REVIEW
Received 6 Jan 2016 | Accepted 14 Dec 2016 | Published 9 Feb 2017
The molecular basis of endothelial cell plasticity
Elisabetta Dejana
1,2,
*
,w
, Karen K. Hirschi
3,
* & Michael Simons
4,
*
The endothelium is capable of remarkable plasticity. In the embryo, primitive endothelial cells
differentiate to acquire arterial, venous or lymphatic fates. Certain endothelial cells also
undergo hematopoietic transition giving rise to multi-lineage hematopoietic stem and
progenitors while others acquire mesenchymal properties necessary for heart development.
In the adult, maintenance of differentiated endothelial state is an active process requiring
constant signalling input. The failure to do so leads to the development of endothelial-to-
mesenchymal transition that plays an important role in pathogenesis of a number of diseases.
A better understanding of these phenotypic changes may lead to development of new
therapeutic interventions.
E
ndothelial cells, that form the early vascular plexus during embryo development, progress
through a number of fate transitions until they achieve their highly differentiated adult
state. At early stages of embryonic development, cells that will form the primitive
vasculature exhibit a primordial, non-specialized endothelial phenotype. As the plexus is
remodelled into specialized vascular structures, these primitive endothelial cells acquire
specialized characteristics typical of arteries, veins or lymphatics. Subsequently, during organ
vascularization, the endothelial cells differentiate further to adapt to the specific needs of the
organ
1
.
In this review, we discuss key emerging concepts and challenges in the rapidly moving field of
endothelial fate transition, including signalling pathways implicated in endothelial-to-
hematopoietic cell transition (EHT) and endothelial-to-mesenchymal transition (EndMT), as
well as physiological and pathological implications of these processes.
Endothelial cell development and fate transitions during embryogenesis
The vasculature is among the first organ systems to develop during embryogenesis, and is
essential for the growth, survival and function of all other organ systems. Blood vessels are
composed of endothelial cells that form the inner, luminal layer and smooth muscle cells that
form the surrounding vessel wall. During blood vessel development, endothelial cells are formed
first, and undergo rapid expansion and coalescence into capillary plexi that are then remodeled
into a circulatory network. Vascular remodelling and maturation involves coordinated
migration, growth control and specification of arterial and venous endothelial subtypes, as
well as smooth muscle cell recruitment.
DOI: 10.1038/ncomms14361
OPEN
1
Vascular Biology Unit, FIRC Institute of Molecular Oncology, Milan 20129, Italy.
2
Department of Immunology, Genetics and Pathology, Uppsala University,
Uppsala 751 85, Sweden.
3
Yale Cardiovasc. Res. Center, Departments of Internal Medicine, Genetics and Biomedical Engineering New Haven, Connecticut
CT06511, USA.
4
Yale Cardiovascular Research Center, Department of Internal Medicine and Department of Cell Biology, Yale University School of Medicine,
New Haven, Connecticut CT06511, USA. * These authors contributed equally to this work. w On leave of absence from Department of Oncology and Onco-
Haematology, University of Milan, Milan, Italy. Correspondence and requests for materials should be addressed to E.D. (email: elisabetta.dejana@ifom.eu) or
to K.K.H. (email: karen.hirschi@yale.edu) or to M.S. (email: michael.simons@yale.edu).
NATURE COMMUNICATIONS | 8:14361 | DOI: 10.1038/ncomms14361 | www.nature.com/naturecommunications 1
As the vasculature is established within distinct organs, the
endothelium therein is further phenotypically specialized to meet
the needs of the tissue. For example, in the brain and retina, tight
junctions are formed to create a barrier against infiltration of
circulating factors and cells. In contrast, in tissues with filtration
functions, such as the kidney and liver, the endothelium can be
discontinuous and develop fenestrae to promote infiltration and
extravasation of circulating factors.
Vascular endothelium also significantly contributes to the
development of other organ systems, including blood and the
heart. In these circumstances, endothelial cells undergo a fate
transition into another cell type; that is, hematopoietic cells, or
cardiac mesenchyme, respectively. The differentiation, specializa-
tion and fate transitions of endothelium during development are
discussed herein.
Endothelial cell differentiation. The emergence of primordial
(non-specialized) endothelial cells is referred to as vasculogenesis
and begins in the developing mammal shortly after gastrulation
in the extraembryonic yolk sac. Endothelial cells are formed from
mesodermal progenitors in response to signals from the adjacent
visceral endoderm and coalescence into vascular plexi that
are remodeled into circulatory networks during the process of
angiogenesis.
Genetic manipulation studies in the mouse revealed that
fibroblast growth factor 2 (FGF2 or bFGF) and bone morpho-
genetic protein 4 (BMP4) are not only critical for mesoderm
formation, but also play an important role in endothelial
cell differentiation.
2
Indian hedgehog (IHH) signalling,
likely mediated via BMP4 (ref. 3) also promotes endothelial cell
development, and is sufficient to induce the formation of
endothelial cells in mouse embryo explants that lack
endoderm
2
. Vascular endothelial growth factor (VEGF-A) is
another key regulator of vasculogenesis. It predominantly binds
two receptors, VEGFR1 (Flt-1), which acts as a sink for bioactive
VEGF-A, and VEGFR2 (Flk-1 or Kdr), which is required for
vascular plexus development
4
. VEGFR2
/
mouse embryonic
stem cells generate endothelial cells, although they fail to
propagate in vitro. Thus, VEGF-A may regulate the survival
and/or propagation of endothelial cells, but not their fate
specification.
Transcriptional regulators in the ETS family are known to play
an important role in endothelial cell development, and the
regulatory regions for almost all endothelial genes contain ETS
binding sites
5,6
. ETS variant 2 (Etv2 or ER71/etsrp), in particular,
regulates the differentiation of mesodermal progenitors toward an
endothelial cell fate. It is restricted to VEGFR2-expressing
mesodermal progenitors, and mice deficient in Etv2 lack a yolk
sac vascular plexus, dorsal aortae and endocardium, despite
normal mesoderm formation
7
. Overexpression of Etv2 in vivo
leads to ectopic expression of endothelial-specific genes,
suggesting it is necessary and sufficient for endothelial cell
development
7
. FGF signalling is known to promote Ets-driven
gene expression
8
, although we have much to learn about the
coordination among signalling pathways and transcriptional
regulators that mediate endothelial cell differentiation.
Endothelial cell specialization. Once formed, primordial vascu-
lature undergoes further differentiation and specialization,
resulting in formation of distinct arterial, venous and lymphatic
systems. Signalling pathways implicated in early endothelial cell
development are also thought to play significant roles in arterial-
venous specification. For example, during arterial-venous speci-
fication, VEGF-A binds to VEGFR2 and co-receptor neuropilin-1
(Nrp1), leading to activation of Notch signalling. Arterial-specific
genes, including EphrinB2, are upregulated downstream of
Notch signalling; whereas, venous-specific EphB4 expression is
suppressed
9
. Inhibition of Notch signalling results in an arterial-
to-venous fate switch
10
. Wnt signalling is also involved in the
specification of arterial endothelial cells; b-catenin, a trans-
criptional co-activator of Wnt signalling pathway, upregulates
Notch ligand Dll4 and promotes arterial specification
11
.In
addition, Hedgehog acts upstream of VEGF-A via smoothened
receptor to drive arterial endothelial cell specification and repress
venous fate
12,13
. Venous endothelial cell specification is induced
by chicken ovalbumin upstream promoter-transcription factor II
(COUP-TFII). Endothelial-specific deletion of COUP-TFII leads
to arterialization of veins, whereas ectopic expression results in
fusion of veins and arteries
14
. Lymphatic endothelial cells are
formed, in part, from a subset of endothelium within the cardinal
vein; wherein, the co-expression of COUP-TF II and SOX18
leads to upregulation of PROX-1 that promotes lymphatic
specification. The propagation of lymphatic endothelial cells is
mediated by VEGFR3 signalling, driven by VEGF-C from the
surrounding mesenchyme. Importantly, further stabilization and
quiescence of the lymphatic system is mediated by FOXC2
and fluid shear stress, providing an essential link between
biomechanical forces and endothelial cell identity
15
.
Transition of endothelial cells to hematopoietic cells. As arterial
and venous endothelial cells are being specified within the yolk
sac, another type of endothelial cell is also developing. These are
hemogenic endothelial cells that will give rise to multi-lineage
hematopoietic progenitors through a process referred to as EHT.
Hemogenic endothelial cells also form in other tissues throughout
the course of gestation, including the placenta and umbilical
vessels, as well as within the embryonic aorta-gonad-mesene-
phros (AGM) region where they give rise to hematopoietic stem
cells (HSC)
16
. Other tissues observed to give rise to hematopoietic
cells during development include the head vasculature
17
and the
endocardium
18
.
The process of blood production from the endothelium
appears to require two distinct steps: hemogenic endothelial cell
specification and EHT (depicted in Fig. 1). The initial specifica-
tion of hemogenic endothelial cells involves some of the same
signalling pathways implicated in arterial-venous specification,
Vascular
endothelium
Hemogenic
endothelium
RA
cKit
Notch
Cell cycle control
Runx1
Gata/Ets/Scl/Lmo2
Wnt/β-catenin
Sox17
Notch
HSPC
EHT in blood development
Figure 1 | Schematic representation of endothelial-to-hematopoietic
(EHT) transition and endothelial-to-mesenchymal (EndMT) transition
during development. During definitive hematopoiesis, a subset of
endothelial cells is specified to become hemogenic (dark red), and these
cells give rise to hematopoietic stem and progenitor cells (HSPC) via EHT.
The specification of hemogenic endothelial cells requires retinoic acid (RA)
signalling, which leads to upregulation of c-Kit. Notch is activated
downstream of c-Kit expression and controls endothelial cell cycle to
enable hemogenic specification via mechanisms that are still unclear. The
subsequent generation of HSPC requires transcription factor Runx1, and
binding partners Gata, Ets, Scl and Lmo-2. Other factors involved in this
process include Wnt/b-catenin, Sox17 and Notch, whose molecular roles
and interactions are under study.
REVIEW NATURE COMMUNICATIONS | DOI: 10.1038/ncomms14361
2 NATURE COMMUNICATIONS | 8:14361 | DOI: 10.1038/ncomms14361 | www.nature.com/naturecommunications
although it appears to be uniquely initiated by retinoic acid (RA)
signalling. During vasculogenesis in the murine yolk sac, the
first site of hemogenic endothelial cell specification, active
RA is generated by retinaldehyde dehydrogenase 2 (Raldh2).
Raldh2
/
mutant embryos exhibit endothelial cell hyperpro-
liferation, lack vascular remodeling and hemogenic endothelial
cell development and die mid-gestation
19
. RA signalling is also
critical for the development of hemogenic endothelial cells within
the AGM
20
. In both tissues,B90% of endothelial cells with active
RA signalling exhibit a hemogenic phenotype, and B90%
of hemogenic endothelial cells are undergoing active RA
signalling
2,20
.
Signalling pathways that function downstream of RA in this
process include c-Kit and Notch. c-Kit (CD117) is a receptor with
tyrosine kinase activity that binds stem cell factor; its expression
is a distinguishing feature of hemogenic endothelial cells, relative
to non-blood forming endothelial cells
19
. c-Kit is known to be
transcriptionally regulated by RA signalling, and re-expression of
c-Kit in Raldh2
/
mutants rescues hemogenic endothelial cell
development and definitive hematopoiesis
21
.
Notch1 expression in endothelial cells is upregulated down-
stream of c-Kit
21
, and is essential for hemogenic endothelial
specification, just as it is for arterial
22
and lymphatic specifi-
cation
23
. Notch signalling is involved in cell-fate decisions and
cell differentiation, and embryos lacking Notch1, or Notch1 and
Notch4, the only Notch receptors expressed by endothelial cells,
exhibit abnormal vascular remodeling similar to Raldh2
/
mutants
24
. Notch1, specifically, is expressed in the ventral wall of
the dorsal aorta where hemogenic endothelium forms
25
and the
AGM of Notch1
/
mutants exhibit decreased hematopoietic
activity. Collectively, these data suggest that Notch signalling is
important in hemogenic endothelial cell development; however,
much needs to be learned about the role of Notch signalling in
EHT. Some studies suggest Notch ligand distribution dictates
the outcome of Notch signalling during definitive hematopoiesis
in the AGM. That is, hemogenic specification is driven by
low levels of Jag1-mediated Notch signalling. In the absence of
Jag1, Dll4-mediated high Notch activity instead drives arterial
specification
26
.
The formation of hematopoietic stem and progenitor cells
(HSPC) from the endothelium has been most intensely studied in
the AGM region. Therein, intra-aortic hematopoietic clusters
are formed from hemogenic endothelial cells during EHT. The
clusters are composed of pre-HSC that differentially express
endothelial and hematopoietic markers including VE-cadherin
c-Kit, Ly6a (Sca-1), CD41 and CD45, suggesting that a post-
hemogenic endothelial cell intermediate may exist along the
transition from endothelial cell to HSPC within these struc-
tures
27–29
. The dorsoventral polarity of the clusters emergence
is thought to be guided by mesenchymally derived pro-
hematopoietic ventralizing (VEGF, FGF2, TGFb, BMP4) or
anti-hematopoietic dorsalizing (EGF and TGFa) factors, which
affect the expression of critical hematopoietic transcription
factors involved in EHT (ref. 30).
One such essential transcriptional regulator of EHT is Runx1
(AML1), a member of a family of transcription factors called core
binding factors
31
. Runx1 is thought to repress the endothelial
program, while activating the hematopoietic program during the
EHT process
27,29
. Deletion of Runx1 does not prevent hemogenic
endothelial cell specification, but prevents the transition of these
cells to CD41 þ CD45 þ hematopoietic cells
32
. The transition
from endothelial to hematopoietic phenotype is in part controlled
by binding of multiprotein complexes containing GATA,
Ets
33
, and SCL factors to Runx1 enhancers that promotes HSC
emergence
34
, as well as the loss of expression of genes associated
with arterial identity
35
, at least within the AGM.
There is further evidence that Notch and Wnt pathways
interact to generate HSC in the zebrafish embryo and also are
involved in driving hematopoietic development from embryonic
stem cells
36,37
. Wnt/b-catenin activity is transiently required in
the AGM for emergence and generation of long-term HSC, as
well as production of hematopoietic cells in vitro from AGM
endothelial precursors
37
. Downstream of Wnt signalling in the
AGM, is expression of transcription factor Sox17. Conditional
loss of Sox17 in endothelial cells in the AGM leads to increased
production of hematopoietic cells
35
, suggesting that Sox17
modulates the fate of hemogenic endothelium by actively
repressing the hematopoietic program. Other studies show that
enforced expression of Sox17 in mouse embryonic stem cells
leads to increased blood-forming endothelial cells that generate T
lymphocytes, through a mechanism involving Notch signalling
38
.
Thus, more work is needed to understand the roles and
interactions among these regulators of EHT.
Other factors, including inflammatory
39–42
and G-protein
coupled receptor signalling
43,44
, purine signalling
45
and chro-
matin remodeling
46,47
are also involved in promoting EHT, and
their coordination with known regulators of this process are
under investigation.
Endothelial-to-mesenchymal transition in development .In
addition to vascular endothelial cells within hematopoietic tissues
undergoing EHT and giving rise to hematopoietic stem and
progenitor cells, the specialized endothelial cells that line the
heart (endocardial cells) also undergo EndMT, and give rise to
mesenchymal cells necessary for proper heart development.
Although all cells in the heart arise from one or more epithelial-
to-mesenchymal transition, EndMT, specifically, generates valve
progenitor cells that give rise to the mitral and tricuspid valves.
EndMT also contributes to endocardial cushion formation, as
well as to generation of cardiac fibroblasts and smooth muscle
cells, but not cardiac myocytes.
These developmental mechanisms may well be recapitulated in
adult valve disease, in cardiac fibrosis, and in myocardial
responses to ischemic injury (see below). Thus, understanding
what regulates EndMT during embryogenesis may provide
insights needed to treat postnatal pathologies. In addition, since
some of the signalling pathways that regulate developmental
EndMT (TGFb, BMP, Notch, Wnt/b-catenin)
48
also play a role in
EHT (Fig. 1), as well as postnatal EndMT (discussed in detail in
subsequent sections), comparing these regulatory pathways may
reveal common targets for therapy.
Maintenance of adult endothelial homeostasis
The enormous phenotypic plasticity exhibited by endothelial cells
is a direct reflection of their exposure to the environment full of
growth factors, cytokines, rich in oxygen and mechanical stresses.
It is not surprising, therefore, that maintenance of endothelial
normalcy is an active processes requiring constant energy
expenditure and signalling input. While still poorly understood,
recent studies shed light on several active endothelial ‘main-
tenance’ pathways. These include regulation of expression of key
proteins such as VEGFR2 and FGFR1, maintenance of endothe-
lial barrier function, suppression of apoptosis and prevention of
the fate drift.
Fibroblast growth factors (FGFs) play a particularly important
role in control of endothelial homeostasis. Even a transient
withdrawal of the FGF signalling input leads to a progressive loss
of endothelial cell-cell contacts, increased permeability and,
eventually, compromise of vascular integrity
8,49,50
while a more
prolonged withdrawal leads to endothelial apoptosis
51
and
vascular rarefication
52
including the loss of vasa vasorum
53
.In
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NATURE COMMUNICATIONS | 8:14361 | DOI: 10.1038/ncomms14361 | www.nature.com/naturecommunications 3
part, these changes are due to increased VE-cadherin phospho-
rylation at Y658 (and thus the loss of VE-cadherin-p120-catenin
association that impairs adherence junctions), and in part to a
decline in VEGFR2 levels (thereby increasing endothelial
apoptosis). The former is a consequence of a decrease in FGF-
dependent expression of the phosphatase Shp2 (ref. 54) while the
latter is due to the requirement for the FGF-driven Ets/FOXC2
VEGFR2 transcription
8
. Since both VE-cadherin phospho-
rylation and VEGFR2 turnover are rapidly occurring processes,
withdrawal of a continuous FGF signalling input rapidly leads to
the above-described consequences
55
.
Another key role played by the FGF signalling input is
maintenance of the endothelial cell fate. FGFs achieve this by
blocking activation of TGFb signalling cascade that is central to
the induction of EndMT transition both in blood
56,57
and
lymphatic endothelial cells
58
that is discussed below. Endothelial
homeostasis is also maintained by VE-cadherin that is the cell
specific major organizer of endothelial cell-to-cell adherens
junctions
59,60
. The expression and clustering of VE-cadherin at
cell–cell junctions not only maintains endothelial cell-to-cell
adhesion but, through the interaction with a complex network
of intracellular partners, transduces intracellular signals that
mediate contact inhibition of cell growth, cell polarity and lumen
formation. Therefore, conditions that disrupt endothelial
junctions not only induce increase in vascular permeability by
opening intercellular gaps but also change the endothelial cell
responses to their environment and to the surrounding cells. For
instance, when VE-cadherin junction organization is dismantled,
gene transcription of endothelial cells is strongly modified, the
cells tend to grow in multiple layers, are unable to form a correct
vascular lumen and establish adhesion contacts with the
surrounding pericytes and smooth muscle cells
61–63
.
At the molecular level, VE-cadherin is linked through its
cytoplasmic domain to p120 catenin, b-catenin and plakoglobin.
Furthermore, it can interact and modulate signalling of several
growth factor receptors to promote contact inhibition of cell
growth. For instance, VE-cadherin expression and clustering
inhibits VEGFR2 and FGFR1 signalling while increases TGFbR
complex organization and signalling
64–66
. Some phosphatases
(VE-PTP, DEP-1, PTPu, Csk and SHP2)
61,67,68
and
kinases (such as Src or FAK)
69–71
may also associate with the
VE-cadherin complex and modulate cell signalling. This complex
signalling system is dynamic and continuously adapting to
different external conditions (shear stress, growth stimulation,
increase in permeability) to maintain endothelial integrity
72
.
Definition and occurrence of EndMT in various pathologies
In the absence of active input endothelial cells may either die or
undergo EndMT, a process with certain similarities with the
better understood epithelial-to-mesenchymal transition. Just as
occurs during normal development, during postnatal EndMT,
endothelial cells acquire mesenchymal characteristics such as an
elongated, fibroblastoid morphology, increased motility, cytoske-
letal modifications and cell-to-cell junction rearrangement.
However, in the adult, they further become proliferative,
thrombogenic and deposit large amounts of extracellular matrix.
EndMT may lead to endothelial cells acquiring a variety
of different mesenchymal fates (Fig. 2). As the result of this
transformation, endothelial cells undergo a profound phenotypic
change assuming the shape and properties of mesenchymal cells
(fibroblasts, smooth muscle cells), including secretion of extra-
cellular matrix proteins such as fibronectin and collagen, and
expression of various leukocyte adhesion molecules. (see Fig. 3
reporting the list of specific markers)
At the cellular level, EndMT consequences include altered
endothelial cell junction organization, loss of cell polarity, and
increased cell proliferation and migratory capacity
73
. This results
in a number of pathological consequences of considerable
clinical significance in diseases ranging from cavernous cerebral
malformations (CCM)
74
to tissue fibrosis
75–77
, heterotopic
ossification
78,79
, neointima formation
56,80–82
, atherosclerosis
81
and cancer
83,84
.
The recognition of EndMT in tissues relies on detection of cells
expressing both mesenchymal and endothelial markers. However,
this approach assumes that cells that have undergone EndMT
retain endothelial marker expression. While this is likely to be
true immediately after the EndMT switch, at later stages
Endothelium lining
CCM Malformations
Smooth muscle cells (SMCs)
Fibroblasts
Osteocytes
Adipocytes Chondrocytes
EndMT
Reversion?
Maladaptation
Inflammatory
cytokines
β-catenin
signalling
miRNAs
FGFVEGF
Figure 2 | Schematic representation of endothelial-to-mesenchymal transition in the adult. In the adult, endothelial cells (flesh coloured) deprived of
growth factors or exposed to inflammatory cytokines may undergo EndMT (see light and intense green cells representing the progression to EndMT) and
acquire characteristics of fibroblasts, smooth muscle cells, osteocytes, adipocytes, chondrocytes or form vascular malformations such as CCM.
Re-establishment of endothelial homeostasis by exposure to growth factors or to specific miRNAs may revert the mesenchymal phenotype (from intense
green to flesh coloured cells).
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endothelial cells may lose their endothelial markers (Fig. 3).
Therefore, any immunocytochemical assessment of EndMT is an
underestimate of the true frequency of this phenomenon. In
addition, EndMT frequency varies depending on location and
type of endothelial cells undergoing this process. Thus, in
transplant settings, over 80% of luminal endothelial cells express
mesenchymal markers while the frequency of this phenomenon
elsewhere (for example, in the neointima) is much less
56,80
.
Similarly, endothelial cells overlying the atherosclerotic plaque
have a much higher EndMT incidence than nearby cells not in
direct contact with the plaque
81
.
Fate-mapping has been employed to assess the true frequency
of this phenomenon. For obvious reasons, however, this is
possible in mice but not in clinical specimens. Fate-mapping
involves activation of endothelial expression of a marker gene
(typically LacZ) in adult mice using a tissue-specific Cre. Cdh5-
CreER
T2
is the most common driver line used for these studies
85
.
It reaches high, but not 100% efficiency in most, but not all,
endothelial beds so even this approach does not provide full
assessment of EndMT frequency.
EndMT in atherosclerosis and transplant arteriopathy. Recent
studies documented EndMT contribution to vascular pathology
both in transplant arteriosclerosis and atherosclerosis lesions.
Both diseases are characterized by the growth of neointima that is
composed of a mixture of smooth muscle cells, fibroblasts,
mononuclear inflammatory and immune cells and extracellular
matrix. In both cases chronic vascular inflammation, induced by
certain immune mismatches in the case of transplant arterio-
pathy
86
and lipid deposition and mechanical forces in the case of
atherosclerosis
87,88
, are thought to play a central role in disease
progression. EndMT has been detected in both conditions and
likely plays a critical role in progression of both disease states.
Fate mapping using Cdh5Cre;mTmG fate mice in a
mouse acute transplant rejection model showed that B10% of
neointimal smooth muscle cells were of endothelial origin 2
weeks after transplant. At the same time, B80% of neointimal
and B60% of luminal endothelial cells expressed mesenchymal
markers indicating EndMT
56,80
. In agreement with these data,
examination of patient samples from explanted rejected hearts
found that B80% of coronary artery luminal endothelial and
neointima cells were undergoing EndMT
56,80
.
Studies in other vascular injury models revealed a lower
incidence of EndMT: 5% of neointimal SMC were of endothelial
origin in the mouse wire injury model and 7% in vein-to-artery
(inferior vena cava to aorta) graft model. Interestingly, in the
latter case the incidence of EndMT increased from 3% at 2 weeks
after grafting to 7% 4 weeks later, indicating that it is an ongoing
process
56,80
. A study utilizing a somewhat different vein graft
model (jugular vein to femoral artery) reported still higher
incidence of EndMT: 28 to 50% at 5 weeks
82
. Similarly high
incidence of EndMT has been observed in a mouse model using
tissue-engineered vascular grafts. Here, EndMT frequency varied
from 38 to 51% in occluded grafts (severe rejection) to 17% in
patent grafts (less severe rejection)
89
. These variations in the
observed EndMT extent likely relate to the severity of injury, the
magnitude of the inflammatory response and differences in
hemodynamic stress.
EndMT is equally prevalent in atherosclerosis, a progressive
disease initiated by lipid deposition in parts of the arterial tree
subjected to disturbed blood flow and is characterized by the
gradual build-up of intraluminal plaques leading to reduction in
distal tissue perfusion
88
. Some of the plaques are prone to
rupture, an event than can lead to thrombosis and sudden
death
90
. Chronic inflammation is thought to play a particularly
important role in the disease progression although molecular
details remain poorly understood
86,88,91
.
Examination of fate-mapped Apoe
/
mice revealed that after
four months of high fat diet B30% of luminal aortic endothelial
cells were undergoing EndMT while no EndMT was detected in
animals on the normal diet
81
. Similarly, examination of
atherosclerotic plaques showed that about 35% of fibroblasts
and ‘‘mesenchymal’’ cells were of endothelial origin
92
. Exami-
nation of human atherosclerotic vessels using immuno-
cytochemical techniques, confirmed frequent occurrence of
EndMT
Early EndMT markers
Downregulated
Upregulated
αSMA, SM22a,
Fsp1/S100a, CD44
Downregulated
Endothelial markers
(moderate or strong
inhibition)
Upregulated
SM-SMHL, SM-calponin,
Smoothelin, Fibronectin,
Tenascin, Collagen III,
THY1, Vimentin, Notch3,
PAI, SCA1, ZEB2,
MMP2 and 9, VCAM,
ICAM1
Late EndMT markers
TGF-β, BMPs,
β-catenin
FGF
Endothelial markers
(partial inhibition),
Dismantled
adherens junctions
Figure 3 | EndMT markers. Endothelial cells may undergo EndMT either from growth factor deprivation (FGF) or from activation of b-catenin, TGFb,BMP
pathways. EndMT progresses through different steps. The early endothelial response is characterized by a partial downregulation of endothelial markers,
junction dismantling and up-regulation of some early mesenchymal markers. At later times, expression of endothelial markers further declines while more
mesenchymal markers including matrix proteins, metallo-proteases or cytoskeletal proteins are up-regulated.
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