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Tissue Factor: An Essential Mediator of Hemostasis and Trigger of

Thrombosis

Article  in  Arteriosclerosis Thrombosis and Vascular Biology · February 2018

DOI: 10.1161/ATVBAHA.117.309846

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 Brief Review

Tissue Factor
An Essential Mediator of Hemostasis and Trigger of Thrombosis
Steven P. Grover, Nigel Mackman

Abstract—Tissue factor (TF) is the high-affinity receptor and cofactor for factor (F)VII/VIIa. The TF-FVIIa complex is the
primary initiator of blood coagulation and plays an essential role in hemostasis. TF is expressed on perivascular cells and
epithelial cells at organ and body surfaces where it forms a hemostatic barrier. TF also provides additional hemostatic protection
to vital organs, such as the brain, lung, and heart. Under pathological conditions, TF can trigger both arterial and venous
thrombosis. For instance, atherosclerotic plaques contain high levels of TF on macrophage foam cells and microvesicles
that drives thrombus formation after plaque rupture. In sepsis, inducible TF expression on monocytes leads to disseminated
intravascular coagulation. In cancer patients, tumors release TF-positive microvesicles into the circulation that may contribute
to venous thrombosis. TF also has nonhemostatic roles. For instance, TF-dependent activation of the coagulation cascade
generates coagulation proteases, such as FVIIa, FXa, and thrombin, which induce signaling in a variety of cells by cleavage of
protease-activated receptors. This review will focus on the roles of TF in protective hemostasis and pathological thrombosis.
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Visual Overview—An online visual overview is available for this article.   (Arterioscler Thromb Vasc Biol. 2018;38:
709-725. DOI: 10.1161/ATVBAHA.117.309846.)
Key Words: blood coagulation ◼ factor VII ◼ hemostasis ◼ thromboplastin ◼ thrombosis

T issue factor (TF) is a transmembrane protein that func-

tions as a high-affinity receptor for factor (F)VII and
FVIIa.1–6 The TF-FVIIa complex is the primary activator of
the coagulation protease cascade. TF is required to stabi-
lize the catalytic site of FVIIa on a plasma membrane (PM)
to allow optimal interaction with its substrates FIX and FX.
There are many excellent reviews on TF.1–6 The goal of this
review is to summarize our current knowledge on the roles of
TF in hemostasis and thrombosis.
Brief History of the Major Advances
in TF Research
The original extrinsic coagulation system consisted of 4 fac-
tors: fibrinogen (FI), prothrombin (FII), thromboplastin (FIII,
TF), and Ca++ (FIV). This basic blood coagulation system is
present in jawless fish that diverged over 450 million years
ago.7 Morawitz was the first to show that conversion of pro-
thrombin to thrombin in plasma was induced by tissue throm-
boplastin.8 Indeed, TF was originally simply a laboratory
reagent derived from tissues, such as the placenta, that was
added to plasma to measure the prothrombin time. Early stud-
ies showed that TF was a protein that required phospholipid
for its activity.9,10 It was subsequently purified by 2 groups
in the 1970s.11,12 In 1987, the TF cDNA was cloned by 4
groups,13–16 which led to an explosion of research on TF. The
TF gene was cloned in 1989, which allowed analysis of its
transcriptional regulation.17 The crystal structure of the extra-
cellular region was found to consist of 2 immunoglobulin-like
domains,18 and was the first structure of a member of the class
2 cytokine receptor family. In 1996, 3 groups reported that
deletion of the TF gene in mice leads to universal death during
embryonic development or shortly after birth, indicating that
TF is essential for survival.19–21
Coagulation Protease Cascade
Hoffman and Monroe proposed a new cell-based model of
coagulation with a TF-positive cell and activated platelets
that involves initiation, amplification, and propagation.22 We
have adapted this model and have divided the phases of blood
coagulation into TF-FVIIa-dependent initiation, inhibition of
the TF-FVIIa complex by TF pathway inhibitor (TFPI) and
amplification of thrombin generation.
After vessel injury or tissue trauma the TF-FVIIa complex
is the spark that triggers blood coagulation by activating both
FX and FIX. FVII bound to TF is rapidly activated by a vari-
ety of coagulation proteases, although autoactivation by trace
amounts of FVIIa may be the primary pathway.23 Several stud-
ies have shown that the TF-FVIIa complex can activate FIX in
vitro.24–26 One study analyzed the activation of FIX and FX in
an in vitro system and found that FIXa was the major substrate

Received on: December 13, 2017; final version accepted on: January 25, 2018.
From the Thrombosis and Hemostasis Program, Division of Hematology and Oncology, Department of Medicine, University of North Carolina at
Chapel Hill.
Correspondence to Nigel Mackman, PhD, Division of Hematology and Oncology, Department of Medicine, 111 Mason Farm Rd, 2312C Medical
Biomolecular Research Bldg, CB No. 7126 University of North Carolina at Chapel Hill, Chapel Hill, NC 27599. E-mail nmackman@med.unc.edu
© 2018 American Heart Association, Inc.
Arterioscler Thromb Vasc Biol is available at http://atvb.ahajournals.org DOI: 10.1161/ATVBAHA.117.309846

709
 710   Arterioscler Thromb Vasc Biol   April 2018
Nonstandard Abbreviations and Acronyms
aPLA antiphospholipid antibody
APS antiphospholipid syndrome
EC endothelial cell
F factor
FVIIai active site–inhibited factor VIIa
hTF human tissue factor
IVC infrarenal vena cava
LPS lipopolysaccharide
NAPc2 nematode anticoagulant protein 2
PAR protease-activated receptor
PCA procoagulant activity
PM plasma membrane
SCD sickle cell disease
TF tissue factor
TFPI tissue factor pathway inhibitor
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of the TF-FVIIa complex.27 Moreover, TF-FVIIa activation of
FIX is important in vivo because a point mutation at residue
182, which led to reduced interaction with the TF-FVIIa com-
plex without affecting activation by FXIa, was associated with
a mild form of hemophilia B.28 In addition to the TF-FVIIa
complex, a recent study showed that the TF-FVIIa-FXa com-
plex can activate FVIII, which would provide additional lev-
els of FVIIIa during the initiation phase.29 Formation of the
prothrombinase complex (FVa-FXa) generates small amounts
of thrombin in the initiation phase. Activated platelets are an
important source of FVa that may contribute to the initiation
phase of blood coagulation.30 The final part of the initiation
phase is thrombin activation of the cofactors FV and FVIII
and platelets via protease-activated receptors (PARs). The
TF-FVIIa complex is then rapidly inhibited by TFPI in a FXa-
dependent manner.
In the amplification phase the tenase complex (FVIIIa-
FIXa) is assembled on the surface of activated platelets and
generates additional FXa. The prothrombinase complex is
also assembled on the surface of activated platelets. Thrombin
activation of FXI also allows additional activation of FIX.31
The net result is the generation of large amounts of throm-
bin. Thrombin cleaves fibrinogen to fibrin monomers that are
cross-linked by the transglutaminase FXIIIa, which is acti-
vated by thrombin. In addition, thrombin activates additional
platelets.
Blood-Borne TF
Blood was not thought to contain functional TF because of its
high procoagulant activity (PCA). However, in 1999, Giesen
et al32 reported the presence of blood-borne TF that contributed
to thrombus formation when native blood was flowed over
collagen-coated slides. In contrast, Butenas et al33,34 could not
detect active TF in whole blood. One difference between these
studies is flow, which may be required to reveal the functional
activity of encrypted or intracellular TF. To date, the accumu-
lated evidence suggests that there are very low levels of TF in
blood but this TF is either encrypted or inhibited and must be
activated in some way to activate coagulation. Interestingly,
one study reported TF staining throughout thrombotic clots
but only at the edges of hemostatic clots in mice, and sug-
gested that blood-borne TF may contribute to thrombosis but
not hemostasis.35
TF Expression by Circulating Cells
and Vascular Endothelial Cells
It has been reported that ≈1.5% of CD14+ monocytes in
humans express low levels of intracellular TF under resting
conditions.36 Exposure of monocytes to bacterial lipopolysac-
charide (LPS) in vitro or in vivo leads to a rapid and transient
expression of TF37–39 (Table 1). This induction is mediated by
the transcription factors AP-1 and NF-κB.59–61 We think that
TF is expressed by monocytes as part of the innate immune
response presumably to facilitate clot formation to limit the
dissemination of pathogens.
Osterud and Bjorklid62 proposed that TF is exclusively
expressed by circulating monocytes in blood of healthy
individuals and that under pathological conditions mono-
cytes release TF-positive microvesicles that bind to other
cells, such as activated platelets, neutrophils, and endothe-
lial cells (ECs). Indeed, in vitro studies support the notion
of the transfer of monocyte-derived microvesicles to plate-
lets and neutrophils.63–66 However, it is difficult to make any
conclusions about the origins of TF on cells in vivo because
of the fact that microvesicles can transfer antigens, such as
TF, between cells. The most definitive way to determine
which cells are making TF is to measure TF mRNA by in
situ hybridization.
TF expression by platelets, neutrophils, and eosinophils is
controversial (Table 1). We found that unstimulated platelets
contain TF pre-mRNA that is spliced to TF mRNA after acti-
vation of the platelets, although this process is relatively slow
(≈30 minutes).42 Other studies have reported variable levels
of TF mRNA and protein in unstimulated platelets but an
increase of TF mRNA and protein in activated platelets.43–45,67
Other groups failed to detect TF expression in platelets.49–51
A recent study convincingly showed that a megakaryocyte
line expressed TF pre-mRNA, TF mRNA and TF protein and
released a subset of TF-positive platelets.46 Interestingly, the
megakaryocyte cell line contained higher levels of TF mRNA
versus TF pre-mRNA whereas platelets derived from the
cells had more TF pre-mRNA. Similar results were observed
with megakaryocytes derived from CD34+ cells. It should be
noted that the megakaryocyte cell line expresses very low
levels of TF (90±15 pg/mL), which is <0.1% of the level of
TF observed in a human pancreatic cell line.68 Several studies
have found TF-positive platelets in patients with acute coro-
nary syndromes and diabetes mellitus.47,48 Because of the time
needed to splice TF pre-mRNA and express TF is seems more
likely that platelet TF contributes to thrombosis rather than
hemostasis.69
TF has been reported to be expressed by neutro-
phils, eosinophils, and T-cells in vitro and in vivo
(Table 1).52,53,55,56,58 However, the Osterud group failed
to detect TF expression in human neutrophils or eosino-
phils.54,57 We could not detect any functional TF in acti-
vated human neutrophils (Alexander W and Mackman N,
 Grover and Mackman   Tissue Factor in Hemostasis and Thrombosis   711
Table 1.  Cell Type-Specific Tissue Factor Expression In Vitro
and In Vivo
Cell Type In Vitro In Vivo
Monocyte Yes 37 39,40
Endothelium Yes 41 40
Platelets Yes 42–46 47,48
No 49–51 …
Neutrophils Yes 52 53
No 54 … phosphatidylserine but not to the complex on unperturbed
Eosinophils Yes 55,56 …
No 57 … substrate.88 Figure 1A shows a summary of the proposed phos-
T-cells Yes 58 58
unpublished data, 2013). However, there is stronger evi-
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dence for TF expression by neutrophils in monkeys, rabbits,
and mice compared with humans70–73 (Erlich J and Mackman
N, unpublished data, 2000). In addition, we found that dele-
tion of TF in neutrophils reduces LPS-induced activation
of coagulation in mice (Grover and Mackman, unpublished
data, 2018).
TF expression by ECs is readily demonstrably in vitro but
it is more difficult to demonstrate TF expression by ECs in
vivo in part because of the very low levels of expression.74–76
Typically, studies are performed with cultured ECs, such as
human umbilical vein ECs. It is often assumed that these cells
can be used as models of the variety of ECs present in differ-
ent vascular beds in vivo, which is unlikely to be true. We and
others have shown that human umbilical vein ECs express TF
in response to a variety of different agonists and this induction
is mediated by activation of NF-κB.41,77,78 The most convincing
examples of TF expression in vivo are by ECs in the splenic
microvascular, at arterial branching areas in septic baboons
and in the lungs of sickle cell disease (SCD) mice.79–82 Another
study reported TF expression in ECs in the glomeruli of endo-
toxemic mice.83 One difficulty with these immunohistochemi-
cal studies is TF staining of a given cell does not mean that the
cell expressed TF because this signal could be because of the
binding of TF-positive microvesicles from other cells, such as
activated monocytes.
Transcriptional and Post-Transcriptional
Regulation of TF
Because of the large amplification of the coagulation cas-
cade, TF-dependent activation of blood coagulation needs
to be tightly regulated to maintain hemostasis and prevent
thrombosis. The first level of control is at the level of TF gene
expression. For instance, high TF expression is restricted to
cells not in direct contact with blood.79,84 Additionally, TF
PCA is regulated at the post-translational level. TF exists in
2 states: encrypted TF present on unperturbed cells and de-
encrypted TF on damaged cells.85 FVIIa can bind to TF on
cells under resting conditions but the level of TF PCA activity
is very low. However, activation or disruption of cells dramati-
cally increases TF PCA.86,87 In terms of hemostasis, this rapid
increase in TF PCA would enhance the activation of coagula-
tion at sites of vessel injury and cell damage.
Several mechanisms have been proposed for the post-
translational control of TF PCA.6 The most popular mecha-
nism is that phosphatidylserine interacts with TF resulting in a
conformational change and enhanced PCA on cell surfaces.88
In support of this mechanism, TF PCA is enhanced by the
addition of phosphatidylserine to reconstituted phospholipid
vesicles.86,89 Additionally, the pseudosubstrate TFPI-FXa
bound to the TF-FVIIa complex on perturbed cells exposing
cells, which suggested that activation of the cells leads to a
change in conformation of TF that enables engagement with
phatidylserine-dependent model of TF de-encryption. Briefly,
the PM of eukaryotic cells has an asymmetrical distribution
of phospholipids with negatively-charged phospholipids, such
as phosphatidylserine and phosphatidylethanolamine, seques-
tered in the inner leaflet and neutrally charged phospholipids,
such as phosphatidylcholine and sphingomyelin, present in the
outer leaflet. Cellular damage leads to loss of the PM asym-
metry and exposure of negatively-charged phospholipid, such
as phosphatidylserine, on the cell surface. In a recent study,
molecular dynamic simulations were used to generate a model
of membrane-bound TF-FVIIa complex using soluble TF,
which lacks the transmembrane domain.90 It was found that
“the direct interaction of the phosphatidylserine headgroups
with specific residues on soluble TF most likely also affect the
conformation of these residues.” Further, they concluded that
“lipids might have an active direct role in de(-en)cryption of
soluble TF.”90 Phosphatidylserine exposure on the cell surface
also facilitates the binding of coagulation proteins (FII, FVII,
FIX, and FX) with positively-charged γ-carboxyglutamic acid
(Gla) domains and assembly of the cofactor/protease com-
plexes. In contrast to phosphatidylserine-rich lipid surfaces,
phosphatidylcholine-rich lipid surfaces confer relatively lim-
ited TF PCA suggesting that phosphatidylcholine maintains
TF in an encrypted state.91,92
More recently it was reported that sphingomyelin, a major
phospholipid in the outer leaflet of the PM, helps maintain
the TF-FVIIa complex in an encrypted state in resting cells.93
Cellular activation leads to translocation of acid sphingomy-
elinase to the PM and hydrolysis of sphingomyelin enabling
de-encryption of the TF-FVIIa complex (Figure 1A).
TF PCA is also regulated by oxidation of an alloste-
ric disulfide bond (Figure 1B). TF has 2 pairs of disulfide
bonds, one in the N-terminal (Cys49-Cys57) and the other
in the C-terminal (Cys186-Cys209) domain. An early study
demonstrated that TF PCA was reduced ≈3-fold when
either Cys186 or Cys209 were mutated whereas there
was no effect of mutating either Cys49 or Cys57.94 This
suggested that formation of the Cys186-Cys209 disulfide
bond was important for TF PCA. A later study proposed
that cryptic TF contains unpaired cysteine thiols at posi-
tions 186 and 209, and that activation involves formation
of a Cys186-Cys209 disulfide bond.95 Protein disulfide
isomerase has been proposed to be an important regula-
tor of TF redox state and activity.96–98 For instance, inhi-
bition of protein disulfide isomerase reduces TF PCA in
 712   Arterioscler Thromb Vasc Biol   April 2018
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Figure 1. Mechanisms of tissue factor (TF) de-encryption. A, The distribution of phospholipids in the plasma membrane of resting cells
is asymmetrical and contains phosphatidylcholine (PC) and sphingomyelin (SM) primarily in the outer leaflet and phosphatidylinositol (PI),
phosphatidylserine (PS) and phosphatidylethanolamine (PE) in the inner leaflet. The presence of SM and PC in the outer leaflet maintains
TF in an encrypted state. Upon cell activation phospholipid scramblase mediates transfer of PS to the outer leaflet. Cellular activation also
leads to release of acid sphingomyelinase (ASM) that catalyzes the conversion of SM to ceramide (Cer). These 2 processes lead to de-
encryption of TF increasing its cofactor activity and enabling efficient activation of FX by the TF-FVIIa complex. B, Encrypted TF contains
2 free thiols at Cys186 and Cys209. It is proposed that protein disulfide isomerase (PDI) catalyzes the formation of an allosteric disulfide
bond (Cys186-Cys209) that de-encrypts TF and enhances its procoagulant activity.

vitro.97,99 However, there are several unresolved issues with
the allosteric disulfide bond hypothesis. First, the disul-
fide bond must be formed before binding of FVII/FVIIa
to TF because this region of TF is buried at the interface
between TF and FVII/FVIIa.100 Second, as noted by Bach
and Monroe100 “there is no direct quantitative evidence for
free thiols in TF.” Finally, protein disulfide isomerase may
also indirectly affect TF PCA by promoting exposure of
phosphatidylserine.101 It should be noted that studies of de-
encryption of TF PCA have largely used cultured cells and
it is difficult to know what mechanism(s) occur in vivo.
TF Inhibitors
After the cloning of the TF cDNA in the late 1980s there was
a lot of excitement about the prospect of using TF inhibitors
as new antithrombotics.102 This is summarized by a com-
ment by Dr Golino in 2002 “many experimental studies have
demonstrated that inhibitors of TF:FVIIa PCA are powerful
inhibitors of in vivo thrombosis and that this approach usually
results in less pronounced bleeding tendency, as compared
with other more classical antithrombotic interventions.”102
There are 5 main classes of human TF (hTF) inhibitors: anti-
TF antibodies that either compete with FVII/VIIa binding or
bind to the TF-FVIIa complex, recombinant TFPI, active site-
inhibited FVIIa (FVIIai), nematode anticoagulant protein C2
(NAPc2), and Ixolaris. NAPc2 is an 85-amino acid protein
isolated from the hematophagous hookworm.103 It acts in a
similar manner to TFPI by first binding to FXa and then the
NAPc2-FXa complex binds to TF-FVIIa. Ixolaris was isolated
from a tick salivary gland and contains 2 Kunitz domains.104
Unlike TFPI, it binds to zymogen FX and then the complex
inhibits the TF-FVIIa complex.
These inhibitors have been used in preclinical and
clinical studies of acute coronary syndromes, sepsis, HIV
infection, and acute lung injury/acute respiratory distress
syndrome. The effect of NAPc2 was examined in patients
with non–ST-segment–elevation acute coronary syndromes
and seems to be safe because it did not significantly affect
major or minor bleeding (3.7% versus 2.5%).105 However, no
further studies have been performed using NAPc2. A recent
study showed that Ixolaris reduced levels of plasma D-dimer
in monkeys infected with simian immunodeficiency virus.106
Recombinant TFPI reduced lethality in a baboon model of
sepsis.107 However, there was no effect on all-cause mortal-
ity in patients with severe sepsis and there was an increased
risk of bleeding.108 Early administration of a monoclonal
 Grover and Mackman   Tissue Factor in Hemostasis and Thrombosis   713

antibody against TF reduced sepsis-induced respiratory

and renal failure in baboons.109 The success of this preclini-
cal model led to a clinical trial that examined the effect of
FVIIai in patients with acute lung injury/acute respiratory
distress syndrome. Unfortunately, the trial was stopped
early by the Safety Committee because of increased mortal-
ity in the group receiving the highest dose of FVIIai.110,111 In
addition, patients with the higher doses of FVIIai also had a
trend to increased risk of serious bleeding. In retrospect, given
the central role of TF in lung hemostasis, these results are not
unexpected. Based on these studies, it seems unlikely that TF
inhibitors will be used clinically in the future.

TF-FVIIa-Dependent Signaling
The TF-FVIIa complex has both PCA and signaling activity.
The localization of TF on the cell surface seems to regulate
these functions with the signaling form of TF being located
in lipid rafts.112 The signaling form of the TF-FVIIa complex
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also interacts with β1 integrins.113 A recent study identified the

binding motif in the FVIIa protease domain that mediates the
interaction with β1.114
The TF-FVIIa complex and downstream coagulation
proteases, such as thrombin, activate cells by cleavage of G
protein-coupled PARs. This family consists of 4 members,
PAR1–4.115 PAR2 can be activated by the TF-FVIIa and
TF-FVIIa-FXa complexes as well as FXa, whereas PAR1 is
activated by thrombin and FXa.116 TF-FVIIa-dependent sig-
naling has been mainly studied in cancer.117 Studies have
shown that PAR2 activation by TF-FVIIa induces proangio-
genic and immune modulating cytokines, whereas PAR1 acti-
vation by thrombin or the TF-FVIIa-FXa complex contributes
to the metastatic tumor environment.117
Alternatively Spliced TF
A soluble form of TF can be generated by alternative splicing
of the TF gene where exon 5 is skipped.118,119 This alternatively
spliced TF lacks residues required for interaction with FX, the
allosteric disulfide in the C-module and the transmembrane
domain that dramatically reduce its PCA.120–122 However,
alternatively spliced TF has PCA in the presence of phospho-
lipids.123 Moreover, alternatively spliced TF plays a role in
cancer biology by promoting angiogenesis and proliferation
of tumor cells via interacting with integrins containing β1 and
β3.124,125
Hemostasis
Hemostatic Defects in Mice and Humans
Deficient in Different Coagulation Proteins
Factors of the coagulation cascade can be divided into 4 groups
based on the phenotype of mice deficient in each of the factors
(Figure 2). Group 1 contains factors of the extrinsic pathway
(TF, FVII, FV, FX, and FII). Complete deficiencies of any of
these proteins in mice leads to universal death during embry-
onic development or shortly after birth.19–21,126–130 Group 2 con-
tains FVIII and FIX. Complete deficiencies of these proteins
in mice is not lethal but is associated with impaired hemo-
stasis in a tail transection model and a saphenous vein injury
Figure 2. Role of different coagulation proteins in hemostasis
and thrombosis. Based on the hemostatic defects of mice with
complete deficiencies in the different coagulation factors, we can
divide the coagulation factors into 4 groups. Group 1 consists of
factors in the extrinsic pathway (tissue factor [TF], factor [F] VII,
FV, FX, FII). Group 2 consists of FIX and FVIII. Group 3 consists
of FXI. Group 4 consists of FXII. Pathological TF expression and
FXII can trigger thrombosis.
model.131–135 Group 3 contains FXI. A complete deficiency of
this protein in mice is not lethal but these mice have a mild
hemostatic defect in a saphenous vein injury model.135,136
Group 4 contains FXII. In contrast to the hemostatic defects
in mice with deficiencies in proteins in the other groups, mice
with a complete deficiency in FXII have no hemostatic defects
(Figure 2).137 However, FXII-deficient mice are protected in
thrombosis models.138,139
Unlike mouse studies where gene inactivation leads to a
complete deficiency of a given protein, humans with deficien-
cies in different coagulation proteins usually have residual
amounts of functional protein. To date, there are no reports
of humans with a deficiency of TF. In contrast, rare inherited
bleeding disorders occur because of deficiencies in FII, FVII,
FX, FXI, and FXIII,140 although the estimated prevalence of
these disorders is very low (between 1 and 2 per 106 individu-
als).140 Individuals with deficiencies in FII, FX, or FV most
often bleed in the skin and mucus membranes and those with
the lowest levels of the different coagulation factors have intra-
cranial bleeds.140 In terms of the coagulation cascade, the most
frequent congenital bleeding disorders in the general human
population are caused by deficiencies in FVIII (hemophilia A)
and FIX (hemophilia B). The prevalence rates of these bleed-
ing disorders is 1/5000 male births.140 Hemophilia A and B
patients are prone to spontaneous bleeding in the joints and
skeletal muscle.141 FXI deficiency is rare (1 per 100 000) and
individuals with FXI deficiency rarely exhibit spontaneous
bleeding but have an elevated risk of bleeding after surgery
such as tooth extraction.142–144
 714   Arterioscler Thromb Vasc Biol   April 2018

TF as a Hemostatic Envelope Around Blood Table 2.  Hemostatic Defects in Low Tissue Factor Mice
Vessels and Body Surfaces Phenotype References
The TF-FVII complex plays an essential role in hemostasis.
Spontaneous
TF is expressed by perivascular cells, such as adventitial fibro-
blasts and pericytes, and at body surfaces, such as keratino-  Lung Fatal hemorrhage and 154,155
cytes in the skin.79,84,145 Drake et al79 proposed that TF formed a hemosiderin
hemostatic envelope around blood vessels and body surfaces.  Brain Fatal hemorrhage 152,154
Initially, it was thought that the physical separation of TF from  Uterus Fatal postpartum hemorrhage 156
blood meant that the TF-FVIIa complex was only formed after
 Placenta Blood pools in the placenta 156
vascular injury. However, a recent study reported that perivas-
cular TF was prebound to FVII/FVIIa in the dermal vascu-  Testis Hemorrhage and calcification 2,154
lature.146 Indeed, coagulation factors, such as FVII, leak into  Heart Hemorrhage and fibrosis 157,158
the extravascular space at levels roughly proportional to their
 Intestine Hemorrhage 152,154
size.147,148 The presence of a preformed TF-FVIIa complex
would allow rapid activation of clotting after vessel injury. In Induced
contrast to dermal vessels, there is a high proportion of free TF  Intratracheal Increased hemorrhage 159
(non-FVII bound) present in the brain parenchyma and cere- lipopolysaccharide
bral vessels.149 This difference is likely because of the lower   Influenza A infection Increased hemorrhage 160
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permeability of the blood–brain barrier. The high levels of TF

 Collagenase-induced Increased hemorrhage 73
in the brain may compensate for this the lower level of pre-
intracerebral hemorrhage
formed TF-FVIIa complex around the cerebral vasculature.
  Tail vein transection Prolonged bleeding 152

Use of Transgenic Mice to Study
the Role of TF in Hemostasis
As noted above, TF−/− and FVII−/− mice die during embryonic
development or shortly after birth, which means that they
cannot be used to study the roles of the TF-FVIIa complex
in hemostasis in different organs in adult mice.19–21,126 To cir-
cumvent the lethal phenotype of these mice, we and others
have generated mice that express low levels of either TF or
FVII.150,151 We rescued mTF−/− embryos by expressing hTF
from a hTF minigene.150 These mice (mTF−/−, hTF+ minigene)
are referred to as low TF mice and express ≈1% to 2% of wild-
type TF levels. Rosen et al151 generated transgenic mice with
low levels of FVII by targeted replacement of the FVII gene
with a FVII cDNA. Remarkably, the phenotypes of low TF
and low FVII mice are similar supporting the notion for a cen-
tral role for the TF-FVIIa complex in hemostasis.152,153
Consistent with a role for TF in hemostasis, low TF mice
exhibit prolonged bleeding after various hemostatic chal-
lenges. For instance, low TF mice have increased bleed-
ing after tail transection (Table 2).152 In addition, low TF
mice had prolonged bleeding times in the saphenous vein
mechanical injury model and after repeated vessel injury in
the saphenous vein laser injury model161 (Table 2; unpub-
lished data, 2018). Interestingly, the hemorrhage observed in
low TF mice in these models is not as severe as FIX−/− mice.
Low TF mice also have excess bleeding after a full thickness
skin punch.162
We determined the effect of combining low levels of TF
with a complete deficiency of other procoagulant factors or
PAR4, which is the main thrombin receptor on mouse plate-
lets. Consistent with the importance of FIX in hemostasis,
mice with a combination of low levels of TF and an absence
of FIX do not survive to wean.163 Our recent studies indicate
that low TF, FIX−/− mice survive embryonic development but
die shortly after birth (Grover and Mackman, unpublished
data, 2018). In contrast, we found that the majority of low TF,
 Saphenous vein laser Prolonged bleeding 161
injury model
 Saphenous vein Prolonged bleeding Unpublished
mechanical injury model
  Skin punch Delayed wound healing 162
FXI−/− mice survive to wean (Grover and Mackman, unpub-
lished data, 2018).
Mice with low levels of TF but lacking PAR4 survive to
wean but exhibit reduced survival because of fatal bleeds in
the lung and brain compared with low TF, PAR4+/+ controls.154
Interestingly, the bleeding phenotype observed in low TF mice
in the laser injury model was increased after administration
of the P2Y12 inhibitor clopidogrel at a dose that had a mini-
mal effect on bleeding time in wild-type mice.161 These data
suggest that TF-dependent thrombin generation leads to both
fibrin deposition and platelet activation that act co-operatively
to form and stabilize the hemostatic plug.
Two other transgenic mouse lines have been made that
express hTF in place of mouse TF. Chromosomal vector hTF
mice (mTF−/−, hTF+ chromosomal vector) express hTF from
a vector consisting of the majority of chromosome 1, whereas
hTF knockin mice were generated by gene targeting that
replaced the mouse TF gene with the hTF gene.157,164 Mouse
FVIIa binds with high affinity to both mouse and hTF but the
association of human FVIIa with mouse TF is significantly
lower than with hTF.165 Indeed, we found a 6-fold reduction
in FXa generation when we used human FVIIa with mouse TF
compared with hTF.166 These studies indicate that hTF can be
used to replace mouse TF but not vice versa, although we do
not know about interactions of hTF with mouse β1 integrins.
Finally, we generated mice with a floxed TF gene (TFfl/fl) that
when crossed with mice expressing Cre recombinase under the
control of a specific promoter leads to cell type-specific dele-
tion of the TF gene (Table 3).157
 Grover and Mackman   Tissue Factor in Hemostasis and Thrombosis   715

Table 3.  Transgenic Mice With Tissue Factor Deleted in Tissue-Specific Hemostasis
Different Cell Types TF is expressed in an organ-specific manner with high levels
Cre Line Cell Type Phenotype References in the brain, lung, heart, uterus, and placenta, and low levels
in skeletal muscle and joints.183 This pattern of expression
Mlc2v Cardiomyocyte Cardiac fibrosis 157
suggests that TF provides additional hemostatic protection
SM22 Vascular smooth Reduced carotid arterial 167 to a select group of vital organs. For instance, blood ves-
muscle cell thrombosis
sels in the heart and lung may need additional hemostatic
LysM Myeloid cell Reduced 168 protection because they are more likely to be injured by the
lipopolysaccharide- mechanical movement of these organs. In contrast, tissues
induced coagulation that have low levels of TF are likely more dependent on
Tie2 Endothelial and Reduced 168 the intrinsic pathway to generate thrombin for hemostasis.
hematopoietic cell lipopolysaccharide- Indeed, hemophilia A and B patients are prone to bleeding
induced coagulation into joints and skeletal muscle and inhibitor patients receive
Alb Hepatocyte Reduced acute 169,170 FVIIa as a bypass agent to boost the extrinsic pathway. Low
liver injury induced TF mice have been used to study the role of TF in tissue-
coagulation specific hemostasis and exhibit spontaneous hemorrhaging
SPC Lung epithelial Reduced acute 160,171 in various tissues (Table 1).
lung injury induced
coagulation
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Nestin Neuroectodermal Increased collagenase- 172
induced intracerebral
hemorrhage
K14 Keratinocyte Delayed wound healing In
preparation
Hemostasis Requires a Balance
Between TF and TFPI
TFPI is the major physiological inhibitor of the TF-FVIIa com-
plex and shuts down the initiation of the clotting cascade.173–177
There are 2 forms of TFPI: TFPIα and TFPIβ. TFPIα contains
3 Kunitz domains where domain 1 binds to FVIIa and domain
2 binds to FXa. TFPIβ is alternatively spliced form that con-
tains Kunitz 1 and 2 with a different C-terminus that mediates
anchoring to the membrane via a glycosylphosphatidyl inosi-
tol linkage. Similar to a TF−/− mice, a complete deficiency of
TFPI is universally lethal during embryonic development.178
TFPI−/− embryos had intravascular thrombi and fibrin(ogen)
deposition in the liver and seem to exhibit a consumptive
coagulopathy with secondary bleeding. There are no reports
of TFPI-deficient humans.
The effect of reducing the level of FVII on the survival
of TFPI−/− embryos was evaluated by intercrossing FVII+/−,
TFPI+/− mice.179 FVII+/−, TFPI−/− mice were rescued from
embryonic lethality but died shortly after birth because of
consumptive coagulopathy.
Similarly, FVII−/−, TFPI−/− survived embryonic develop-
ment but succumbed to perinatal bleeding because of an
absence of FVII. We found that low levels of TF rescued
TFPI−/− embryos from embryonic lethality and allowed the
generation of viable TFPI−/− adult mice that survived for
>6 months without any adverse phenotype.155 Conversely,
reducing the levels of TFPI dose-dependently reduced the
hemostatic defects in low TF mice.155 A similar principle of
rebalancing clotting has been applied to restore hemostasis in
hemophilic mice. For instance, inhibition of TFPI in hemo-
philia A mice reduced bleeding in a tail vein model.180 TFPI
inhibitors are currently being developed for the treatment of
patients with hemophilia.181,182
Lung
The lung contains high levels of TF.184 Epithelial cells are
the primary site of TF expression in the lung.61 Mice with TF
deleted in epithelial cells (TFfl/fl, SPCcre; Table 3) have sig-
nificantly reduced baseline levels of TF compared with con-
trols.160,171 LPS and influenza A virus infection increase TF
expression in the lung of mice and specifically in epithelial
cells.61,159,160,171,184 LPS and influenza A virus did not increase
TF expression in TFfl/fl, SPCcre mice. These studies indicate
that the increased TF expression in epithelial cells is response
to infection may provide additional hemostatic protection in
the face of an infection.
Low TF mice exhibit a high incidence of fatal lung hem-
orrhages (≈50%) when backcrossed 7 generations onto the
prohemorrhagic C57BL/6 background. (Table 2).154,155 This
indicates that lung hemostasis is impaired in low TF mice.
In addition, our standard low TF (backcrossed 6 genera-
tions to the C57BL/6 background) exhibit increased alveolar
hemorrhage compared with control mice when challenged
with intratracheal LPS (Table 2).159 As expected, both low
TF and TFfl/fl, SPCcre mice had increased alveolar hemor-
rhage after influenza A virus infection (Tables 2 and 3).160
However, significantly more bleeding was observed in low
TF mice compared with TFfl/fl, SPCcre mice, and we hypoth-
esize that additional cell types, such as fibroblasts, also con-
tribute to hemostasis in the lung. We are currently testing this
hypothesis.
Brain
The brain contains high levels of TF with astrocytes being
the major source.184,185 Low TF mice occasionally succumb
to spontaneous fatal brain hemorrhages (Table 2).152,154 In
addition, administration of an antihuman TF antibody to TF
knockin mice expressing hTF resulted in cerebral hemor-
rhage.164 Importantly, FVII−/− and FII−/− neonates also have
fatal cerebral hemorrhages.126,129,130 In addition, genetic elimi-
nation of FII in adult mice led to spontaneous hemorrhage in
the brain.186,187
Recently, the Pawlinski group generated mice with TF
deleted in all neuronal cells (TFfl/fl, nestin-cre; Table 3).172
 716   Arterioscler Thromb Vasc Biol   April 2018
These mice express a very low level of TF (≈1%) in their
brains compared with a slightly higher level of TF (≈2%) in
the brains of low TF mice. A model of collagenase-induced
intracerebral hemorrhage was used to determine the role of
TF in brain hemostasis. Both low TF and TFfl/fl, nestin-cre
mice exhibited increased intracerebral hemorrhage com-
pared with controls.172 However, more hemorrhage was
observed in TFfl/fl, nestin-cre compared with low TF mice
and this was associated with death of some of these mice.
These studies suggest that the high levels of TF in the brain
are required to limit intracerebral hemorrhage after trau-
matic brain injury.
Heart
TF is expressed by both cardiac fibroblasts and cardiac myo-
cytes in the heart.184 Administration of an inhibitory anti-
human TF antibody to TF knockin mice expressing HTF
resulted in cardiac hemorrhage.164 We have observed occa-
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sional spontaneous hemorrhages in the heart of low TF mice
and hearts of older low TF mice are fibrotic.158 Mice with
only ≈20% expression of hTF in the heart and those with
TF deleted in cardiac myocytes (TFfl/fl, Mlc2v; Table 3) also
exhibit cardiac fibrosis.157 Importantly, a similar pattern of
cardiac fibrosis has been observed in low FVII, low FX, and
low FII expressing mice but not FIX deficient mice.133,153,188,189
Spontaneous hemorrhage was also observed in the heart
after genetic elimination of prothrombin in adult mice.186
These results indicate that the extrinsic pathway, but not the
intrinsic pathway, mediates cardiac hemostasis.153,158,186,188,189
Uterus, Placenta, and Testis
Female low TF mice exhibit excess bleeding after delivery and
14% of the mice die, which indicates that TF is required for
uterine epithelium hemostasis postpartum (Table 2).156Low
TF placentas contain blood pools in the labyrinth (Table 2).156
At present, it is unclear whether this is because of a hemo-
static defect or a structural defect in the placental labyrinth.
Interestingly, FVIII−/− and FIX−/− mice have no hemostatic
defects in the uterus or placenta.132,133
We observed that male low TF mice lost fecundity at a
relatively young age and had shrunken and fibrotic testes.
We also observed spontaneous hemorrhage in the testes of
low TF mice that may have been induced by trauma or dur-
ing retraction of the testes into the abdomen. We think that
hemorrhage leads to fibrosis of the testes and the loss of
fertility.
Thrombosis
Thrombosis is a pathological process that occludes blood flow
in vessels. TF is involved in the pathogenesis of both arte-
rial and venous thrombosis.190 Sources of TF that activate
coagulation in different diseases include macrophages and
microvesicles in atherosclerotic plaques, activated cells within
the vasculature and perivascular cells that are exposed because
of an increase in vascular permeability.
There is strong evidence that monocyte TF contributes to
thrombosis in many diseases (Table 4).
Table 4.  Cellular Sources of Tissue Factor That Drive
Thrombosis and Disseminated Intravascular Coagulation
Cellular Source
Disease Model of Tissue Factor References
Atherosclerosis Macrophage foam cells and 191–193
macrophage-derived microvesicles
Sepsis/endotoxemia Monocytes 39,40,168
Surgery Peripheral blood mononuclear cells 194
Cancer Tumor cell-derived microvesicles 195–197
Antiphospholipid Monocytes 198–200
syndrome
Liver injury Hepatocytes 169,170
Sickle cell disease Monocytes 73,201
Viral infection Peripheral blood mononuclear 202,203
cells/monocytes
Atherosclerosis
In humans, rupture of advanced atherosclerotic lesions trig-
gers arterial thrombosis leading to myocardial infarction
and stroke. Atheroma contain high levels of TF that likely
plays a central role in atherothrombosis (Figure 3A).191,204–210
Macrophage foam cells are the main source of TF in plaques
with apoptotic macrophages releasing TF-positive microves-
icles into the necrotic core.192,193,211 In addition, patients with
acute coronary syndromes have increased levels of TF expres-
sion in monocytes, platelets, and leukocyte-platelet aggre-
gates that may also contribute to thrombosis after plaque
rupture.48,212 Many studies have reported increased levels of
TF antigen in the plasma of cardiovascular disease patients.213
However, we could not detect increased levels of microvesicle
TF activity in these patients.214
In mice a 50% reduction of TFPI increased thrombo-
sis after photochemical disruption of carotid artery plaques
in ApoE−/− mice, presumably because of an increase in TF
PCA.215 Importantly, inhibition of TF with FVIIai markedly
reduced thrombosis after ultrasonic disruption of carotid
plaques in ApoE−/− mice.216 Similarly, FVIIai or recombinant
TFPI reduced thrombosis after balloon-mediated disrup-
tion of carotid plaques in hypercholesterolemic rabbits.217,218
These studies indicate that TF within atherosclerotic plaques
plays a major role in the formation of occlusive thrombi after
rupture.219
Sepsis/Endotoxemia
Disseminated intravascular coagulation is a life-threatening
disorder caused by diffuse activation of coagulation that
can result in microvascular occlusion and organ dysfunc-
tion.220 Disseminated intravascular coagulation is frequently
observed in patients with bacterial sepsis. TF has been shown
to play a central role in disseminated intravascular coagulation
(Figure 3B).221 In a human model of LPS-induced endotoxemia
TF mRNA and TF protein expression were induced in blood
cells (presumably monocytes) and this was associated with
increased levels of thrombin-antithrombin complex, a marker
of activation of coagulation.39 TF expression is also observed
 Grover and Mackman   Tissue Factor in Hemostasis and Thrombosis   717
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Figure 3. Cellular sources of tissue factor (TF) that trigger thrombosis and disseminated intravascular coagulation. A, TF within athero-
sclerotic plaques triggers thrombosis after plaque rupture. Other sources of TF in patients with cardiovascular disease include circulating
activated monocytes and activated platelets. B, In endotoxemia and sepsis, pathogens induce TF expression in monocytes and endothe-
lial cells and the release of TF-positive microvesicles (MVs). In addition, increased vascular permeability exposes TF on perivascular cells
to blood. All these sources of TF may contribute to disseminated intravascular coagulation. C, Pancreatic tumors express high levels of
TF and release TF-positive MVs that circulate in the blood and trigger venous thrombosis. D, Antiphospholipid antibodies (aPLA) induce
TF expression in monocytes and the release of TF-positive MVs that trigger thrombosis. TF expression by endothelial cells may also con-
tribute to local thrombosis. Administration of the antioxidant ubiquinol reduced Coenyzme Q10 (CoQ10) reduces monocyte TF expression.

in monocytes isolated from septic baboons.40 As noted earlier,
immunohistochemistry studies demonstrated TF staining on
ECs in the spleen and aorta of septic baboons and ECs in the
glomerulus of endotoxemic mice.38,80,83 Therefore, EC TF may
also play a role the activation of coagulation during sepsis and
endotoxemia. Importantly, early inhibition of TF reduced the
activation of coagulation and mortality in septic baboons.107,222
We analyzed the role of TF in the activation of coagulation
in a mouse model of endotoxemia. As expected, LPS activa-
tion of coagulation was lower in low TF mice compared with
controls.223 Similarly, wild-type mice containing bone marrow
from low TF mice or mice with TF deleted in myeloid cells
(TFfl/fl, LysMCre; Table 2) had lower levels of activation of
coagulation compared with controls.168 However, bone mar-
row cells only account for ≈50% of TF that activates coagula-
tion in endotoxemic mice. We found that deleting TF on ECs
did not affect activation of coagulation in a mouse endotox-
emia model.168 Another study found that inducible expression
of a degradation resistant form of IκBα in ECs blocked the
induction of TF expression in ECs in the glomerulus of endo-
toxemic mice.83 However, the specific impact of this blockade
of EC TF expression could not be evaluated because expres-
sion of the mutant IκBα also prevented the downregulation
of the anticoagulant proteins EC protein C receptor and
thrombomodulin.83 Taken together, we propose that activated
monocytes are the major source of TF that activates coagula-
tion during endotoxemia and sepsis. However, we speculate
that TF expressed on perivascular cells exposed by increased
vascular permeability and possibly TF induced on ECs also
contribute to the activation of coagulation in sepsis.
Surgery
Surgery is associated with an increased risk of venous throm-
bosis. Three studies have reported an increase in monocyte
TF expression 1 day after surgery to remove tumors as well
as after coronary artery bypass surgery and total knee arthro-
plasty.194,224,225 This suggests that surgery triggers an inflamma-
tory response surgery that induces monocyte TF expression.
Importantly, the increase in monocyte TF occurs before the
peak of clinical venous thrombosis, which is 7 days after sur-
gery.226 These studies suggest that induction of TF expression
on monocytes contributes to postsurgery venous thrombosis.
Cancer
Thromboembolism represents a leading cause of death in
patients with cancer and is associated with poor prognosis.227
 718   Arterioscler Thromb Vasc Biol   April 2018
Venous thrombosis, in particular, is frequently observed
in cancer patients with reported annual incidences ranging
between 1.5% to 20% per annum depending on the site of can-
cer and treatment.228 In vitro, cancer cell lines shed TF-positive
microvesicles.68,229–231 Moreover, increased levels of circulat-
ing TF-positive microvesicles are observed in patient with dif-
ferent types of cancer, including pancreatic, brain, colorectal,
lung, breast, and prostate.195,232–235 Importantly, elevated levels
of MV TF activity are associated with venous thromboembo-
lism in pancreatic cancer patients but not in brain, gastric, or
colorectal cancer patients.195–197,228
Preclinical studies of cancer-associated thrombosis have
largely focused on pancreatic cancer, given the high rates of
thrombosis observed in patients with this type of malignancy.
Immunocompetent and immunodeficient mouse models have
been used to study the role of tumor-derived, TF-positive
microvesicles in thrombosis (Figure 3C). Studies have
used the mouse pancreatic cell line Panc02 that expresses
TF. Injection of mice with Panc02-derived, TF-positive
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microvesicles enhanced thrombosis in a laser injury model
and an infrarenal vena cava (IVC) stenosis model, whereas
microvesicles from Panc02 cells with reduced TF expres-
sion were less thrombotic.236,237 Moreover, immunocompetent
mice bearing Panc02 tumors had enhanced thrombus forma-
tion in a mesenteric ferric chloride model and an IVC stenosis
model.236,237
Immunodeficient mice are required for studies with
human pancreatic cancer cell lines. We and others have
used several TF-expressing human pancreatic cells lines,
including L3.6pl, HPAF-II, and BxPc-3.68,238,239 Injection
of TF-positive microvesicles derived from these cell lines
activated coagulation and increased thrombus formation
in the IVC stenosis model.68,238,239 In addition, inhibition of
TF on BxPc-3 microvesicles with an antihuman TF anti-
body abolished the enhanced thrombosis. Mice bearing
HPAF-II tumors also demonstrated markedly shortened
time to occlusion in a saphenous vein ferric chloride injury
model but there was no increase in thrombosis in the IVC
stenosis model.68 However, mice bearing BxPc-3 tumors
had an increase in thrombosis in the IVC stenosis model.238
Recently, we evaluated the contribution of tumor derived,
TF-positive microvesicles to venous thrombus in tumor
bearing mice.240 Mice with orthotopic BxPc-3 tumors had
larger thrombi in an IVC stasis model compared with con-
trols and administration of an antihuman TF antibody sig-
nificantly reduced the size of the thrombi.240 Taken together,
these studies suggest that tumors release TF-positive
microvesicles into the circulation and that these microvesi-
cles enhance venous thrombosis.
Antiphospholipid Syndrome
Antiphospholipid syndrome (APS) is an autoimmune disorder
caused by the development of antibodies against many phos-
pholipids, such as cardiolipin, phospholipid-associated pro-
teins, including β2-glycoprotein I, and plasma proteins, such
as prothrombin.241 Collectively, these antibodies are called
antiphospholipid antibodies (aPLA). APS patients with per-
sistently high titers of aPLA are at significant risk of arterial
and venous thromboembolic events and fetal loss.242
The exact mechanism by which aPLA induce a hyperco-
aguable state remains unclear. However, results from differ-
ent studies suggest that aPLA induction of TF expression in
monocytes may underpin the pathogenesis of APS-associated
thrombosis. Importantly, human monocytes isolated from
patients with APS express TF.198–200 Moreover, incubation
of primary human monocytes from healthy individuals with
anti-β2-glycoprotein I IgG leads to increased monocyte TF
gene expression, PCA and release of TF-positive microves-
icles.243–246 In vitro studies showed that aPLA induces oxi-
dative stress in monocytes, and levels of cell surface TF
exposure correlate with intracellular peroxide generation.247
Furthermore, pretreatment of the monocytes with the antioxi-
dant ubiquinol, reduced Coenzyme Q10, decreased oxidative
stress and TF expression.248 In a follow-up study, adminis-
tration of Coenzyme Q10 to APS patients reduced oxidative
stress and TF exposure on monocytes.248,249 Further clinical
studies are required to determine if antioxidant therapies can
effectively prevent thrombotic events in patients with APS by
reducing monocyte TF expression.
Animal models have been used to gain insights into
the involvement of TF in APS-associated thrombosis
(Figure 3D).250,251 For instance, low TF mice were protected
from the thrombotic microangiopathy induced by the pas-
sive transfer of a murine monoclonal aPLA or purified
patient aPLA.252 Interestingly, aPLA were shown to induce
TF expression in monocytes and ECs via a pathway involv-
ing endosomal NADPH oxidase and TLR7/8.253 Importantly
induction of TF was absent in aPLA-treated monocytes from
NOX2−/− mice. A follow-up study with mice showed that
aPLA enhanced thrombosis in an IVC stenosis model.254
Enhanced thrombosis in this study required NOX2 and leu-
kocytes but not TLR4 or platelets.250 These 2 studies sug-
gest a role for monocyte TF in aPLA induced thrombosis.254
Further studies are needed to understand how TF contributes
to thrombosis in patients with APS.
Acute Liver Injury and Cirrhosis
Acute liver injury and cirrhosis are associated with a pro-
thrombotic state. One study found very high levels of circulat-
ing MV TF activity in patients with acute liver injury.255 We
found that MV TF activity was increased with cirrhosis and
levels were associated with the Child–Pugh score and markers
of activation of coagulation and fibrin degradation.256
The liver expresses low levels of TF compared with other
organs, such as the brain and lung. The Luyendyk group deter-
mined the contribution of hepatocytes to basal TF expression
in the liver by comparing levels in mice with TF deleted in
hepatocytes (TFfl/fl, Albcre) to controls (Table 3). Importantly,
we found a dramatic reduction of TF in TFfl/fl, Albcre mice
indicating that hepatocytes are the major source of TF in
the liver.169 Furthermore, administration of acetaminophen
to wild-type but not TFfl/fl, Albcre mice led to a rapid activa-
tion of coagulation as measured by increased levels of plasma
thrombin-antithrombin complexes.169 In a second study, we
used a mouse model of bile duct ligation to determine the cel-
lular source of TF responsible for activation of coagulation in
chronic liver injury. As expected, activation of coagulation was
lower in low TF mice compared with controls.170 Interestingly,
 Grover and Mackman   Tissue Factor in Hemostasis and Thrombosis   719
activation of coagulation was reduced by deletion of TF in
hepatocytes but not myeloid cells.170 These 2 mouse models
indicate that de-encryption of hepatocyte TF mediates the
rapid activation of coagulation during liver injury.
Sickle Cell Disease
SCD is an autosomal recessive disorder caused by a single
point mutation in the β-globin gene. Under hypoxic condi-
tions aggregation of the hemoglobin variant leads to the for-
mation of sickled erythrocytes. Patients with this disorder have
an increased risk of both arterial and venous thromboembolic
events.257–259 SCD patients have increased numbers of mono-
cyte- and EC-derived TF-positive microvesicles and levels
were positively correlated with both thrombin-antithrombin
and D-Dimer, which suggested that these particles may con-
tribute to the procoagulant state observed in SCD.201
Mouse models have been used to study the role of TF
in the activation of coagulation and inflammation in SCD.
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Inhibition of TF reduced the chronic procoagulant state and
inflammation observed in SCD mice.73 Furthermore, SCD
mice had increased thrombosis in a light-dye–mediated injury
to the cerebral microcirculation and this was inhibited by an
inhibitory anti-TF antibody.260 Several studies have investi-
gated the cellular sources of TF expression that contribute to
the activation of coagulation in SCD mice.261 TF expression
has been reported in monocytes, neutrophils, and ECs in SCD
mice.73,81,82 An early study observed increased TF expression
on pulmonary veins of moderate and severe models of SCD
whereas hypoxia and reoxygenation was required to increase
EC TF expression in a mild model of SCD.81 We found that
deletion of TF in ECs did not affect activation of coagulation
in mice with sickle cell bone marrow but reduced levels of
interleukin6.262 Similar results were observed in SCD mice
that received the FXa inhibitor rivaroxaban and SCD mice
lacking PAR2 in nonhematopoietic cells.262 These data sug-
gest that a TF-FXa-PAR2 complex on ECs does not contribute
to the activation of coagulation but drives interleukin6 expres-
sion in SCD.
Viral Infections
Viral infections are associated with activation of coagulation
and thrombosis. For instance, infection with hemorrhagic
fever viruses, such as Ebola virus, is associated with DIC,
whereas HIV infection increases the risk of thrombosis.
Viruses induce TF expression in a variety of cells, includ-
ing monocytes and ECs.263 Importantly, Ebola virus induced
TF mRNA expression in peripheral blood mononuclear
cells in vitro and peripheral blood mononuclear cells iso-
lated from infected monkeys.202 Moreover, inhibition of TF
reduced mortality in monkeys infected with Ebola virus.264
Similarly, monocyte TF expression correlated with levels of
HIV and D-dimer in plasma from HIV infected patients.203
More recently, it was shown that TF is expressed by a sub-
set of monocytes (CD14+, CD16−, CCR2+) in HIV infected
patients, and inhibition of TF reduced D-dimer levels in mon-
keys infected with simian immunodeficiency virus.106 These
studies suggest that TF expression by monocyte drives the
coagulopathy in HIV infected patients.
Conclusions
It is clear that the TF-FVIIa complex is essential for hemostasis
and provides additional protection to vital organs, such as the
heart, lung, and brain. Further studies are needed to understand
the cell types that express TF in different tissues and control
hemostasis under basal and pathological conditions. TF con-
tributes to arterial and venous thrombosis. TF also plays a
major role in the formation of thrombi after rupture of athero-
sclerotic plaques. Infection induces TF expression on mono-
cytes and this can trigger DIC. Inhibition of all sources of TF is
associated with increased bleeding. However, targeted inhibi-
tion of pathological TF expression, for instance in monocytes,
may reduce thrombosis associated with different diseases.
Acknowledgments
We would like to acknowledge Drs D. Monroe, J. Erlich, and Y.
Hisada for helpful comments.
Sources of Funding
This work was supported by funding from the John C. Parker
Professorship.
Disclosures
None.
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 Tissue Factor: An Essential Mediator of Hemostasis and Trigger of Thrombosis
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Steven P. Grover and Nigel Mackman

Arterioscler Thromb Vasc Biol. 2018;38:709-725; originally published online February 8, 2018;
doi: 10.1161/ATVBAHA.117.309846
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