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Blood, Vol. 94 No. 5 (September 1), 1999:
pp. 1657-1664
Discordant Expression of Tissue Factor and Its Activity in Polarized
Epithelial Cells. Asymmetry in Anionic Phospholipid Availability as a
Possible Explanation
Carsten B. Hansen,
Bo van Deurs,
Lars C. Petersen, and
L. Vijaya
Mohan Rao
From the Department of TF/VIIa Research, Health Care Discovery, Novo
Nordisk A/S, Denmark; Structural Cell Biology Unit, the Department of
Medical Anatomy, The Panum Institute, University of Copenhagen,
Denmark; and the Department of Biochemistry, the University of Texas
Health Center at Tyler, Tyler, TX.
Recent studies have shown a discrepancy between the level of tissue
factor (TF) expression and the level of TF procoagulant activity on the
apical and basolateral surface domains of polarized epithelial cells.
The present investigation was performed to elucidate possible reasons
for the discordant expression of TF and its activity on the surface of
polarized epithelial cells using a human intestinal epithelial cell
line, Caco-2 and Madin-Darby canine kidney epithelial cells, type II
(MDCK-II). Functional activity of coagulation factor VIIa (VIIa) in
complex with TF was 6- to 7-fold higher on the apical than the
basolateral surface in polarized Caco-2 cells. In contrast, no
significant difference was found in the formation of TF/VIIa complexes
between the apical and basolateral surface. Confocal microscopy of
Caco-2 cells showed TF expression on both the apical and the
basolateral surface domains. Studies with MDCK-II cells showed that the
specific functional activity of TF expressed on the apical cell surface
was 5-fold higher than on the basolateral surface. To test whether
differential expression of TF pathway inhibitor (TFPI) on the apical
and basolateral surface could account for differences in TF/VIIa
functional activity, we measured cell-surface-bound TFPI activity in
Caco-2 cells. Small but similar amounts of TFPI were found on both
surfaces. Further, addition of inhibitory anti-TFPI antibodies induced
a similar enhancement of TF/VIIa activity on both surface domains.
Because the availability of anionic phospholipids on the outer leaflet
of the cell membrane could regulate TF/VIIa functional activity, we
measured the distribution of anionic phospholipids on the apical and
basolateral surface by annexin V binding and thrombin generation. The
results showed that the anionic phospholipid content on the basolateral
surface, compared with the apical surface, was 3- to 4-fold lower. Mild
acid treatment of polarized Caco-2 cells, which markedly increased the
anionic phospholipid content on the basolateral surface membrane,
increased the TF/VIIa activity on the basolateral surface without
affecting the number of TF/VIIa complexes formed on the surface.
Overall, our data suggest that an uneven expression of TF/VIIa activity
between the apical and basolateral surface of polarized epithelial
cells is caused by differences in anionic phospholipid content between
the two surface domains and not from a polar distribution of TFPI.
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