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l l l THROMBOSIS AND HEMOSTASIS
Comment on Heestermans et al, page 2630
Mouse models of venous
thrombosis
are not equal
----------------------------------------------------------------------------------------------------Nigel Mackman
UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL
In this issue of Blood, Heestermans et al1 evaluated the roles of platelets, neutrophils,
and factor XII (FXII) in a new mouse model of spontaneous venous thrombosis.
V
enous thrombosis is a major medical
problem in industrialized countries.2,3 It
is multifactorial with both genetic and acquired
risk factors. Genetic factors include either
deficiencies in the anticoagulants antithrombin,
protein C, or protein S, or the presence of
FV or prothrombin variants (FV Leiden and
prothrombin 20210A).2 Acquired risk factors
include older age (older than age 50 years),
use of hormonal therapy (birth control or
replacement), trauma, major surgery,
Immune
thrombus
Vessel wall
Endothelium
prolonged immobilization, and certain
diseases, such as cancer and antiphospholipid
antibody syndrome.3 Vein damage and/or
activation of the endothelium, changes in blood
flow, and the presence of procoagulant factors
in blood appear to be the triggers of venous
thrombosis. In patients with pancreatic cancer,
tumor-derived, tissue factor–positive
extracellular vesicles may induce thrombosis,
whereas in patients with artificial heart valves,
FXII may drive thrombosis. Most venous
NETs
Platelet
FXII
Neutrophil
?
Blood
Thrombus
Comparison of two mouse models of venous thrombosis. In the inferior vena cava stenosis model of venous thrombosis, the
inferior vena cava is ligated to reduce blood flow by ;90%. Platelets, FXII, and neutrophils contribute to immune thrombosis. In
the spontaneous venous thrombosis model, mice are treated with small interfering RNAs to reduce levels of the anticoagulants
antithrombin and protein C. Platelets contribute to thrombosis in this model, whereas there is no role for neutrophils, and
decreasing FXII increases thrombosis. NET, neutrophil extracellular trap. Professional illustration by Somersault18:24.
2510
thrombi are thought to originate in valve
pockets as a result of the low oxygen tension,
and they develop over a period of hours to days.
Mouse models are used to provide new
insights to human disease. There are a variety
of mouse models of venous thrombosis
available to researchers that use large (inferior
vena cava [IVC]), medium (femoral vein), or
small (mesenteric and cremaster venules)
vessels.4 The IVC has become the most popular
vessel for studying venous thrombosis because
it is the largest vein available in the mouse.
Some mouse models are better than others.
Adding ferric chloride to the IVC and
measuring thrombosis at 30 minutes could be
considered a good model of trauma-induced
thrombosis, but is not a good model of
nontrauma-related venous thrombosis in
humans. Although complete ligation of the
IVC leads to reproducible thrombosis, it is not
ideal for studying nontrauma-related venous
thrombosis because of the absence of blood
flow.
The IVC stenosis model reduces blood flow
by ;90% via partial ligation of the IVC, and
thrombi form upstream of the site of stenosis
because of activation of the endothelium by
the disturbed blood flow and possibly because
of some vessel injury induced during the
surgery.5,6 Thrombi can be detected as early as
3 hours but are usually measured at 48 hours.
Von Bruhl et al6 described roles for several
different cell types and clotting factors in
this model, including platelets, FXII, and
neutrophils (see figure). Indeed, the
contribution of neutrophils was a particularly
novel finding that led to the concept that
the innate immune response is involved
in this form of thrombosis, and the term
“immunothrombosis” was coined. Neutrophils
are proposed to enhance immunothrombosis
via several mechanisms, including the
generation of neutrophil extracellular traps.
A mouse model of spontaneous venous
thrombosis was recently described that
involves reducing levels of antithrombin and
protein C with small interfering RNAs.7 Mice
develop thrombi in veins of the head, leg, and
liver with fewer thrombi in vessels in the heart,
lung, and brain and no thrombi in the kidney
or gastrointestinal tract. This pattern of
thrombosis is consistent with the notion of
differential expression of anticoagulants that
regulate tissue-specific hemostasis. The
prothrombotic phenotype in the mice develops
within 3 days and is fatal. Anticoagulating the
BLOOD, 26 MAY 2016 x VOLUME 127, NUMBER 21
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mice with the thrombin inhibitor dabigatran
etexilate prevents the coagulopathy.7 A
selective reduction of antithrombin in mice
produced a milder prothrombotic phenotype,
whereas reducing protein C alone did not
induce thrombosis.7 One unanswered question
with the model and in individuals with
deficiencies in anticoagulants is, What is
triggering the spontaneous thrombosis?
Heestermans et al examined the roles of
platelets, neutrophils, and FXII in this new
spontaneous venous thrombosis model (see
figure). Depletion of platelets prevented the
thrombotic coagulopathy and fibrin deposition
in the liver, which is consistent with a role
of platelets in the IVC stenosis model.5,6
However, depletion of neutrophils did not
reduce thrombosis in mice, in contrast to the
results with the IVC stenosis model.6 This
indicates that neutrophils do not contribute to
all forms of venous thrombosis. FXII has been
shown to contribute to thrombosis in a variety
of mouse models, including the IVC stenosis
model.6,8 Surprisingly, Heestermans et al
found that decreasing levels of FXII increased
thrombosis. How do we explain this result?
Heestermans et al explored the possibility that
a deficiency of FXII enhances secondary
bleeding changes in fibrinolysis and edema
without any definitive conclusions. It is
possible that reducing levels of FXII leads to
BLOOD, 26 MAY 2016 x VOLUME 127, NUMBER 21
destabilization of clots with increased
embolization.
In summary, all mouse models have
limitations, but it is always important to
remember that we are modeling human disease.
Venous thrombosis is a complex disease that
may be triggered by different risk factors and
combinations of risk factors. Further studies
are needed to determine the role of neutrophils
and other immune cells in different forms
of venous thrombosis in mouse models and
humans. FXII is an attractive target for
anticoagulant drug therapy8 and therefore
we need to understand why it was apparently
antithrombotic in the mouse model of
spontaneous venous thrombosis. It would
be very interesting to determine the effect
of reducing FXI levels in this spontaneous
thrombosis model, given the close relationship
between FXII and FXI and results from
a recent study showing reduced thrombosis
without hemorrhage in patients undergoing
total knee arthroplasty.9 Finally, platelets have
been shown to play roles in different mouse
models of thrombosis, and clinical studies have
shown that aspirin reduces the recurrence of
venous thrombosis in patients.10 These results
suggest that antiplatelet therapy may reduce
the incidence of venous thrombosis in patients.
Conflict-of-interest disclosure: The author declares
no competing financial interests. n
REFERENCES
1. Heestermans M, Salloum-Asfar S, Salvatori D, et al.
Role of platelets, neutrophils, and factor XII in
spontaneous venous thrombosis in mice. Blood. 2016;
127(21):2630-2637.
2. Dahlbäck B. Advances in understanding pathogenic
mechanisms of thrombophilic disorders. Blood. 2008;
112(1):19-27.
3. Mackman N. New insights into the mechanisms of
venous thrombosis. J Clin Invest. 2012;122(7):2331-2336.
4. Diaz JA, Obi AT, Myers DD Jr, et al. Critical review of
mouse models of venous thrombosis. Arterioscler Thromb
Vasc Biol. 2012;32(3):556-562.
5. Brill A, Fuchs TA, Chauhan AK, et al. von Willebrand
factor-mediated platelet adhesion is critical for deep vein
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6. von Brühl ML, Stark K, Steinhart A, et al. Monocytes,
neutrophils, and platelets cooperate to initiate and
propagate venous thrombosis in mice in vivo. J Exp
Med. 2012;209(4):819-835.
7. Safdar H, Cheung KL, Salvatori D, et al. Acute and
severe coagulopathy in adult mice following silencing of
hepatic antithrombin and protein C production. Blood.
2013;121(21):4413-4416.
8. Kenne E, Nickel KF, Long AT, et al. Factor XII:
a novel target for safe prevention of thrombosis and
inflammation. J Intern Med. 2015;278(6):571-585.
9. Büller HR, Bethune C, Bhanot S, et al; FXI-ASO
TKA Investigators. Factor XI antisense oligonucleotide for
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10. Warkentin TE. Aspirin for dual prevention of venous
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DOI 10.1182/blood-2016-03-704270
© 2016 by The American Society of Hematology
2511
From www.bloodjournal.org by guest on June 17, 2017. For personal use only.
2016 127: 2510-2511
doi:10.1182/blood-2016-03-704270
Mouse models of venous thrombosis are not equal
Nigel Mackman
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