This article is the introduction of a new thematic series on Wnts in Cardiovascular Development and Disease, which

includes the following articles:

An Updated Overview on Wnt Signaling Pathways: A Prelude for More [Circ Res. 2010;106:1798 –1806]
The Multiple Phases and Faces of Wnt Signaling During Cardiac Differentiation and Development [Circ Res.
2010;107:186 –199]
The Role of Wnt Signaling in Physiological and Pathological Angiogenesis

Wnt Signaling in Cardiac Hypertrophy and Remodeling: Lessons Learned From Cardiac Development

Wnt Signaling and Aging-Related Heart Disorders

Wnt Signaling and Stem Cells Michael Kühl, Guest Editor

WNT Signaling in Adult Cardiac Hypertrophy and Remodeling

Lessons Learned From Cardiac Development
Martin W. Bergmann
Abstract: On pathological stress, the heart reactivates several signaling pathways that traditionally were thought
to be operational only in the developing heart. One of these pathways is the WNT signaling pathway. WNT
controls heart development but is also modulated during adult heart remodeling. This review summarizes the
currently available data regarding WNT signaling during left ventricular (LV) remodeling. Upstream, soluble
frizzled-related proteins (sFRPs) block WNT-dependent activation of the canonical WNT pathway. By inhibition
of WNT activation, these factors also reduce ␤-catenin– dependent transcription by altering the ratio of
cytoplasmic/nuclear ␤-catenin. In experimental settings, sFRPs injected into the heart attenuated LV remodeling.
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sFRPs are secreted from autologous bone marrow– derived mononuclear cells. Disheveled is a signaling
intermediate of both the canonical and noncanonical WNT pathway. Similarly to the effect of sFRP, depletion of
a disheveled isoform attenuated LV remodeling. In contrast, disheveled activation led to progressive dilated
cardiomyopathy. Inhibition of nuclear ␤-catenin signaling downstream of the canonical WNT pathway
significantly reduced postinfarct mortality and functional decline of LV function following chronic left anterior
descending coronary artery ligation. WNT signaling also affects mobilization and homing of bone marrow–
derived vasculogenic progenitor cells. Finally, heart-specific WNT/␤-catenin interaction partners have been
identified that will possibly allow targeting this pathway in a tissue-specific manner. In summary, the WNT
pathway plays a pivotal role in adult cardiac remodeling and may be suitable for therapeutic interventions.
Currently, several molecular and cellular mechanisms whereby WNT inhibition attenuates LV remodeling are
proposed. Reactivation of the developmental program to restore functional LV myocardium from resident
precursor cells may significantly contribute to this process. (Circ Res. 2010;107:1198-1208.)
Key Words: heart failure 䡲 progenitor cells 䡲 angiogenesis 䡲 WNT 䡲 ␤-catenin

O n pathological stress, the heart reactivates several sig-

naling pathways that traditionally were thought to be
operational only in the developing heart.1 Recently published
data prove the adult heart to be capable of limited regenera-
tion from cardiac-endogenous precursors.2 Such adult cardiac
progenitor cells (CPCs) appear similar to prespecified cardiac
stem cells in the embryo.3 Canonical WNT signaling controls
the proliferation and differentiation of embryonic cardiac
precursors toward mature cardiomyocytes.4,5 Both positive
and negative WNT signaling is observed during cardiac

Original received July 10, 2010; revision received September 14, 2010; accepted September 21, 2010. In September 2010, the average time from
submission to first decision for all original research papers submitted to Circulation Research was 13.1 days.
From the Experimental and Clinical Research Center, Charité Campus Buch & Max Delbrück Center for Molecular Medicine, Berlin, Germany. Present
address: Department of Cardiology, ASKLEPIOS Klinik St Georg, Hamburg, Germany.
Correspondence to Priv-Doz Dr med Martin W. Bergmann, Department of Cardiology, ASKLEPIOS Klinik St Georg, Lohmühlenstr. 5, 20099
Hamburg, Germany. E-mail docbergmann@mac.com
© 2010 American Heart Association, Inc.
Circulation Research is available at http://circres.ahajournals.org DOI: 10.1161/CIRCRESAHA.110.223768

1198
 Bergmann WNT and Cardiac Remodeling 1199

remodeling of the adult heart on stress or injury: WNT

Non-standard Abbreviations and Acronyms
factors, as well as frizzled-related proteins inhibiting WNTs,
are detected.6 – 8 This review summarizes the available data Ang II angiotensin II
regarding WNT signaling in the adult heart. Because the role BMP bone morphogenic protein
of WNT in cardiac development may allow understanding of CPC cardiac progenitor cell
the mechanisms also relevant in adult cardiac remodeling, the CT control (mice)
review begins with a brief summary of the existing knowl- cTnT cardiac troponin T
edge regarding WNT signaling in cardiac precursor cells. DKK dickkopf
E embryonic day
WNT/␤-Catenin Signaling Axis in Embryonic Frz frizzled (WNT receptor)
and Adult Cardiac Precursor Cells IGF insulin-like growth factor
The heart develops from a common precursor. One of the IGFBP insulin-like growth factor– binding protein
earliest cardiac-specific transcription factors expressed both Isl-1 islet-1
in cultured embryonic stem cells, as well as in situ, is the KLF Krüppel-like factor
basic helix–loop– helix factor MesP1 (Figure 1).9 It is appear- LAD left anterior descending coronary artery
ing at approximately embryonic day (E)5.5 in the murine LV left ventricular
cardiac development and vanishes by E8.5.10 In vitro, MesP1 MHC myosin heavy chain
is required and sufficient to induce cardiovasculogenesis
MSC mesenchymal stem cell
from embryonic stem cells. The key factor downstream of
NLK nemo-like kinase
MesP1 is the inhibitor of the WNT/␤-catenin canonical
PKC protein kinase C
signaling cascade, dickkopf-1. Whether dickkopf-1 is the sole
PLC phospholipase C
transcriptional target of MesP1 responsible for cardiac cell
sFRP soluble frizzled related protein
fate determination remains a matter of debate: following
TG transgenic (mice)
MesP1 activation, the whole panel of cardiac transcription
machinery is found to be upregulated.9 –11 More precisely,
forced expression of MesP1 in ES cells leads to the appear-
ance of cardiomyocytes with intact gap junctions, direct
expression of Nkx2.5, MEF2c, and GATA4, as well as early differentiation of several independent cardiac precursor
sprouting of vasculogenic cells positive for von Willebrand cell pools from a common cardiac stem cell marked by
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factor (vWF) and CD31.11 In summary, inhibition of the MesP1 expression (Figure 1). These cells have the capacity to
canonical WNT- pathway via dickkopf appears to mediate restore functional myocardium in the adult heart following

Figure 1. Schematic summary of WNT/␤-catenin signaling regarding developmental and adult CPC proliferation and differentia-
tion. CPCs expressing markers of first and second heart field progenitors were identified in both the murine embryo and adult heart. To
date, no MesP1pos multipotent CPCs have been described in the adult heart. WNT/␤-catenin activation is required for amplification of
CPCs in both the embryo and the adult heart. WNT/␤-catenin inhibition, together with activation of other signaling pathways (ie, BMP),
not depicted in the scheme controls cell fate determination (ie, first vs second heart field or vasculogenic) and differentiation toward
contractile LV cardiomyocytes.
 1200 Circulation Research November 12, 2010
damage: a population of CPCs from rhesus monkey pluripo-
tent embryonic stem cells was able to integrate into an infarct
scar without forming teratomas. This maneuver replaced
⬇20% of the scar tissue in rhesus monkey heart with
contractile myocardial tissue.12
Similarly to the role of dickkopf downstream of MesP1, the
insulin-like growth factor binding protein (IGFBP)-4 was
found to induce cardiomyocyte differentiation from P19CL6
cells. Independent of IGF, IGFBP-4 leads to the expression of
cardiac troponin T and other markers of cardiomyocyte
differentiation in P19CL6 cells. IGFBP-4 attenuated activa-
tion of a ␤-catenin– dependent reporter gene on stimulation
with Frizzled 8. Several independent experiments then proved
IGFBP-4 to inhibit WNT signaling directly at the cell
membrane receptor complex by binding to Frz8 and LRP6.
Other pathways known to be involved in cardiomyocyte
differentiation, namely the bone morphogenic protein (BMP)
signaling cascade, were not affected. IGFBP-4 was not only
sufficient but also required for cardiomyocyte differentiation
in this system: knockdown of IGFBP-4 abrogated cardiomyo-
cyte differentiation.13 Interestingly, we identified IGFBP-5 to
be a target for ␤-catenin– dependent transcription: gene array
experiments with adult left ventricular (LV) tissue from mice
with heart-specific, conditional ␤-catenin stabilization (␤-
catenin␣MHC-⌬exon3) revealed IGFBP-5 to be a transcriptional
target of ␤-catenin.14 The role of this observation is currently
unclear because IGFBP-1, -2, -4, and -6 but not IGFBP-5 and
-3 were found to inhibit WNT-dependent signaling. Possibly
the IGF pathway itself is involved; forced, cardiac-specific
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expression of mIGF was found to attenuate LV remodeling
following cryoinjury. This was not mediated by the AKT
pathway but via the alternate pathway of phosphoinositide-
dependent protein kinase (PDK1) and serum glucocorticoid
kinase (SGK)1. The authors suggested endogenous regener-
ation to be involved in this process.15
In cardiac development, WNT/␤-catenin has a stage-
specific, at least biphasic, role.4,5 This state of affairs is
covered in more detail in another review of this series.16
Experiments with embryonic stem cell cultures revealed
WNT/␤-catenin inhibition plus activation of the BMP path-
way to enhance differentiation toward a cardiomyocyte phe-
notype but block hematopoietic and vascular lineage devel-
opment in the early stage (Figure 1).17 Similar observations
were made in vivo, where at approximately E5.5, upregula-
tion of WNT/␤-catenin is required to amplify the cardiom-
yogenic precursor cell pool of both the first and second heart
fields, giving rise to the left and right ventricle, respectively
(Figure 1).18 The segregation of these 2 distinct cardiogenic
lineages appears early and results in very distinct populations
of CPCs possibly also relevant to adult heart CPCs (Figure
2).19 The second heart field is marked by the expression of
Islet (Isl)1.20 Using conditional mice, where Cre expression is
driven by the Isl1 promoter, Notch1 was found to negatively
regulate ␤-catenin– dependent transcription.21 Inhibition of
␤-catenin resulted in upregulation of cardiac transcription
factors Myocd and Smyd1. In turn, these 2 factors were
required to drive differentiation of Isl1 CPCs to a cardiomyo-
cyte phenotype (Figure 3). In summary, ␤-catenin upregula-
tion causes expansion of different CPC lines including Isl1pos
second heart field progenitors. Negative regulation of
␤-catenin appears to drive CPC cell fate and differentiation.22
Similarly, the nuclear protein Chibby was found to facili-
tate cardiomyocyte differentiation from ES cells. Chibby
directly binds to ␤-catenin and antagonizes its transcriptional
activity (Figure 3). Chibby expression was associated with
upregulation of Nkx2.5, ␤-myosin heavy chain (␤MHC), and
Mef2c. Chibby expression is also found at high levels in adult
cardiomyocytes. Apparently, the cardiac-specific transcrip-
tion factor Nkx2.5 controls Chibby expression. The authors
suggested the clinical perspective that antagonizing the
WNT/␤-catenin pathway by this cardiac signaling molecule
will allow to promote CPC differentiation toward a cardio-
myocyte phenotype similar to cardiac development.23
Several other independent lines of evidence find positive
regulation of WNT/␤-catenin to be required for CPC ampli-
fication, whereas negative regulation of WNT/␤-catenin is
required for CPC differentiation toward adult cardiomyocytes
(Figure 1). In the murine embryo development, upregulation
of WNT/␤-catenin is required at the time of MesP1 activation
(approximately E5.5) to start amplification of the different
CPC pools. Most prominently, this was observed for first and
second heart field CPCs marked by expression of Tbx5 and
Isl1, respectively.24 At later stages (from approximately E7.5)
negative regulation of the WNT/␤-catenin pathway is re-
quired to drive differentiation of the 2 CPC populations to
adult cardiomyocytes.18,25 Other proteins regulated at this
time point of cardiac development include NKx2.5, the first
peak of ␣MHC, and GATA4 and eHand (first heart field) as
well as dHand (second heart field).26,27 It is often overlooked
that ␣MHC is long known to have 2 expression peaks: the
first peak is observed around E7.5 in the developing left
ventricle.28,29 Subsequently, ␣MHC is downregulated and is
replaced by ␤MHC. After birth, ␣MHC is upregulated again
and is replacing ␤MHC.30
We have developed a technique that allows for flow
cytometric analysis of adult heart CPCs in the noncardiomyo-
cyte cell fraction of either murine heart tissue or human left
atrial appendage.31 In brief, heart tissue is mechanically
minced and then subjected to 2 steps of size filtration where
the smallest mesh will filter for cells larger than 30 ␮m. This
technique is almost completely filtering adult cardiomyo-
cytes. This allows analyzing the noncardiomyocyte cell
fraction of the adult heart without alteration of protein
expression associated with the use of collagenase.16 Analysis
for several markers known from the developing heart to be
characteristic for CPCs revealed the results depicted in Figure
2. In the adult heart, to date, no MesP1pos CPCs have been
described. Similarly, only a few CPCs of the second heart
field marked by islet-1 expression have been detected. How-
ever, we readily detected CPCs positive for several markers
of the first heart field (Tbx5, ␣MHC, eHand) in adult murine
left ventricle, as well as human left atrial appendage (Figure
2). Modulation of proliferation and differentiation of first
heart field CPCs in the adult heart may allow boosting
endogenous regeneration. The precise knowledge of these
developmental pathways has implications for adult cardiac
remodeling as discussed below.
 Bergmann WNT and Cardiac Remodeling 1201
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Figure 2. Qualitative and quantitative analysis of adult heart cardiac progenitor cells. Murine adult heart tissue or human adult left
atrial appendage tissue was homogenized and size-filtrated for depletion of adult cardiac myocytes as described.31 The flow-through
containing the noncardiomyocyte cell fraction of heart tissue was analyzed by flow cytometry and immunofluorescence for expression
of cardiac progenitor cell markers. A, Quantitative assessment of CPC marker expression in the noncardiomyocyte cell fraction from
murine adult heart as analyzed by flow cytometry. Tbx5, ␣MHC, and eHand are specific for the developmental first heart field, giving
rise to the left ventricle. Islet-1 and dHand are specific for the second heart field, giving rise to the right ventricle and LV outflow tract.
cTnT is specific for mature adult cardiomyocytes. Nkx2.5 is expressed in both first heart field (FHF) and second heart field (SHF) pro-
genitor cells. Sca-1 is an unspecific marker of mesenchymal stem cells. Data summarize analysis of nⱖ6 animals. B, Qualitative analy-
sis of murine adult heart CPCs. FACS analysis finds ⬇14% of ␣MHCpos nonadult cardiomyocyte cells to have proliferative capacity as
detected by coexpression of Ki67. The cytoplasm/nuclear ratio is close to 1; also, immunofluorescence detects cells with costaining of
Ki67 as a marker of cell proliferation and ␣MHC as a marker of cardiomyogenic cell commitment. This confirms the presence of first
heart field CPCs in the adult heart. C, Qualitative analysis of human adult heart CPCs isolated from left atrial appendage (LAA). ␣MHCpos/
Tbx5pos/eHandpos CPCs with a cytoplasm/nuclear ratio of close to 1 and evidence for cell division by depicting an S-phase CPC can
be found. This suggests the presence of first heart field CPCs to be present also in human adult heart.

Soluble Frizzled-Related Proteins Attenuate
Adult Cardiac Remodeling Following
Experimental Infarct
WNT proteins are secreted glycoproteins that act locally in a
paracrine fashion. WNT proteins bind to frizzled receptors.
These are 7-pass transmembrane proteins characterized by an
extracellular N-terminal domain.32 sFRPs lack the transmem-
brane domain and compete locally for WNT binding (Figure
3). Interestingly, sFRP-3 and sFRP-4 levels were found to be
elevated in hearts of patients with both dilated cardiomyop-
athy and coronary heart disease.33 FrzA, related to mouse
sFRP1, has been detected during cardiovascular maturation
and in the adult heart. More specifically, sFRP1 is transiently
upregulated following experimental infarct predominantly in the
infarct border zone together with frizzled receptors 1, 2, 5, and
10, as well as WNT 10b. These data suggest sFRPs to be
involved in infarct healing and tissue homeostasis following
injury.7
FrzA/sFRP1 was proven to regulate vascular cell prolifer-
ation and to induce an angiogenic response.34 Moreover, a
genome-wide screen revealed sFRP2 to be the key stem cell
paracrine factor that mediates myocardial survival and repair
after experimental myocardial infarct treated with intracardial
injection of AKT-modified mesenchymal stem cells. This
paracrine secretion of sFRP2 lead to a dramatic reduction of
infarct size and restoration of cardiac function. These effects
were observed as early as 72 hours after AKT–mesenchymal
 1202 Circulation Research November 12, 2010
canonical WNT signaling upstream of GSK3␤. KLF15, Notch1, and Chibby are nuclear interaction partners of ␤-catenin blocking
␤-catenin/Tcf3-activated gene transcription. (Illustration Credit: Cosmocyte/Ben Smith).
stem cell implantation, clearly demonstrating the paracrine
secretion of sFRP2 to be the crucial mechanism whereby
AKT-modified mesenchymal stem cells attenuate maladap-
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tive LV remodeling following myocardial infarct.35
Similar to the observation described above concerning genet-
ically modified mesenchymal stem cells, an endogenous repair
mechanism was identified by comparing 2 different mice strains
inbred for the capability of accelerated healing following tissue
injury (“superhealer”). These mice are capable of increased
myocardial regeneration following cryoinjury in association
with a 3-fold increase of bone marrow– derived mononuclear
cells found in the heart. Comparing bone marrow– derived
CD45neg, CD11bneg, Sca-1pos, CD44pos mesenchymal stem cells
from the “superhealer” strain to a comparable wild type strain
identified sFRP2 to be the key paracrine factor secreted by
superhealer-derived mesenchymal stem cells. Wild-type mesen-
chymal stem cells transduced with sFRP2 and injected into the
periinfarct region recapitulated the superhealer phenotype fol-
lowing experimental myocardial infarct.8 In conclusion, several
lines of evidence suggest sFRP2 to be required and sufficient to
mediate myocardial repair following ischemic injury. In line
with the general function of sFRPs known to sequester WNTs
away from the active receptor complex, sFRP2 was shown to
significantly inhibit ␤-catenin transcriptional activity as assessed
by reporter gene activation and target gene expression.36
Global knockout of sFRP2 lead to reduced cardiac fibrosis
and improved function after experimental myocardial infarction.
The proposed mechanism was increased activation of metallo-
proteinases that are inhibited in the presence of sFRP2. It
remains unclear in this study whether knockout of sFRP2
affected WNT pathway activation. In addition, a global knock-
out of sFRP2 might affect WNT signaling in other tissue
Figure 3. Schematic summary of sig-
naling molecules described to inhibit
WNT/␤-catenin signaling in the car-
diac compartment. Inhibition of WNT
and several downstream signaling
components including ␤-catenin– de-
pendent transcription was found to
attenuate LV remodeling and improve
adult cardiac function following ische-
mia or pressure overload. This scheme
therefore summarizes both positive
and negative signaling components of
the WNT pathway in the cardiovascular
system. These signaling intermediates
may be suitable for therapeutic inter-
ventions. For example, sFRPs were
found to be released from bone mar-
row– derived mononuclear cells homing
to the heart after injury. Experimentally
injected or genetically upregulated
sFRPs generate similar responses.
sFRPs scavenge paracrine-secreted
WNTs, preventing them from binding to
membrane-bound Frz receptors. sFRPs
therefore inhibit both the canonical, as
well as the noncanonical, WNT path-
way. Disheveled binds to FrzR and is
required for activation of the nonca-
nonical and probably also the canoni-
cal pathway (see text). Dickkopf blocks
compartments like the bone marrow important for mobilization
of endothelial progenitor cells that indirectly could influence LV
remodeling following experimental infarct.37
Transgenic mice with cardiac-specific overexpression of FrzA
(similar to mouse sFRP1) resembled the phenotype described
above: ␣MHC-dependant FrzA expression increased survival by
preventing myocardial rupture following experimental infarct.
Mechanistically, this phenotype was associated with reduced
infarct size, less apoptosis, increased collagen deposition, and
reduced activity of metalloproteinase-9 activity. This latter point
is in line with the finding in global sFRP2 knockout mice,
although the functional phenotype is opposite.37 Interestingly,
the percentage of muscularized vessels was significantly higher
in mice with ␣MHC-dependent FrzA/sFRP1 overexpression,
together with an increase in open vessel area.7 In addition,
FrzA/sFRP1 is sufficient and required for the effect of precon-
ditioning on infarct area: in the absence of FrzA/sFRP1, no
effect of preconditioning is observed, which can be restored by
the conditional, ␣MHC-dependent FrzA expression.38 Similarly,
direct injection of sFRP2-secreting mesenchymal stem cells in to
the border zone of an experimental infarct decreased infarct area,
increased ejection fraction and reduced dilation of the left
ventricle in association with increased vascular density. No
increase in angiogenic-positive (PECAM-1) or cardiomyocyte
marker–positive (anti-␣-actinin) cells was observed, implicating
a paracrine mechanism rather than any form of direct transdif-
ferentiation of mesenchymal stem cells.8 Direct treatment of
isolated adult rat ventricular cardiomyocytes with sFRP2 re-
duced caspase activity and exerted a cytoprotective effect via
upregulation of Birc1b.35
In summary, the majority of data finds expression of sFRPs
in the heart following experimental infarct to be beneficial for

Figure 4. Schematic summary of cellular mechanisms con-
tributing to the observed attenuation of LV remodeling on
inhibition of WNT signaling at different levels. Several mecha-
nisms have been proposed whereby WNT signaling inhibition
improves cardiac function in the adult following damage. Inhibi-
tion of cardiac hypertrophy is possibly related to the Ca2⫹-
dependent pathways as part of noncanonical WNT signaling.
WNT signaling may influence cardiomyocyte apoptosis and sur-
vival via PKB/AKT and G␣q. Mobilization of vasculogenic pro-
genitors via canonical ␤-catenin may enhance angiogenesis.
Increased differentiation of cardiac-endogenous first heart field
CPCs toward functional adult cardiomyocytes may contribute to
improved contraction. In addition, WNT signaling affects secre-
tion of metalloproteinases, thereby altering cardiac fibrosis and
extracellular matrix composition.
cardiac remodeling (Figure 4).7,8,34 –36,38 – 40 The mechanism is
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not entirely clear, and the published literature suggests a
cardiomyocyte cytoprotective effect, as well as increased
vascular density in the infarct area (Figure 4). However, the
general role of WNTs in other tissue compartments always
found activation rather than inhibition of the WNT pathway
to mediate antiapoptotic effects via activation of PKB/AKT.41
In addition, WNT activation is crucial for angiogenesis in
heart development.42 The mechanism whereby WNT inhibi-
tion by secreted frizzled related proteins improves cardiac
remodeling therefore remains unclear. A detailed analysis of
signaling molecules downstream of WNT/frizzled interaction
may allow identifying this missing link.
WNT activates the noncanonical pathway including the
axis of phospholipase C (PLC). PLC in turn leads to calcium
release within the cell. Calcium release induces Ca/calmod-
ulin kinase (CaMK)II and calcineurin activation (Figure 3).32
Inhibition of CaMKII and calcineurin including downstream
transcription factors like NF-AT and MEF2 was demonstrated to
inhibit cardiac hypertrophy and attenuate LV remodeling.43
However, the dramatic effects exerted by sFRP-expression on
LV-remodeling following experimental infarct are significantly
more pronounced than those observed in transgenic mice with
CaMKII, NF-AT, or MEF2 deletion. Inhibition of cardiac
hypertrophy and/or cardiomyocyte apoptosis is not sufficient
to explain the phenotypes observed.44 Therefore, the WNT-
mediated activation of ␤-catenin-dependent transcription in
the canonical pathway appears to be central to the observed
phenotype. WNT leads to inhibition of GSK3␤ via intracel-
lular binding of axin, thereby preventing ␤-catenin phosphor-
ylation and subsequent degradation by the proteasome.32 The
Bergmann WNT and Cardiac Remodeling 1203
available data regarding the WNT signaling intermediate
protein disheveled and ␤-catenin in LV remodeling are
summarized below.
Disheveled Is Required and Sufficient to
Induce Maladaptive Adult Heart Remodeling
Disheveled is an intracellular protein that is an essential
positive signal intermediate of both the WNT-initiated canon-
ical but also noncanonical signal transduction cascade.45
These branches are interconnected as described in detail in
another part of this review series.46 Some data support the
notion that the canonical/noncanonical branches separate at
the level of disheveled where activation of disheveled is
specific for the noncanonical pathway.32 Mutants in drosoph-
ila however indicate disheveled to be part of the membrane-
bound protein complex of frizzled receptor regarding all
downstream signaling pathways (Figure 3). Additional stud-
ies may clarify this point in mammalian cells.
More precisely, downstream of the frizzled receptor 3
branches are activated: the canonical WNT/␤-catenin axis
and the 2 noncanonical pathways Rho/Rac/JNK and WNT/
Ca2⫹ (Figure 3). In the noncanonical Jun-kinase pathway,
disheveled activates Rho/Rac via Daam1. This pathway has
been linked to cytoskeletal organization and cell polarity.
Disheveled also links the intracellular domain of the Frizzled
receptor to activation of phospholipase C (PLC). Downstream
of PLC and the subsequent intracellular calcium release,
Calcium/calmodulin kinase II is activated linking Ca-
dependent signaling to frizzled/disheveled stimulation.
CaMKII is required and sufficient to drive the hypertrophic
response in adult LV remodeling on injury and stress.47
Disheveled expression was shown to be upregulated on
transaortic banding (rat) and atrial-fibrillation induced heart
failure (porcine).6 Cardiac-specific overexpression of dishev-
eled under the control of an ␣-MHC promoter results in
severe cardiomyopathy at 3 months of age with significantly
enhanced mortality attributable to pulmonary congestion. The
phenotype resembles a dilated cardiomyopathy with grossly
enlarged hearts (end-diastolic diameter of 5.5⫾0.2 mm [TG]
versus 4.4⫾0.1 mm [CT]) and reduced ejection fraction. A
compensatory increase in myocyte cross-sectional area and
myocardial fibrosis was observed. Downstream of dishev-
eled, the 2 noncanonical pathways of JNK and CaMKII, as
well as c-Myc, were constitutively activated. Mechanistically,
cell culture experiments with small interfering RNA–medi-
ated knockdown of disheveled suggest the protein to be
required and sufficient for ␤-adrenergic (isoproterenol)-
induced cardiomyocyte hypertrophy. Activation of the canon-
ical WNT pathway with ␤-catenin as the downstream target
may also contribute to this phenotype.48 Interestingly, myo-
fibroblasts known to contribute to myocardial wound healing
are also regulated by WNT signaling. Migration of cardiac
fibroblasts immortalized by stable transfection of telomerase
was attenuated by active WNT signaling. The study indicates
that myofibroblast migration and differentiation, but not
proliferation, can be modulated by interventions in Wnt/Frz
signaling.49 In summary, attenuation of cardiac fibrosis after
infarct may also contribute to the positive effects on LV
remodeling seen by WNT inhibition (Figure 4).
 1204 Circulation Research November 12, 2010
Similarly to this heart-specific gain-of-function mutation
regarding disheveled, a disheveled isoform knockout strain
was characterized regarding an adult cardiac phenotype. No
baseline phenotype was observed. Because ␤-catenin knock-
out mutations are embryonic lethal this either indicates that
disheveled is predominantly involved in the noncanonical
WNT pathway or other isoforms compensate for the loss of
disheveled 1. Interestingly, cardiac hypertrophy as assessed
by heart weight, LV wall thickness in echocardiography and
gene expression (atrial natriuretic factor, brain natriuretic
peptide) on pressure overload was attenuated in disheveled 1
knockout animals.50 The cellular mechanism of this observa-
tion remains unclear; an association to increased GSK3␤ and
AKT phosphorylation was described. The phenotype though
is also similar to knockout models of the noncanonical
pathway, ie, modulations of CaMKII.51
The phenotype in disheveled-1 knockout mice might also
be linked to the role of WNT signaling in cardiac develop-
ment and adult cardiac remodeling already summarized
above. Specifically, disheveled is both necessary and suffi-
cient for the fusion of early heart precursors. Diversin links
the Frz/disheveled membrane complex to the downstream
noncanonical pathway with activation of Rho and Rac.
Diversin depletion resulted in 2 independently beating hearts
in the zebrafish. This phenotype of cardia bifida was rescued
by RhoA injection.52 The data prove the noncanonical WNT
pathway to be important for cardiac development. Given the
role of disheveled /diversin in WNT signaling, these signaling
molecules may be interesting therapeutic targets because the
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downstream effectors RhoA, JNK, and CaMKII are known to
be important mediators of maladaptive cardiac remodeling.
Frizzled receptors couple to heterotrimeric G proteins includ-
ing G␣i and G␣q.53 This state of affairs links WNT signaling to
the activation of MAPK JNK. The role of G␣i and G␣q, both
classic 7-transmembrane receptors, in cardiac hypertrophy is
well established.54 WNT activation also leads to recruitment of
factors like ␤-arrestins, which mediate signal transduction from
G␣i to the MAPK signaling cascade.53 In summary, several lines
of evidence support both the noncanonical and the canonical
WNT pathway as involved in WNT/Frz-mediated maladaptive
cardiac hypertrophy.
Evidence for ␤-Catenin Inhibition to
Attenuate LV Remodeling
More than 90% of the cellular ␤-catenin protein content is
bound in the membrane. Loss of ␤-catenin, however, does not
lead to a “membrane” phenotype because plakoglobin (␥-
catenin) functionally substitutes for ␤-catenin in the mem-
brane in various tissue compartments including the heart.55
Modulations of ␤-catenin levels in the heart by WNT signal-
ing or genetic modifications therefore predominantly affect
␤-catenin– dependent transcription.
Both gain- and loss-of-function mutations of ␤-catenin are
embryonic lethal between E5.5 and E10.5.32 To study the role
of ␤-catenin in the adult heart, several conditional mutants
have been generated. Such mice exhibited no phenotype
under baseline conditions.14,56 On pressure overload, chronic
angiotensin (Ang) II stimulation or experimental infarct
␣MHC-dependent ␤-catenin loss-of-function mutations led to
attenuated LV remodeling and improved ventricular func-
tion.14,31,55 Conversely, increased mortality and a phenotype
of dilated cardiomyopathy were observed in ␤-catenin gain-
of-function mutations.14,31 Conditional deletion of ␤-catenin
was achieved by either using the ␣MHC-MerCreMer or
␣MHC-CrePR1 strains. When studied in a TAC model of
pressure overload, mice with ␣MHC-MerCreMer mediated
␤-catenin deletion subjected to the same gradient as control
mice had significantly reduced cardiac hypertrophy (heart
weight CT: 138⫾11 g versus TG: 111⫾4 g) with increased
fractional shortening (CT: 36.2⫾2.7%; TG: 39.1⫾2.4%) and
less LV dilation (echo LV end diastolic dimension: CT,
3.56⫾0.14 mm; TG, 3.5⫾0.09 mm).56
We studied both gain-of-function (␣MHC-CrePR1⫻⌬N-
catenin) and loss-of function (␣MHC-CrePR1⫻␤-cat⌬ex3-6) mu-
tants on 2 weeks of Ang II stimulation. Mice with stabilized
␤-catenin in ␣MHC-positive cells had severely impaired
fractional shortening at the end of the treatment course (TG:
22.⫾4.5% versus CT: 36.2⫾1.6%), whereas mice with
␤-catenin depletion had no significant phenotype compared
to their control littermates.14 Following chronic ligation of the
left anterior descending coronary artery, ␤-catenin loss-of-
function mutants had significantly decreased mortality, re-
duced infarct size, and improved fractional shortening four
weeks after the infarct.31 No significant effect on ejection
fraction was observed at 2 weeks post infarct, suggesting that
modulation of LV remodeling rather than an acute effect on
the infarct size is responsible for this functional effect. This
phenotype is very similar to the observations following
expression of soluble frizzled receptors in the border zone of
the infarct.7,8,35
Several mechanisms whereby ␤-catenin depletion im-
proves LV function following pressure overload or ischemia
have been described (Figure 4). Among other targets, we
consistently found ␤-catenin to regulate Tbx5, a key tran-
scription factor in differentiation of first heart field progeni-
tors toward a cardiomyocyte phenotype.31 In association with
the maladaptive LV remodeling observed in gain-of-function
␣MHC-restricted ␤-catenin mutants subjected to chronic Ang
II stimulation, heart Tbx5 gene expression was reduced. In
contrast, Tbx5, as well as GATA4, was upregulated in heart
tissue of transgenic mice with ␤-catenin loss-of-function
mutations restricted to cells with activation of the ␣MHC-Cre
construct. This phenotype was associated with increased
differentiation of cardiac-endogenous progenitor cells toward
a cardiomyocyte cell type in cell culture experiments with the
noncardiomyocyte fraction of heart cells from such trans-
genic mice. The cells affected expressed ␣MHC and Tbx5
protein but not troponin T or islet-1.31 This suggests first heart
field progenitors to be a target for negative WNT signaling in
the adult heart. Such cells could be identified both in the
murine left ventricle and in specimens from human left atrial
appendage (Figure 2). The amount of cells present in the adult
heart as quantified by FACS of the noncardiomyocyte cell
fraction from murine left ventricle is sufficient to link
differentiation of these cells to the phenotype of attenuated
LV remodeling in ␣MHC/␤-catenin– depleted mice. The
identification of specific markers for first heart field CPCs in
the adult heart is required to further analyze this hypothesis;

Tbx5, ␣MHC, and other currently known markers are not
suitable because they are also expressed in adult
cardiomyocytes.
WNT Inhibition Enhances Mobilization of
Endothelial Lineage Progenitor Cells From
Bone Marrow
As described above, the embryonic heart develops from
several different CPCs. Aside from first heart field CPCs
marked by ␣MHC and Tbx5, cells from the endothelial
lineage are also affected by WNT signaling.57 WNTs play a
key role in embryogenic vasculogenesis by modulating ex-
pansion of primitive VEGF receptor2pos and postnatal angio-
genesis.40 Mobilization and homing of vasculogenic progen-
itor cells have shown potential to enhance myocardial tissue
repair via enhanced neovascularization in the adult.58 Modu-
lation of the WNT/␤-catenin signaling axis in the adult might
allow for enhanced mobilization and homing of such cells
from bone marrow to the heart.59
The intracellular WNT signaling antagonist dickkopf
(Dkk)-1 was studied in the context of vasculogenic progenitor
cells resident in the bone marrow. Recombinant Dkk1 injected
intraperitoneally induced mobilization of Flk1pos/sca-1pos pro-
genitor cells. Secretion of the osteoclast differentiation factor
RANKL was identified as the molecular mechanism.
RANKL in turn induced release of cathepsin K responsible
for selective mobilization of vasculogenic progenitor cells.
The inhibitory effect of Dkk-1 on WNT signaling was
measured by expression analysis of axin-2. This mobilization
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was sufficient to enhance in vivo vascularization of Matrigel
plugs implanted subcutaneously. Different from other growth
factors stimulating release of bone marrow– derived vascular
CPCs like G-CSF, Dkk1 did not lead to an induction of
inflammatory cells from the bone marrow. Even more intrigu-
ing was that the mechanism of vasculogenic cell mobilization
is quite different between the 2 stimuli: whereas G-CSF
reduces the amount of osteoclasts, Dkk1 indirectly cleaves
osteoclast-derived SDF1 via cathepsin K. SDF1 is required
for homing of such progenitor cells to ischemic areas includ-
ing the heart (Figure 4).59
Interestingly, Flk1pos cells are multipotent cardiovascular
progenitors that also form the endocardium and directly
respond to both BMP and WNT signaling regarding differ-
entiation once homed to the myocardium.60 In vitro, WNT
activation of such cells suppressed myocyte specification,
whereas early Noggin⫹Dkk1 exposure significantly en-
hanced cardiomyocyte specification. These data suggest that
the WNT pathway not only controls mobilization, prolifera-
tion, and differentiation of CPCs but also determines cell fate
decisions from early progenitors.60 Key markers of cell fate
decision monitored in this study were similar to other studies:
Tbx5 and GATA4 for first heart field CPCs, Isl1 for second
heart field CPCs and Flk1, and CD31 for hematopoietic/
vascular progenitor cells.
The canonical WNT pathway is activated by ischemic
events. In addition to affect endothelial cells it was found to
regulate smooth muscle cell proliferation and apoptosis.61
Several studies have analyzed the role of WNTs in peripheral
ischemia, which probably can be transferred to the ischemic
Bergmann WNT and Cardiac Remodeling 1205
situation in the heart regarding angiogenesis but not myogen-
esis. These studies support the notion that paracrine effects of
implanted cells are sufficient and required to enhance wound
healing and neoangiogenesis. One such study evaluated the
healing potential of human aorta-derived CD133pos progeni-
tor cells and their conditioned medium in an experimental
model of ischemic diabetic ulcer.39 The CD133pos cells were
found to initially secrete high levels of WNT molecules,
which were downregulated on differentiation into CD133neg
cells. The latter instead secreted soluble frizzled-related
proteins sFRP1, -3, and -4 known to downregulate WNT-
dependent signaling. Consistently, conditioned medium from
CD133pos cells accelerated wound healing and reparative
angiogenesis in comparison to vehicle.39 As depicted above,
sFRP2 was identified to be the key paracrine factor secreted
by mesenchymal stem cells injected into the heart to mediate
myocardial survival and repair after an ischemic event.
CD133pos cells have been studied in human trials on
chronic ischemic cardiomyopathy. One of the studies ana-
lyzed the effect parallel to surgical revascularization: patients
were subjected to autologous CD133pos cell transplantation
into the ischemic border zone during the operating procedure
after autologous bone marrow harvest the day before. The
therapy was safe and resulted in increased ejection fraction
and other parameters of LV function compared to a well-
matched control group.62
In summary, the WNT pathway is pivotal for vascular and
angiogenic development. Yet different conclusions depend-
ing on the experimental settings have been drawn whether
stimulation of WNTs ie, by paracrine activation or inhibition
of this pathway by dickkopf or sFRP is sufficient to improve
angiogenesis. One possible explanation is that biphasic control
of WNT signaling is required for a beneficial effect where WNT
activation promotes proliferation of the vasculogenic precursors,
whereas inhibition is required for mobilization, homing, and
differentiation. This interpretation would be consistent with the
observation that constitutive WNT stimulation leads to stem cell
exhaustion and multi-lineage blockade.63
Nuclear Negative Interaction Partners of
␤-Catenin Attenuate LV Remodeling in
Association With Cell Fate Determination
Krüppel-like factors (KLFs) are a large family of zinc
finger– containing transcription factors involved in regulating
cell differentiation, cardiac remodeling, hematopoiesis, an-
giogenesis, and stem cell fate determination by interacting
with coactivators and corepressors.64 Recent studies revealed
the important role of KLFs as regulators of cardiac biology.65
The family of KLFs was found to control ␤-catenin– depen-
dent transcription in several tissue compartments like the gut
and the heart. Possibly, KLF proteins allow for tissue specific
control of ␤-catenin transcription because KLF4 regulates
transcription in the small intestine whereas we found another
member of the same family, namely KLF15, to control
␤-catenin– dependent transcription in the heart.
Specifically, we searched for cardiac-specific ␤-catenin inter-
action partners able to modify the Wnt/␤-catenin transcriptional
activity specifically in the heart using a yeast 2-hybrid system
with a cardiac-specific library. Our own unpublished data
 1206 Circulation Research November 12, 2010
describe a novel interaction between the KLF15 and ␤-catenin
resulting in an inhibition of ␤-catenin/TCF3 transcriptional
activity via the N-terminal domain of KLF15 in cardiomyocytes.
Moreover, Nemo-like kinase (NLK), a Wnt/␤-catenin signaling
inhibitor, was found to interact with KLF15. NLK is activated
downstream of the noncanonical WNT pathway via CaMKII
linking the 2 pathways.
KLF15 is expressed at high levels only in the heart. KLF15
was previously reported to modulate cardiomyocyte hyper-
trophy and fibrosis under stress.66,67 We found that mice with
global KLF15 deletion develop normally with no apparent
phenotype at baseline. However, on aging and after patho-
logical stress the mice exhibit progressive cardiac deteriora-
tion whereas no apparent defects were observed in other
organs. The cardiac phenotype was associated with a reduc-
tion of cardiogenic precursor cells (Sca1pos/␣MHCpos and
Tbx5pos/cTnTneg)60 and upregulation of the vascular (CD31)
and endocardial Flk1 progenitor pool: the total CD31pos and
CD31pos/Ki67pos, as well as the Flk1pos/Ki67pos, CPC pool
was increased, as revealed by FACS analysis of the noncar-
diomyocyte fraction of the left ventricle from KLF15 knock-
out mice as compared to control mice. We conclude that
KLF15 determines CPC cell fate via ␤-catenin in the adult
heart. Moreover, these data support the concept that mainte-
nance of CPC-driven cardiomyocyte regeneration is required
for cell homeostasis during cardiac aging. In addition, cell
fate regulation is important for adaptive remodeling on
cardiac injury by ischemic events or increased afterload as
observed in arterial hypertension.
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As a note of caution to this concept, one must recognize
that until the identification of an unequivocally accepted
marker of adult LV cardiac progenitor cells, these data only
provide associations of heart physiology with biochemical
and cellular events in the CPC pool but no causal relationship.
New concepts are warranted to decipher the role of CPCs in
the adult heart. Interestingly, second heart field progenitors
giving rise to the right ventricle are marked by islet-1 and
have been characterized in detail.20 –22,68 Proliferation and
differentiation of islet-1pos cells isolated from the embryo are
also controlled by WNT/␤-catenin, however only few islet-
1pos cells could be isolated from the adult heart suggesting a
limited role of this precursor cell pool after birth. In contrast,
we found ⬇10% of the noncardiomyocyte cell fraction of the
left ventricle to express a reporter gene controlled by the
␣MHC-promoter with 2 different conditional mouse strains,
namely the ␣MHC-CrePR1 and the ␣MHC-MerCreMer.31
This cell fraction overlaps in part with previously described
adult heart CPCs, namely the c-kitpos cells, as well as the
sca-1pos cell population.31 However, both c-kit and sca-1 are
global markers of stem and progenitor cell pools rather than
being heart-specific. Future research needs to identify specif-
ically the cardiac progenitor cell pool; currently available
data suggest several different cell pools including locally
resident cardiomyogenic CPCs, as well as angiogenic cells
possibly homing from the bone marrow, to be involved in
adult cardiac remodeling.
On the basis of the currently available data from our group
and others, we believe that negative WNT/␤-catenin signal-
ing enhances first heart field CPC differentiation toward
functional cardiomyocytes in the adult heart. This is sufficient
for functionally relevant adult heart regeneration in the
context of aging as well as following injury.
Clinical Implications of Available Data
Regarding WNT Signaling in Adult
Cardiac Remodeling
The data summarized above prove the WNT pathway to be
required and sufficient for adult LV remodeling. Namely,
inhibition of the WNT pathway on several levels (soluble
frizzled receptors, dickkopf, inhibition of disheveled, ␤-catenin
depletion) has proven beneficial for cardiac remodeling (Figures
3 and 4). The noncanonical WNT pathway links WNT signaling
to well known players in adult cardiac remodeling, namely
RhoA and CaMKII. Given the new findings of an interplay
between the noncanonical and canonical pathways, the latter
comes into focus.46 The role of ␤-catenin in cardiac development
is now well established. The canonical WNT/␤-catenin pathway
has a conserved role in vertebrate heart development, regulating
and restricting cardiac precursor cell formation and subsequent
heart muscle differentiation.
Several lines of evidence, both in vitro and in vivo, suggest
cardiac CPCs and their responsiveness to WNT signaling to
be a possible therapeutic target. Some authors suggest the ex
vivo amplification by WNT stimulation and reinsertion of
amplified cells to be a favorable model.3 Given the high
prevalence of cells resembling first heart field cardiac pro-
genitors in the adult (Figure 1), boosting endogenous repair
mechanisms also appears feasible. One such approach build-
ing on the above summarized data would aim to inhibit WNT
signaling temporarily in the heart. This can be achieved by
local injection of bone marrow– derived mononuclear cells or
subpopulations thereof like CD133pos cells, which were
found to be capable of secreting sFRP.39 These molecules
scavenge WNT proteins and prohibit binding of WNTs to
membrane-bound frizzled receptors.
In summary, the WNT pathway plays a pivotal role in adult
cardiac remodeling and may be suitable for therapeutic
interventions. Among several currently discussed molecular
and cellular mechanisms whereby WNT inhibition attenuates
LV remodeling (Figure 4), reactivation of the developmental
program building the left ventricle from first heart field
progenitors in the embryo appears feasible. This maneuver
may restore functional LV myocardium from resident precur-
sor cells without the need of exogenous cell therapy. To prove
this hypothesis, consensus must be found regarding which
markers are specific for adult heart LV CPCs. It is now clear
that several different adult heart CPCs can be identified
similar to the situation in the developing heart. Tissue-
unspecific markers like c-kit or sca-1 are indirect markers that
not suitable for cell tracking. Islet-1pos cells will only be
relevant for the second heart field progenitors restricted to the
right ventricle and the left ventricle outflow tract in the
developing heart. The population of ␣MHCpos/cTnTneg/
Tbx5pos/eHandpos cells similar to first heart field progenitors
in the developing heart (Figure 2) is a possible candidate for
adult LV cardiac precursor cells. These cells are responsive to
negative WNT signaling and appear to be able to regenerate
functional LV myocardium in vivo.31 Whatever the mecha-

nism, inhibition of WNT in the adult heart may allow for
additional therapeutic interventions on top of currently avail-
able therapy.
Acknowledgments
M.W.B. apologizes for the many interesting articles that have not
been covered in this review because of space restrictions. M.W.B.
thanks Claudia Noack, MSc; Maria-Patapia Zafiriou, PhD; and Laura
Zelarayan, PhD (ECRC, Charité Campus Buch, & Max-Delbrück
Center, Berlin, Germany) for expert scientific work and fruitful
discussions. Invaluable technical assistance over many years by
Bärbel Pohl (MDC Berlin) formed the solid foundation of this work.
M.W.B. thanks Kai Jaquet, PhD (Department of Cardiology, St
Georg, Hamburg, Germany) and Stephan Geidel, MD (Department
of Cardio-thoracic surgery, St Georg, Hamburg, Germany) for
fruitful cooperation regarding human cardiac progenitor cells. Inter-
esting scientific discussions with Leon de Windt (Maastricht Uni-
versity Medical Center, The Netherlands) and Jean-Luc Balligand
(Unit of Pharmacology and Therapeutics, University of Louvain
Medical School, Brussels, Belgium) contributed to the opinions
expressed in this article.
Sources of Funding
Supported by Deutsche Forschungsgemeinschaft grant BE 2025/8-2
and grants from the Jürgen Manchot and the Grimmke Foundation.
Disclosures
None.
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