 hours. Further experiments were performed
with endothelial cells incubated for 4 hours with TNF. Factor Xa was
produced at a rate of 7 fmol factor Xa/min/cm2 on perfusion of the
capillaries with factor X (100 nmol/L) and factor VII (0.1 U/mL) at a shear
rate of 34 s-1. The extracellular matrix preparations of these cells
produced factor Xa at a 20-fold higher rate (150 fmol factor Xa/min/cm2).
In both cases factor Xa formation was dependent on the presence of factor
VII and was completely inhibited when the perfusate also contained 5 nmol/L
recombinant tissue factor pathway inhibitor (rTFPI). Pre-perfusion with
factor Xa-TFPI complex in the absence of factor VIIa caused a much lesser
inhibitory effect, suggesting that TFPI-mediated neutralization of
endothelial cell and matrix tissue factor activity requires the presence of
factor VIIa in addition to the presence of factor Xa.
Volume 79,
Issue 11,
pp. 2909-2916,
06/01/1992
Copyright © 1992 by The American Society of Hematology
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