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Expression of tissue factor pathway inhibitor by cultured endothelial cells
in response to inflammatory mediators
A Ameri, MN Kuppuswamy, S Basu and SP Bajaj
Department of Medicine, St Louis University School of Medicine, MO.
We recently proposed that endothelium may represent the primary physiologic
site of synthesis of the tissue factor pathway inhibitor (TFPI). In support
of this conclusion, we have now found that the poly(A)+ RNAs obtained from
rabbit and bovine lung tissues contain abundant amounts of TFPI messenger
RNAs (mRNAs), whereas the poly(A)+ RNAs obtained from the liver of these
animals contain less than 5% of that found in the lung tissues. Because
inflammatory mediators are known to upregulate tissue factor (TF)
expression by the endothelium, we have examined the effect of these agents
on the TFPI expression by the cultured endothelial cells. When cultured
human umbilical vein endothelial cells were stimulated (in 10% fetal bovine
serum) with phorbol myristate acetate (PMA), endotoxin, interleukin-1, or
tumor necrosis factor-alpha, the TF mRNA increased approximately 7- to 10-
fold within 2 to 4 hours. Unstimulated cells constitutively expressed TFPI
mRNA and its levels either did not change or increased slightly (up to
1.5-fold) upon stimulation with these inflammatory agents. TF mRNA abruptly
declined to a negligible level and the TFPI mRNA returned essentially to
the basal level at approximately 24 hours. The membrane- bound TF clotting
activity of induced cells peaked between 4 and 8 hours, and finally
declined. The cumulative TFPI activity secreted into the media was either
unchanged or slightly higher in the induced cell cultures as compared with
that present in the noninduced cultures. Endothelial cells were also
cultured in 10% heat-inactivated human serum derived from plasma or whole
blood. TFPI secreted into the media containing whole blood serum was
consistently higher (approximately 1.5- fold at 8 hours) than that secreted
into the media supplemented with serum obtained from plasma lacking the
formed elements; these cells also expressed similarly increased levels of
TFPI mRNA. Moreover, PMA- stimulated cells cultured in whole blood serum
expressed modestly increased levels of TFPI mRNA (approximately 1.5-fold);
supernatants from these cells also contained similarly increased TFPI
activity. Cumulatively, our data indicate that, unlike thrombomodulin and
fibrinolytic enzymes synthesized by the endothelial cells, TFPI synthesis
is not downregulated and may be slightly upregulated during an inflammatory
response. Inspection of the 5' flanking region of the TFPI gene showed a
conserved GATA-binding motif located approximately 400 bp upstream of the
proposed transcription initiation site(s). This motif by binding to the
GATA-2 transcriptional factor may keep the endothelium in an 'on' state for
constitutive expression of TFPI.(ABSTRACT TRUNCATED AT 400 WORDS)
Volume 79,
Issue 12,
pp. 3219-3226,
06/15/1992
Copyright © 1992 by The American Society of Hematology
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