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The functional expression of tissue factor by fibroblasts and endothelial
cells under flow conditions
EF Grabowski, DB Zuckerman and Y Nemerson
Department of Pediatrics, Massachusetts General Hospital, Boston 02114.
The expression of tissue factor (TF) by a variety of vascular cell types
under physiologic flow conditions is critical to factor X activation and in
vivo clotting. Therefore, in a parallel-plate flow chamber (volume 40
microL) we mounted monolayers of human embryonic fibroblasts (FBs) or
interleukin-1 alpha (IL-1 alpha) (5 U/mL x 4 hours)-stimulated human
umbilical vein endothelial cells (ECs). Inflow buffer contained 10 nmol/L
factor VIIa, 100 nmol/L factor X, and 2.0 mmol/L CaCl. With FBs, production
of factor Xa (product of outflow concentration of factor Xa-and flow rate)
increased 200-fold over the range of shear stress from 0 to 2.7 dynes/cm2.
Production values (mean +/- SE (N)) were 7.93 +/- 0.024 (6), 312 +/- 7.3
(6), 688 +/- 33.1 (8), 1,033 +/- 119 (6), and 1,601 +/- 183 (7)
fmol/cm2.minute at shear stresses of 0, 0.27, 0.68, 1.35, and 2.7
dynes/cm2, respectively. Further experiments at 0.68 dynes/cm2 indicated
that factor Xa production increased with factor X concentration over the
range from 3 to 100 nmol/L, but changed little from 300 to 1,000 nmol/L.
With ECs, production was 0.13 +/- 0.86 (6), 8.17 +/- 1.65 (13), and 1.66
+/- 1.66 (5) fmol/cm2.minute at 0, 0.68, and 2.7 dynes/cm2, respectively.
However, in the presence of an antibody directed against tissue factor
pathway inhibitor (TFPI) production with ECs was augmented to 16.46 +/-
0.80 (8), 149.8 +/- 18.6 (8), and 48.9 +/- 10.3 (10), respectively, at
these same shear stresses. Control experiments with factor VIIa, factor X,
or both absent confirm for both cell types the specificity of the reaction
for the TF pathway. Similarly, specificity for TF itself is shown by the
virtual absence of factor Xa generation in the presence of the monoclonal
antibody HTF1-7B8 directed against human TF. We conclude that ECs, even
when activated, are normally unable to generate significant quantities of
factor Xa in the presence of factors X and VIIa. However, significant
quantities of factor Xa are possible in the presence of an inhibitor of
TFPI. On the other hand, production of factor Xa by fibroblasts is markedly
augmented by shear stress, yet independent of the availability of substrate
factor X above an inflow concentration of 100 nmol/L. The latter suggests a
direct effect of flow on the fibroblast monolayers, not substrate
limitation by convective diffusion.
Volume 81,
Issue 12,
pp. 3265-3270,
06/15/1993
Copyright © 1993 by The American Society of Hematology
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