Editorial
Reprogramming the Beat
Kicking It Up a Notch
Sean M. Wu, MD, PhD; David J. Milan, MD
T
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regulation of transcript levels of many proteins typically
expressed in specialized conduction tissue. In the atrioventricular node, which had been shown previously to become
expanded in size after Notch activation,30 the increase in size
was shown to be the result of increased developmental
recruitment of cells into the conduction system, and not
because the cells were simply dividing more rapidly.
The timing of these effects is particularly interesting. At
birth, conduction system markers were not increased in the
ventricles of the Notch-activated hearts. However, by 3
weeks of age, these markers were observed outside the
regions where conduction tissues are normally found. These
results suggest that ongoing Notch activation not only influenced the cell fate decisions during early development, but
also appeared to do so even after birth.
These intriguing findings raise a number of questions that are
central to cardiac cell and developmental biology. For example,
are these cells truly conduction cells, or could they represent
ventricular cardiomyocytes that have been altered experimentally to express conduction cell proteins ectopically? To address
this question, the investigators characterized these Notch-activated cells functionally using patch clamp techniques. They
demonstrate that the Notch-activated cells not only express
markers of conduction cells, but they also exhibit action potential profiles with a characteristic plateau found uniquely in
Purkinje cells. Of interest, the degree of phenotypic conversion
was heterogeneous because there were also some cells that were
caught in transition with only a partial plateau, and some that
were complete nonresponders. These observations about the
relative efficiency of the conversion process are important and
raise interesting questions regarding the determinants of the
response to the Notch signal.
Although the in vivo data on Notch activation in developing cardiomyocytes address predominantly the question of
cardiac cell lineage decision, the question of cardiac cell
phenotype conversion (ie, reprogramming) by Notch activation remained open. To examine this, the investigators went
on to demonstrate further that Notch signaling in committed
neonatal cardiomyocytes also leads to increased conduction
tissue marker expression and prolonged action potentials,
supporting the notion that Notch signaling may be able to
reprogram working chamber cardiomyocytes toward a conduction cell fate. These results add to a growing body of
evidence that Notch signaling is a key determinant for cardiac
conduction system development, and that Notch activation
can influence cell fate decisions of the developing cardiomyocytes and may potentially convert mature postnatal cardiomyocytes into conduction-like cells.
These findings are particularly relevant given the intense
interest in direct cell lineage reprogramming in recent years.
he explosion of stem cell research in cardiology has
yielded an increasing recognition that understanding
developmental cell fate decisions is critical for everything
from disease models to cellular therapeutics. However, the
biology is particularly rich and complex and has not yielded
easily to traditional investigative methods. One area of
intense focus is the study of the cues responsible for the
development of specialized conduction tissues in the heart.
Direction of native cells or exogenous cells into the conduction system lineage might offer therapeutic insights into
degenerative conduction disease.
Article see p 1058
The most distal subendocardial fibers of the cardiac conduction system were first described by Jan Purkinje in 1839.1 Over
100 years ago, in painstakingly delicate dissections, Sunao
Tawara2 characterized the cardiac conduction system further,
describing the atrioventricular node, the penetrating bundle of
His, the bundle branches, and the Purkinje fiber network. During
the past 20 years, however, the pace of discovery of this highly
specialized tissue has accelerated. Beginning with the observations that peripheral and central conduction tissue derives
from cardiomyocyte progenitors in the chick,3,4 investigators have now identified key roles for many important
molecular signals and transcription factors in conduction
system development. Signaling molecules implicated in the
formation of the conduction system include endothelin,5–9
neuregulin,10 –13 Wnts,14 and bone morphogenetic proteins,15
and complex transcriptional roles have been identified for
NKx2.5,16 –20 members the T-box transcription factor family,18,21,22 shox2,23 msx2,24,25 and hop.26 –28
In this issue of Circulation, Rentschler et al29 take a
significant step toward understanding the determinants of
conduction cell fate. In a series of elegant experiments, they
demonstrate that activation of the Notch signaling pathway
can direct developing cardiomyocytes to become specialized
conduction cells. To do this, the investigators used a conditional transgenic mouse, one that turns on the Notch signal
only in developing cardiomyocytes, to demonstrate the upThe opinions expressed in this article are not necessarily those of the
editors or of the American Heart Association.
From the Cardiovascular Research Center, Division of Cardiology,
Department of Medicine (S.M.W. and D.J.M.), and Center for Human
Genetic Research (D.J.M.), Massachusetts General Hospital and Harvard
Stem Cell Institute, Cambridge, MA (S.M.W.).
Correspondence to David Milan, MD, Cardiovascular Research Center, Massachusetts General Hospital, 55 Fruit Street, Boston, MA 02115.
E-mail [email protected]
(Circulation. 2012;126:1009-1011.)
© 2012 American Heart Association, Inc.
Circulation is available at http://circ.ahajournals.org
DOI: 10.1161/CIRCULATIONAHA.112.126482
1009
1010
Circulation
August 28, 2012
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Inspired by the groundbreaking work of Takahashi et al31,32 to
generate embryonic stem cell-like cells from the enforced
expression of pluripotency-associated transcription factors,
investigators have now shown the ability of lineage-specific
transcription factors to induce neuronal, hematopoietic, pancreatic, and cardiomyogenic phenotypes in somatic cells.33–36
Although the primary intention of activating Notch in cardiomyocytes in this study was to examine the role of this
developmentally critical signaling pathway on the proper
formation of the cardiac conduction system, the finding of an
induced conduction cell phenotype in an otherwise committed cardiomyocyte places the results of this study in an
entirely different context. Namely, the triggering of a signaling pathway in committed cells may also lead to phenotypic
reprogramming.
To take this one step further, one could envision the
possibility that by manipulating signaling pathways alone, we
may be able to generate pluripotent stem cells, cardiomyocytes, neurons, pancreatic ␤-cells, and hematopoietic stem
cells without the need for viral or transfection-mediated
overexpression of transcription factors. Indeed, the requirement for bone morphogenetic proteins and fibroblast growth
factor 2 supplementation to maintain human pluripotent stem
cells, and the enhancement of pluripotent phenotype in the
presence of chemical inhibitors of transforming growth
factor-␤, mitogen activated protein kinase kinase, and glycogen synthase kinase 3-␤ all support the notion that signaling
pathway modulation plays an important role in direct cell
lineage reprogramming.37 In this regard, the dramatic difference between the efficiency of fibroblast-to-cardiomyocyte
reprogramming in vitro (1%–5%) and in vivo (10%–30%) by
the overexpression of Gata4, myocyte-specific enhancer factor 2C, and T-box 5 may be the result of the presence of
critical growth factors that are present only in vivo.36,38
However, to examine the findings from Rentschler et al29
in a reprogramming context, a number of key issues need to
be addressed. For example, are the Notch-activated cells truly
reprogrammed epigenetically or do they manifest a conduction system phenotype only under the continuous stimulus of
Notch signaling? A demonstration of the maintenance of
conduction system gene expression and functional phenotype
in the absence of continuous Notch signaling would be
important. Furthermore, the presence of epigenetic silencing
at sarcomeric gene loci and epigenetic activation at ion
channel gene loci would be essential. Also, are the cardiomyocytes that are induced in this study truly mature or are
they still immature and somewhat plastic for conduction cell
development? It has been reported that the cardiac conduction
system in mice has not finished developing at birth, based on
elegant experiments that have demonstrated postnatal requirements for NKX2.519 and T-box 518 in conduction system
maturation. Hence, it is possible that results from the current
study support the role of Notch signaling on lineage commitment and not reprogramming per se. In addition, what are the
barriers to conversion of cardiomyocytes to conduction cells
given that only 2 out of 11 patched cells were found to exhibit
the full conduction cell phenotype? Because multiple other
signals are involved in conduction system development, it is
likely that modulation of these other pathways will be
necessary to improve the efficiency of the reprogramming
process. Also, it would be important to investigate the ability
of Notch signaling to induce a conduction cell phenotype in
adult cardiomyocytes or even cardiac fibroblasts given the
potential utility of this strategy to create replacement cells for
the degenerative conduction system in the elderly.
The reprogramming biology of cardiovascular cells is in
its infancy. Although the pioneering studies from Takahashi
et al.31,32 have shown the power and possibilities of what can
be achieved with transcription factor overexpression, how
much these findings can be adapted to generate mature
cardiomyocytes and conduction cells remains to be seen. We
envision a day in the future when electrophysiologists will
discuss with their patients the reprogramming of their cardiomyocytes instead of their permanent pacemaker. The work of
Rentschler et al29 reported here provides a new dimension to
this arena by showing that modulation of signaling pathways
can lead to direct phenotype conversion of cardiomyocytes
toward conduction cells.
Acknowledgments
This work was supported by National Institutes of Health grants
HL100408 and OD004411 to Dr Wu, and HL109004 and DA026982
to Dr Milan.
Disclosures
None.
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KEY WORDS: Editorials
signal transduction
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conduction
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development genes
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genetics
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Reprogramming the Beat: Kicking It Up a Notch
Sean M. Wu and David J. Milan
Circulation. 2012;126:1009-1011; originally published online July 26, 2012;
doi: 10.1161/CIRCULATIONAHA.112.126482
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