n engl j med 359;13 www.nejm.

org september 25, 2008

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clinical implications of basic research
The new engl and journal o f medicine
Duchennes muscular dystrophy is a devastating,
progressive, X-linked muscle-wasting disease. It is
the most common form of muscular dystrophy,
affecting 1 in 3500 boys. With an onset in early
childhood, the disease progresses to final stages
that are characterized by cardiorespiratory fail-
ure and death, which usually occurs in the teen-
age years or early 20s. The disease affects stri-
ated muscles of the limbs, diaphragm, and heart
and is associated with a progressive loss of mus-
cle mass, leading to impaired ambulation and pa-
ralysis. Skeletal myofibers degenerate and undergo
necrosis; muscles progressively accumulate calci-
um deposits and are replaced by connective tissue
and fat.
Twenty years ago, mutations in the dystro-
phin gene were found to be responsible for the
disease. Such mutations lead to the production
of defective structural proteins and the loss of
muscle-membrane integrity. No effective therapy
is available, although therapeutic interventions
that include pharmacologic agents and genetic
alterations to replace the missing dystrophin by
exon skipping or viral gene delivery are in clini-
cal trials.
An attractive alternative is a cell-based therapy,
and a study recently described by Cerletti et al.
1

demonstrates an interesting strategy. Muscle is
composed of multinucleated cells, called myofi-
bers, to which myogenic precursors fuse. If these
precursors harbor a normal, healthy gene, they
could provide the missing dystrophin protein
and rescue the fiber by taking advantage of the
normal biology of muscle-tissue formation. Sev-
eral cell-based strategies have been tested with
variable success.
Myoblasts are mononucleated myogenic pre-
cursors capable of extensive proliferation and
fusion to form multinucleated fibers in tissue
culture. The discovery of methods for their puri-
fication has led to several clinical trials. Although
the injected human cells fused with resident
muscle fibers and synthesized the appropriate
gene products in patients muscles, the cells and
their products remained extremely localized.
2
Given the limitations inherent in this native
muscle-cell source, researchers have turned to
other cell sources and modes of delivery. Bone
marrow stromal cells are among the cell types
that have been isolated on the basis of their ad-
hesive properties. These cells can be expanded
extensively in tissue culture, providing an ample
source of cells capable of contributing to mus-
cles of the mdx mouse (a model of Duchennes
muscular dystrophy) after intramuscular injection.
In another approach, cells with an exceedingly
active efflux pump can be isolated from blood
or muscle by flow cytometry. An attractive fea-
ture of these cells is that they can be delivered
intravenously because they are capable of extrav-
asating from the vasculature and engrafting into
myofibers after tail-vein injection. A limitation
of these cells is that they cannot be grown in
tissue culture.
3
Efficient transplantation has been
observed with the use of another cell type, the
blood vesselassociated mesangioblast, which can
be expanded extensively in culture and effectively
delivered by femoral-artery catherization.
4
Me-
sangioblasts are isolated from the outgrowth of
small-vesselcontaining tissue fragments from
muscle-biopsy specimens. Such cells are advanta-
geous because they contribute to muscles through-
out the body and have been shown to restore dys-
trophin and strength to muscles of dystrophic
dogs (which best recapitulate the human disease).
In parallel, researchers have become increas-
ingly interested in the satellite cell, which is in-
trinsic to skeletal muscle and is the natural cell
source for muscle regeneration. More than four
decades ago, Alexander Mauro first identified
Cell Therapies for Muscular Dystrophy
Helen M. Blau, Ph.D.
Copyright 2008 Massachusetts Medical Society. All rights reserved.
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The new engl and journal o f medicine
n engl j med 359;13 www.nejm.org september 25, 2008
1404
Copyright 2008 Massachusetts Medical Society. All rights reserved.
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n engl j med 359;13 www.nejm.org september 25, 2008
1405
clinical implications of basic research
satellite cells using electron microscopy as mono-
nucleated cells ensheathed in their own mem-
brane compartments along the length of the
muscle fiber. Recently, isolation of satellite cells
has been achieved with the use of diverse proto-
cols. Remarkably, if satellite cells are transplant-
ed as soon as they are isolated, they are capable
of extensive dissemination throughout the mus-
cle into which they are injected, a property that
is lost as soon as the cells are exposed to culture
conditions, whereupon they give rise to more
specialized myoblasts.
5
The study by Cerletti et al. represents an ad-
vance in several respects. The authors describe a
subpopulation of freshly isolated satellite cells ob-
tained with the use of a combination of molecu-
lar markers, of which CXCR4 (a receptor that
responds to signals emitted by muscle damage)
may be key. Genetically marked skeletal muscle
progenitors, which express green fluorescent pro-
tein (GFP), can be recovered from the muscles to
which they contribute (Fig. 1). Using a dual mark-
er system, the investigators showed that the cells
contributed not only to preexisting muscle fibers
but also to newly formed fibers. Most remark-
ably, this subpopulation of satellite cells contrib-
uted substantively to the injected muscles (the
authors observed that 90% of muscle fibers were
composed of cells labeled with GFP) and led to
an unprecedented increase in muscle force in the
mdx mouse.
Are CXCR4+ skeletal-muscle progenitors or
other satellite cells the long-sought stem cells of
adult muscle? To be so, they need to fulfill stan-
dard criteria. A single cell must be transplanted
and shown both to reproduce itself and to give
rise to more specialized progeny. A means of
propagating the cells in culture without a loss
of their regenerative properties is necessary; with-
out expansion, the low cell numbers would limit
therapeutic applications of either genetically cor-
rected autologous cells or heterologous cells used
in conjunction with immunosuppression. More-
over, if systemic delivery were possible, it would
not be necessary to inject each muscle individu-
ally. Finally, the human counterpart of the spe-
cialized satellite cell remains to be identified and
shown to have similar properties. Nonetheless,
the findings of Cerletti et al. are notable and will
no doubt fuel interest in a cell-based therapy for
Duchennes muscular dystrophy.
No potential conflict of interest relevant to this article was re-
ported.
From Stanford University School of Medicine, Stanford, CA.
Cerletti M, Jurga S, Witczak C, et al. Highly efficient, func- 1.
tional engraftment of skeletal muscle stem cells in dystrophic
muscles. Cell 2008;134:37-47.
Gussoni E, Pavlath GK, Lanctot AM, et al. Normal dystrophy 2.
transcripts detected in Duchenne muscular dystrophy patients
after myoblast transplantation. Nature 1992;356:435-8.
Gussoni E, Soneoka Y, Strickland CD, et al. Dystrophin ex- 3.
pression in the mdx mouse restored by stem cell transplanta-
tion. Nature 1999;401:390-4.
Sampaolesi M, Blot S, DAntona G, et al. Mesoangioblast 4.
stem cells ameliorate muscle function in dystrophic dogs. Nature
2006;444:574-9.
Montarras D, Morgan J, Collins C, et al. Direct isolation of 5.
satellite cells for skeletal muscle regeneration. Science 2005;
309:2064-7.
Copyright 2008 Massachusetts Medical Society.
Figure 1 (facing page). Repair of Muscle Function
in a Mouse Model.
Cerletti et al. isolated mouse skeletal muscle progeni-
tors (SMPs) from mice that are transgenic for the ex-
pression of green fluorescent protein (GFP
+
) from
skeletal muscles and enriched these cells by flow cy-
tometry using antibodies to cell-surface markers. They
observed that after injection of these purified cells into
muscles damaged by toxins or genetically deficient
(mdx), the cells contributed to more than 95% of mus-
cle fibers and restored dystrophin expression and func-
tion, as shown by increased normalized force (the ratio
of maximal force to the area of the muscle) exerted by
excised soleus muscles (lower panels). (Elements for
the figure courtesy of Stephane Corbel, Ph.D., Stanford
University, Stanford, CA.)
Copyright 2008 Massachusetts Medical Society. All rights reserved.
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