Highlights
- •Protein disulfide isomerase (PDI) contributes to TF regulation in monocytes.
- •Propynoic acid carbamoyl methyl-amide-31 (PACMA) is a potent PDI inhibitor.
- •PACMA regulates monocyte TF by transcriptional and posttranscriptional mechanisms.
- •PACMA can convert preformed procoagulant TF into its cryptic state.
- •PACMA amplifies LPS-induced monocyte TF production in a PAR2-dependent manner.
Abstract
Introduction
Protein disulfide isomerase (PDI) contributes to tissue factor (TF) regulation in
monocytes. While bacitracin and quercetin-3-rutinoside mitigate myeloid TF production,
the effect of PACMA-31, a more specific PDI inhibitor with distinct pharmacologic
properties, remains unclear.
Materials and methods
Lipopolysaccharide (LPS) stimulation of peripheral blood mononuclear cells (PBMCs)
or citrate-anticoagulated whole blood was carried out in the presence of PACMA-31
or DMSO vehicle before monocytes were analyzed for TF expression, including antigen,
procoagulant activity (PCA) and mRNA, release of IL-6 and TNFα, and LPS-induced signaling
pathways.
Results
While PACMA-31 alone had no effect, coincubation with LPS and PACMA-31 (25 μM) enhanced
LPS-induced monocyte TF production in whole blood. The effect was at least partially
regulated on the transcriptional level and could not be explained by increased phosphatidylserine
membrane exposure. In contrast, the same PACMA-31 concentrations were cytotoxic in
isolated PBMCs. A lower dose of PACMA-31, however, restored the stimulating effect
by enhancing IκB-NFκB signaling that also increased the release of IL-6 and TNFα.
The protease-activated receptor 2 (PAR2) inhibitor ENMD547 but not TF antibody 10H10 or factor Xa inhibitor rivaroxaban prevented the stimulatory
effect of PACMA-31 on inflammatory monocytes. In sharp contrast, short time incubation
of LPS-stimulated PBMCs with 25 μM PACMA-31 was non-cytotoxic and significantly inhibited
cellular TF PCA but not surface antigen expression.
Conclusions
PACMA-31 regulates monocyte TF in a concentration-dependent manner by opposing transcriptional
and posttranscriptional mechanisms. While low concentrations of PACMA-31 augment monocyte
TF production by amplifying LPS-dependent PAR2 signaling, high concentrations convert
monocyte TF into its non-coagulant state.
Graphical abstract
Graphical Abstract
Keywords
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Article info
Publication history
Published online: September 30, 2022
Accepted:
September 28,
2022
Received in revised form:
September 9,
2022
Received:
July 25,
2022
Identification
Copyright
© 2022 Elsevier Ltd. All rights reserved.
