lin superfamily that is expressed on platelets, most leukocyte
subsets, and at endothelial cell (EC) junctions.1–5 PECAM-1
sustains both homophilic interaction (PECAM-1–PECAM-1)
and heterophilic interactions with integrin αvβ3, CD38, and
CD177 and is a highly efficient signaling molecule mediat-
ing both outside-in and inside-out signaling.5,6 Because of its
identification within the plasma membrane of EC,7 and its clon-
ing soon thereafter,8 and subsequent generation of PECAM-1
knockout mouse,9 the role of PECAM-1 in the vascular system,
as well as during inflammatory and autoimmune responses, has
been extensively studied.1–5,9,10 For example, antibody blockade
of PECAM-1 function in in vitro adhesion assays limits leuko-
cyte migration through EC monolayers, as well as the direc-
tionality of migrating neutrophils under flow conditions.2,11–14
PECAM-1 has also been shown to mediate leukocyte motility
and extravasation through the basement membrane in vivo, in
response to interleukin-1β but not tumor necrosis factor-α.5,15–17
A mechanosensory role for PECAM-1 was first demonstrated
in EC that had been mechanically stimulated or exposed to
latelet-endothelial cell adhesion molecule (PECAM)-1 is a
type-1 transmembrane glycoprotein of the immunoglobu-
osmotic stress.18,19 Antibody-conjugated magnetic bead ligation
of PECAM-1 followed by exposure to a mechanical force
resulted in tyrosine phosphorylation of the cytoplasmic tail
within 30 seconds.18,19 Downstream kinases such as extracellular
signal-regulated kinases-1/2 were subsequently activated.19
EC-expressed PECAM-1 can also form a mechanosensory
complex with vascular endothelial growth factor receptor-2
and vascular endothelial-cadherin, which responds to fluid
shear stress.20 Disturbed flow profiles with relatively low and
oscillatory shear stress are found in the inner curvature of the
aortic arch or at bifurcations in the arterial system, and are
associated with localized formation of atheroma. These areas
also show activation of the proinflammatory transcription
factor nuclear factor kappa-light-chain-enhancer of activated
B cells in EC, compared with regions experiencing laminar
flow with high levels of shear stress. Importantly, in PECAM-
1–deficient mice, EC positioned in the inner curvature of the
aortic arch were protected against nuclear factor kappa- light-
chain-enhancer of activated B cells activation.20 However,
cultured EC, which had been subjected to PECAM-1 small
interfering RNA knock down, did not align in response to
Received on: September 5, 2012; final version accepted on: January 17, 2013.
From the College of Medical and Dental Science, The Medical School, University of Birmingham, UK (M.H., E.S., E.R., C.D.B., G.B.N., G.E.R.);
Bioengineering, Imperial College, Campus South Kensington, London (R.K.); and Cardiology, Erasmus MC, Rotterdam, UK (D.S.).
*These authors contributed equally to the experimental and written content of this manuscript.
The online-only Data Supplement is available with this article at http://atvb.ahajournals.org/lookup/suppl/doi:10.1161/ATVBAHA.112.300379/-/DC1.
Correspondence to M. J. Harrison, Centre for Cardiovascular Sciences, The Medical School, University of Birmingham, Birmingham, UK, B15 2TT.
© 2013 American Heart Association, Inc.
Arterioscler Thromb Vasc Biol is available at http://atvb.ahajournals.org DOI: 10.1161/ATVBAHA.112.300379
Objective—Polymorphisms in the platelet-endothelial cell adhesion molecule (PECAM-1)-1 gene are linked to increased
risk of coronary artery disease. Because PECAM-1 has been demonstrated to form a mechanosensory complex that can
modulate inflammatory responses in murine arterial endothelial cells, we hypothesized that PECAM-1 contributes to
atherogenesis in a shear-dependent and site-specific manner.
Approach and Results—ApoE–/– mice that were wild-type, heterozygous, or deficient in PECAM-1 were placed on a high-
fat diet. Detailed analysis of the aorta at sites with differing hemodynamics revealed that PECAM-1–deficient mice had
reduced disease in areas of disturbed flow, whereas plaque burden was increased in areas of steady, laminar flow. In
concordance with these observations, bone marrow chimera experiments revealed that hematopoietic PECAM-1 resulted
in accelerated atheroma formation in areas of laminar and disturbed flow, however endothelial PECAM-1 moderated
disease progression in areas of high sheer stress. Moreover, using shear stress–modifying carotid cuffs, PECAM-1 was
shown to promote macrophage recruitment into lesions developing in areas of low shear stress.
Conclusions—PECAM-1 on bone marrow cells is proatherogenic irrespective of the hemodynamic environment, however
endothelial cell PECAM-1 is antiatherogenic in high shear environments. Thus, targeting this pathway therapeutically
would require a cell-type and context-specific strategy. (Arterioscler Thromb Vasc Biol. 2013;33:694-701.)
Key Words: ApoE–/– ◼ atherosclerosis ◼ carotid cuffs ◼ low shear ◼ platelet endothelial cell adhesion molecule-1
The Role of Platelet-Endothelial Cell Adhesion Molecule-1 in
Atheroma Formation Varies Depending on the Site-Specific
Matthew Harrison,* Emily Smith,* Ewan Ross, Robert Krams, Dolf Segers,
Christopher D. Buckley, Gerard B. Nash, G. Ed Rainger
Harrison et al Site-Specific Effects of PECAM-1 in Atheroma 695
laminar high shear, but remained polygonal,20 a phenotype
found in areas of arteries prone to atherogenesis. Generally,
laminar shear stress suppresses the inflammatory responses of
EC,21,22 and we have recently shown that this protective effect
also requires expression of PECAM-1.23 Thus, taken together,
these results indicate that PECAM-1 may play roles in the
proinflammatory response believed to promote atherogenesis
in regions of disturbed flow and in the protective effects of
laminar, high shear stress.
The expression of PECAM-1 on EC, platelets, and leu-
kocytes has led investigators to hypothesize that it may play
a key role in cardiovascular disease. In humans, population
studies have linked single nucleotide polymorphisms of the
PECAM-1 gene and elevated levels of soluble PECAM-1 to
severe coronary artery stenosis as well as myocardial infarc-
tion.24–27 In mice, the role of PECAM-1 in atherogenesis
remains controversial. Recent studies in the Pecam-1–/– ApoE–/–
mouse28,29 or Pecam-1–/–ldlr–/– mouse30 on high-fat diet (HFD)
have produced conflicting results. Although all the studies
agree that PECAM-1 is proatherogenic in the inner curvature
of the aortic arch, the site-specific effects of PECAM-1 in the
descending aorta are inconsistent between models, with a net
proinflammatory28 or antiinflammatory30 role of PECAM-1 in
lesion formation being observed over the whole aorta.
To investigate the site-specific effects of PECAM-1, we
generated Pecam-1–/– ApoE–/– mice and placed them on a HFD
to promote atherogenesis. The disease phenotype was domi-
nated by the descending aorta, which delivered a net eleva-
tion in plaque burden in the whole aorta. However, a detailed
analysis of disease burden at sites subject to different hemo-
dynamic forces showed a reduction of plaque formation in
the inner curvature of the aortic arch where low shear stress
and complex patterns of flow are apparent. There was also
a concomitant elevation in plaque burden in the descending
aorta, which is subject to laminar high shear flow. Formation
of radiation chimeras revealed that increased plaque burden
in the descending aorta was primarily a result of loss of the
EC pool of PECAM-1; the hematopoietic pool appeared to
be more important for disease progression in the aortic arch.
Experiments using shear stress–modifying carotid cuffs in
Pecam-1–/– ApoE–/– and ApoE–/– mice also showed a reduction
in plaque burden in areas of low shear stress in mice deficient
in PECAM-1. Interestingly, in this model, we also found a
strong dependence on PECAM-1 for recruitment of mono-
cytes into the atheromatous environment.
Effects of PECAM-1 Ablation on the
Hematology and Blood Chemistry of
the Pecam1–/– ApoE–/– Mouse
Total white blood cell counts did not vary between ApoE–/––
deficient mice with a Pecam-1–/– genotype or the control groups
of ApoE–/– mice, which were Pecam-1+/– or Pecam-1+/+ on either
dietary regimen (chow versus HFD; Tables I–III in the online-
only Data Supplement). However, ablation of PECAM-1 did
result in a small but significant increase in the percentage of
circulating peripheral blood lymphocytes, which in chow-fed
animals was associated with a significant reduction in the
percentage of circulating monocytes (Table I in the online-
only Data Supplement). In animals on HFD, there was a trend
toward lower phagocyte (monocytes and polymorphonuclear
neutrophils) numbers in the circulation, but this was not signifi-
cant. Triglyceride and cholesterol levels were elevated signifi-
cantly by HFD; although triglyceride levels were not sensitive
to alterations in genotype, cholesterol levels in ApoE–/– mice
that were Pecam-1+/– were significantly reduced compared with
other genotypes in both dietary regimes. The loss of PECAM-1
did not alter the rate of weight gain over the duration of these
experiments (Table II in the online-only Data Supplement).
Site-Specific Effect of PECAM-1 Ablation
on Atheroma Formation in the Aorta
of the Pecam1–/– ApoE–/– Mouse
In mice on a chow diet, plaque burden over the whole aorta was
low (≈7%), and was not affected by genotype. A full analysis
of the animals on this dietary regimen can be found in Figure
I the online-only Data Supplement. ApoE–/– animals fed a HFD
showed an increase in plaque burden compared with chow-
fed animals. Interestingly, the loss of PECAM-1 led to a net
elevation (≈25%) in disease burden over the whole aorta (Figure
1A–1C). However, detailed analysis showed that the effects of
PECAM-1 ablation were complex and site-specific. Thus, there
was significant reduction in plaque burden in the aortic arch,
particularly in the inner curvature (Figure 1D). In the same
animals, the loss of PECAM-1 led to a significant elevation
in disease burden in the thoracic and abdominal aortas (ie, the
descending aorta; Figure 1E). No effect of the loss of PECAM-1
was found on atheroma formation in the aortic sinus (Figure
II the online-only Data Supplement). In the descending aorta,
atheroma often occurs at the sites where branching arteries form
areas of disturbed flow. In a more detailed analysis (Figure 2), we
observed that plaques in the descending aorta were, in general,
associated with branch points of smaller arteries, although in
some instances, especially in the abdominal aorta, plaques were
large enough to encompass an area of vessel wall that included
number of branching arteries (data not shown). Interestingly, the
number of plaques in the descending aorta did not vary with the
loss of PECAM-1; however, the average size of the plaques was
significantly larger in Pecam-1–/– ApoE–/– mice. Interestingly, the
site-specific effects of the ablation of PECAM-1 on atheroma
formation were maintained for an extended period. Thus, in
Pecam-1–/– ApoE–/– mice fed a HFD for 24 weeks, there was a
≈23% increase in plaque burden over the whole aorta compared
with the control group (Figure IIIA–IIIC in the online-only Data
Supplement). Importantly, plaque burden remained significantly
reduced in the arch of the aorta (Figure IIID in the online-
only Data Supplement), whereas disease was more extensive
in the descending aorta (Figure IIIE in the online-only Data
Supplement) in the Pecam-1–/– ApoE–/– mice. Taken together,
these experiments show that the loss of PECAM-1 has either
proatherogenic or antiatherogenic effects, which appear to be
dependent on the local hemodynamic environment.
Effect of Losing Either Endothelial or Bone
Marrow PECAM-1 on Plaque Burden
Because PECAM-1 is expressed widely in cells of hema-
topoietic origin, as well as EC, we generated bone marrow
696 Arterioscler Thromb Vasc Biol April 2013
(BM) chimeras to differentiate the function of these pools.
First, in control experiments, it was important to show that
patterns in the change of disease burden in irradiated animals
was similar to that reported for nonirradiated animals. Thus,
ablation of BM in the ApoE–/– mouse followed by reconstitution
with ApoE–/– BM allowed formation of atherosclerosis (Figure
3A). Importantly, the ablation of BM in the Pecam-1–/– ApoE–/–
followed by reconstitution with BM from Pecam-1–/– ApoE–/–
animals (Figure 3A–3C) showed an increase in disease across
the whole aorta, with a decrease in disease burden in the aortic
arch and increased disease in the descending aorta (Figure 3A–
3C), which phenocopied the nonirradiated animals. Further
analysis in chimeric animals indicated that differences in disease
susceptibility were based on the presence of the different pools
of PECAM-1. Thus, in the arch (Figure 3B), reconstitution of
the Pecam-1–/– ApoE–/– animals with Pecam-1+/+ ApoE–/– BM
(Figure 3B) significantly exacerbated disease, when compared
with Pecam-1–/– ApoE–/– mice receiving Pecam-1–/– ApoE–/– BM
(Figure 3B) and Pecam-1+/+ ApoE–/– mice receiving Pecam-1+/+
ApoE–/– BM (Figure 3B). In contrast, the opposite construct (ie,
reconstituting EC PECAM-1 in the double knockout mice) had
no significant effect when compared with Pecam-1–/– ApoE–/–
mice receiving Pecam-1–/– ApoE–/– BM. We interpret this to
show a dominant effect of the BM pool in disease progression
at this site. Interestingly, in the descending aorta (Figure 3C),
reconstitution of Pecam-1–/– ApoE–/– with BM from Pecam-1+/+
ApoE–/– mice (Figure 3C) had no significant effect on disease
compared with Pecam-1–/– ApoE–/– mice receiving Pecam-
1–/– ApoE–/– BM (Figure 3C). However, the opposite construct
(ie, reconstituting the Pecam-1+/+ ApoE–/– mice with Pecam-
1–/– ApoE–/– BM) significantly reduced disease in comparison
with Pecam-1–/–ApoE–/– mice receiving Pecam-1–/– ApoE–/– BM
(Figure 3C). Thus, in the descending aorta, the presence of
PECAM-1 in the BM pool had no significant effect on disease
progression when compared with the Pecam-1–/– ApoE–/– mice
receiving Pecam-1–/– ApoE–/– BM (Figure 3C). However,
the EC pool had antiatherogneic properties at this site and
conferred rescue from disease. Interestingly, however, the
BM pool significantly increased disease progression in the
descending aorta when compared with the Pecam-1+/+ ApoE–/–
mice receiving Pecam-1+/+ ApoE–/– BM.
PECAM-1 Is Proatherogenic in Regions of Low
Shear Stress Generated by Restrictive Carotid Cuffs
We used surgically implanted shear stress–modifying carotid
cuffs to generate areas of defined low shear stress in the
common carotid artery in which plaque was evident (Figure
4A).31 Extensive control studies for this model have been
described in detail in Cheng et al.32 Briefly, these studies used
nonflow-altering control cuffs (constant diameter) that were
placed around the vessel touching the adventitial layer, to test
for nonspecific effects. Short-lasting placement (7 days) did not
change endothelial nitric oxide synthase expression, and long-
lasting placements (9 weeks; unpublished data) did not induce
plaques or inflammatory responses. Analysis after 9 weeks of
HFD showed that there was no disease in the common carotid
artery of ApoE–/– control mice undergoing sham surgery
(Figure 4B). In the ApoE–/– mouse, a large plaque developed in
the low shear environment, no plaque was evident in the high
shear region, and a small plaque was found downstream of the
cuff in the region with oscillatory-like patterns of flow (Figure
4A and 4B), patterns in agreement with other reports using
this methodology.31 In the common carotid artery of ApoE–/–
/Pecam-1–/– mice, a significant reduction in plaque burden was
Figure 1. Site-specific effects of platelet-endothelial cell adhesion molecule (PECAM)-1 in ApoE–/– mice on high-fat diet (HFD) for 13
weeks. ApoE–/– mice that were wild-type (WT), heterozygous, or deficient in PECAM-1 were placed on a HFD at 10 weeks of age for a
total of 13 weeks. A, Plaque burden and oil red O (ORO) extraction over the whole aorta was analyzed (B). C, Representative images
of the whole aorta. D, Image analysis of plaque burden found in the aortic arch, inner and outer curvature, and the whole descending
aorta as well as its parts, thoracic and abdominal aorta (E). *P<0.05; and **P<0.01 Pecam-1–/– ApoE–/– mice compared with ApoE–/– mice.
$P<0.05; and $$P<0.01 Pecam-1–/– ApoE–/– mice compared with Pecam-1+/– ApoE–/– mice by unpaired Student t test; n=7 to 11.
Harrison et al Site-Specific Effects of PECAM-1 in Atheroma 697
seen in the low shear region (Figure 4B). Cross sections of
the carotid arteries were also examined, and oil red O staining
confirmed that lack of PECAM-1 dramatically diminished
the deposition of lipid within the artery wall in the low shear
region (Figure 5A and 5B). Interestingly, even though there
was no change in plaque burden or in lipid deposition in the
region subjected to oscillatory shear, collagen deposition was
significantly reduced in this area (Figure 5C and 5D). Analysis
of the common carotid artery for macrophage content also
revealed interesting variations between hemodynamic
environments. Thus, plaques in the low shear environment
were rich in CD68+ cells, but ablation of Pecam-1 significantly
reduced the presence of macrophages in the double knockout
mouse (Figure 6). No significant differences were found in
smooth muscle cell actin content or CD3+ T-cells between
mouse groups (data not shown).
PECAM-1 is expressed by EC and hematopoietic cells,
such as leukocytes and platelets. It is well known to act as a
transducer of mechanical signals in EC,19,20,33–35 and also to
regulate migration of leukocytes through endothelium.12,36,37
In the past decade, evidence that PECAM-1 is important
in cardiovascular disease has emerged. Polymorphisms of
PECAM-1 have been linked to coronary artery disease and
myocardial infarction in various ethnic groups.24–27 However,
the functional basis of such pathogenesis is unclear, and to
attempt to clarify this, a number of studies have now been
conducted in whole-body PECAM-1 knockouts superimposed
on murine models of atherosclerosis (the low-density
lipoprotein receptor [LDLR]–/– or apolipoprotein E [ApoE]–/–
mice). Unfortunately, even separate studies conducted on
mice with the same genetic background have used different
dietary regimen and assessed disease at different time points,
which makes comparative interpretation problematic.
However, previous reports do agree that regulation of
disease burden by PECAM-1 is strongly associated with
differences in the local hemodynamic environment.
Unfortunately, even these interesting variations are not
consistent between models, or even between studies using
the same model. Thus, to understand the role of PECAM-1
in atherogenesis more thoroughly, we generated Pecam-
1–/– ApoE–/– mice and evaluated 2 models: natural variation
in disease in different regions of the arterial tree, which
experience different patterns of blood flow; and placement
of a carotid cuff to generate disease in defined areas of low
Figure 2. Increased plaque burden in the descending aorta after
platelet-endothelial cell adhesion molecule (PECAM)-1 aboli-
tion in apolipoprotein E (ApoE)–/– mice on high-fat diet (HFD) for
13 weeks. ApoE–/– mice that were wild-type (WT) or deficient in
PECAM-1 were placed on a HFD at 10 weeks of age for a total of
13 weeks. A, Total number of individual plaques, average plaque
size of the descending aorta was analyzed (B). *P<0.05, Pecam-
1–/– ApoE–/– mice compared with ApoE–/– mice by unpaired Stu-
dent t test; n=7 to 11.
Figure 3. The endothelial cell (EC) pool of platelet-endothelial
cell adhesion molecule (PECAM)-1 is responsible for the elevation
in plaque burden under high laminar flow regions of the aorta.
Bone marrow (BM) chimeras were generated by lethally irradiat-
ing 10-weeks-old female Pecam-1+/+ apolipoprotein E (ApoE)–/– or
Pecam-1–/– ApoE–/– mice, and reconstituting them with BM from
male Pecam-1+/+ ApoE–/– or Pecam-1–/– ApoE–/– mice. A, Plaque
burden over the whole aorta, the aortic arch (B), and the descend-
ing aorta were analyzed for all mouse groups (C). *P<0.05; and
**P<0.001, compared by Student t test; n=3 to 6.
698 Arterioscler Thromb Vasc Biol April 2013
shear stress and oscillating shear stress, which would be free
of disease otherwise.
Effects of PECAM-1 Ablation on Disease
Burden in the Whole Aorta
Using the first model to assess natural variation in dis-
ease, mice were subjected to a HFD-feeding regimen for
either 13 or 24 weeks. On general analysis, ApoE–/– mice
lacking PECAM-1 in EC and leukocytes showed elevated lev-
els of plaque in a simple analysis of total disease burden in the
whole aorta. This is in agreement with the study utilizing the
ldlr–/–/Pecam-1–/– mouse by Goel et al,30 however it contradicts
the reported decrease in total burden arising from PECAM-1
ablation in the ApoE mouse.28,29 When we used BM chimeras
to discriminate between the function of the EC and hemato-
poietic pools (BM), the EC pool had a strong regulatory role
Figure 4. Restriction of blood flow using carotid cuffs demonstrates the proinflammatory role platelet-endothelial cell adhesion mol-
ecule (PECAM)-1 has in regions of low shear. At 10 weeks of age, apolipoprotein E (ApoE)–/– mice that were wild-type (WT) or deficient
for PECAM-1 were placed on high-fat diet (HFD) for 2 weeks, when restrictive carotid cuffs were placed around the left common carotid
artery (LCCA). The mice were continued on HFD for a further 9 weeks. A, Schematic diagram of the carotid arteries and cuff placement,
and pictures of the carotid arteries from PECAM-1 WT or deficient ApoE–/– mice. B, Analysis of the plaque burden found in the low shear
(LS), high shear (HS), and oscillatory shear (OS) regions induced by the placement of the carotid cuff. *P<0.05, Pecam-1–/– ApoE–/– mice
compared with ApoE–/– mice by unpaired Student t test; n=3.
Figure 5. Cross section of left common carotid artery (LCCA) after the placement of carotid cuffs demonstrates that platelet-endothelial
cell adhesion molecule (PECAM)-1 has proinflammatory effects on plaque formation and collagen deposition. A, Cross sections of LCCA
stained with oil red O (ORO) (red) and hematoxylin (blue). B, Analysis of plaque burden as percentage of cross section. C, Cross sections
of LCCA stained with Sirius Red for collagen deposition (dark pink) and hematoxylin (blue). D, Analysis of plaque burden as percent-
age of cross section. *P<0.05; and **P<0.01, Pecam-1–/– apolipoprotein E (ApoE)–/– mice compared with ApoE–/– LCCA cross sections by
unpaired Student t test; n=5.
Harrison et al Site-Specific Effects of PECAM-1 in Atheroma 699
on disease progression so that, in the presence of EC PECAM-
1, disease was significantly reduced compared with animals
completely lacking in PECAM-1, however the presence of
BM PECAM-1 significantly increased disease compared with
animals either completely lacking or completely expressing
PECAM-1. Importantly, as the aorta is a complex vessel that
is subjected to variation in the hemodynamic environment in
a site-specific manner, such a crude analysis of plaque burden
is insufficiently stringent to describe the effects of PECAM-1
ablation on the anatomical patterning of atherosclerosis.
Effects of PECAM-1 Ablation on Disease
Burden in Areas of Low Shear Stress
In the aortic arch, which is an anatomic site subject to low
shear and oscillatory flow, we observed that total ablation
of PECAM-1 in ApoE–/– mice fed a HFD for 13 or 24 weeks
resulted in decreased plaque burden. More detailed analysis
of the aortic arch at the sites of the inner and outer curva-
ture revealed a decrease in plaque formation at the inner cur-
vature, but no significant difference at the outer curvature.
These observations are in broad agreement with all previous
reports, irrespective of the geneotype of the models used (ie,
ApoE–/–/Pecam-1–/– or ldlr–/–/Pecam-1–/– mice).28,30 Here, the use
of BM chimeras revealed that the presence of BM PECAM-1
significantly increased disease burden compared with both
PECAM-1 null mice and mice expressing PECAM-1 on both
the BM and EC pools. In contrast, however, abolition of EC
PECAM-1 had no significant effect on disease. We interpret
these data to show a strong proatherogenic role for the hema-
topoietic (BM) pool of PECAM-1 in the aortic arch.
In addition to analysis of the aorta, we also used shear-
modifying carotid cuffs to generate an area of low flow and
an atherosclerotic plaque in the carotid artery. This model
demonstrated broadly similar results on ablation of PECAM-
1. Thus, plaque burden was significantly reduced in ApoE–/–
Pecam-1–/– compared with ApoE–/– animals. Interestingly,
analysis of the content of these plaques demonstrated a
much reduced representation of macrophages in the cellular
infiltrate. These data, taken together with the demonstration
of the importance of the hematopoietic pool of PECAM-1
in our chimera studies, imply that monocyte PECAM-1 may
be important in supporting the trafficking of these cells into
plaques that develop in areas of low shear stress.
The above observations made in vivo are well supported by
previous studies of the effects of flow on EC, and the roles of
PECAM-1 in endothelial responses and in leukocyte migration.
First, exposure of EC to increasing levels of shear stress in vitro
leads to progressive inhibition of their response to inflamma-
tory cytokines; EC cultured at low shear stress are less prone to
recruit leukocytes than those cultured under static conditions,
but more prone than those cultured at high shear stress.21,22
Interestingly, oscillatory shear stress, which is often associated
with areas of low shear stress in the aorta, is itself proinflamma-
tory, inducing increased expression of adhesion molecules and
ability to bind monocytes.38,39 Acute exposure to oscillatory flow
itself can lead to nuclear translocation of nuclear factor kappa-
light-chain-enhancer of activated B cells, a response which is
reduced when PECAM-1 is absent.20,40 In addition, nuclear fac-
tor kappa-light-chain-enhancer of activated B cells activation
in EC, which can be observed in the aortic arch of mice, was
reduced in Pecam-1–/– mice.20 Thus, there is strong evidence
that loss of PECAM-1 should protect from atheroma in such
regions. The literature on the effects of PECAM-1 on leukocyte
migration is well established.12,36,37 In the case of monocytes,
especially, lack of PECAM-1 would be expected to reduce
migration and hence be protective against atheroma, although
the magnitude of this effect would depend on level of EC activa-
tion, which is the primary driver of leukocyte recruitment.
Effects of PECAM-1 Ablation on Disease
Burden in the Descending Aorta
Examination of the descending aorta revealed that PECAM-1
total knockout mice exhibited an increase in plaque deposition
within both the thoracic and abdominal aorta, leading to a net
increase in deposition within the descending aorta, a pheno-
type previously reported in the ldlr–/– model.30 However, other
studies in ApoE–/–/Pecam-1–/– models either report no change in
disease burden,28 or failed to investigate disease burden at this
anatomical site.29 Although the descending aorta is generally
thought of as an atheroprotected environment because of the
Figure 6. Platelet-endothelial cell adhesion molecule (PECAM)-1 contributes to macrophage recruitment into atherosclerotic lesion for-
mation in regions of low shear. A, Cross sections of left common carotid artery (LCCA) stained with for CD68 (brown) and hematoxylin
(blue). B, Analysis of macrophage content as percentage of cross section. *P<0.05, apolipoprotein E (ApoE)–/– mice deficient in PECAM-1
compared with ApoE–/– LCCA cross sections; and $$P<0.05, low shear regions compared with oscillatory regions by unpaired Student t
test; n=3 to 4.
700 Arterioscler Thromb Vasc Biol April 2013
levels of high shear lamina flow that are present, atheromatous
disease does develop, and this generally occurs at the points
where the intercostal, mesenteric, and renal arteries branch
from the aorta. At these points, blood flow is disturbed, and
areas of low shear stress and complex flow are present, which
appear to act as foci for the development of disease. When we
conducted a more detailed analysis of the descending aorta,
we observed that the number of disease foci was the same in
ApoE–/– and ApoE–/–/Pecam-1–/– animals. However, the average
size of plaques was significantly increased in the absence of
PECAM-1. Moreover, the distribution of disease was consis-
tent with development of atheroma at arterial branch points.
These data strongly indicate that atherosclerosis develops
at branch points in the descending aorta in the presence of
PECAM-1; however, in its absence, the burden of atheroma at
these foci of disease is exacerbated, so that individual plaques
occupy a greater surface area of the aorta.
When BM chimeras were used to discriminate between
the function of the EC and hematopoietic (BM) PECAM-1
in the descending aorta, it was clear that both the BM and EC
pools of PECAM-1 played a role in regulating disease burden.
Within the descending aorta, as within the aortic arch, the BM
pool of PECAM-1 appears to be proinflammatory, leading
to a net increase in plaque burden in comparison with both
the PECAM-1 null mice and the mice expressing PECAM-1
on both the BM and EC pools. However, in addition, and at
variance with the analysis in the aortic arch, the EC pool of
PECAM-1 also played a strong role in regulating disease bur-
den within the descending aorta. Indeed, EC PECAM-1 effec-
tively rescued PECAM-1 null mice from the additional burden
of atheroma. In other words, in the presence of EC PECAM-
1, disease progression was suppressed within the descending
aorta. The data from the whole-body PECAM-1 null mice
and from our chimeric studies strongly imply that the EC
pool of PECAM-1 provides antiatherogenic signals. As dis-
cussed above, exposure of EC to increasing levels of shear
stress in vitro leads to progressive inhibition of their response
to inflammatory cytokines. Importantly, we recently showed
that the inhibitory effect of laminar shear stress in such assays
was lost, when the experiments were conducted on human
EC in which PECAM-1 expression was reduced by treatment
with small interfering RNA duplexes, or when ECs isolated
from the Pecam-1–/– mouse were compared with those of wild-
type animals.23 Thus, loss of endothelial PECAM-1 would be
expected to make disease worse in any situation of laminar
flow, although these effects via EC would be more marked the
greater the local shear stress. Bearing in mind the observations
of our own studies and those in the ldlr–/–, 30 it is appropriate to
speculate that this aspect of PECAM-1 function is an impor-
tant regulator of disease progression, with high shear-induced
signaling into ECs limiting the extent of plaque formation.
Our findings, and those of a previous studies, including one
conducted in the ldlr–/–/Pecam-1–/– mouse, suggest that the
flow characteristics of the aortic arch (ie, oscillatory dis-
turbed flow) induce a proinflammatory environment in which
PECAM-1 expressed on hemopoetic cells (BM) promotes the
migration of leukocytes into tissue at sites of inflammation,
which result in increased disease burden. The presence or
absence of endothelial PECAM-1 at this site appears to have
little influence on the progression of disease. In regions with
high shear stress, such as the descending aorta, a proinflam-
matory environment is also produced and, similar to the situa-
tion in the aortic arch, PECAM-1 on leukocytes promotes their
migration into developing lesions. However, an additional
regulatory role for EC PECAM-1 is also evident. Thus, EC
PECAM-1 appears to send antiinflammatory signals into (or
to promote the antiinflammatory properties of) ECs, thereby
moderating the burden of disease at this anatomic site. Thus,
targeting EC PECAM-1 therapeutically would be expected
to worsen disease. However, concentrating on inhibiting the
proinflammatory properties of PECAM-1 in BM-derived cells
could potentially lessen the disease profile in both high shear
stress and oscillating flow regions of the aorta.
Sources of Funding
This work was supported by a British Heart Foundation project grant
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Platelet-endothelial cell adhesion molecule (PECAM)-1 is an adhesion and signaling molecule found on circulating blood cells (platelets and
leukocytes) and on endothelial cells which line blood vessels. PECAM-1 regulates the traffic of inflammatory leukocytes into tissue during
inflammation, and endothelial cells (EC) can sense patterns of blood flow through a mechanosensory receptor complex, which requires PE-
CAM-1. Here, we show that PECAM-1 plays a role in susceptibility or protection from atheroma formation dependent on the local patterns of
blood flow in the arterial circulation. Importantly, under all conditions of blood flow, PECAM-1 on circulating blood cells is proinflammatory.
However, in areas usually protected from atherosclerosis, which are marked by blood flow with a laminar profile and high shear stress, the
PECAM-1 on ECs induces protective responses, which moderate the burden of disease. Targeting PECAM-1 therapeutically in atherosclerosis
would thus need to be done with care and in a cell-specific manner.