Molecular Exercise Physiology Skeletal Muscle Hypertrophy Presentation 8 Henning Wackerhage

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Molecular Exercise PhysiologySkeletal Muscle
HypertrophyPresentation 8Henning Wackerhage
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Learning outcomes

• At the end of this presentation you should be
  able to
• Explain the effects of myostatin knockout on
  muscle growth.
• Explain how myostatin signalling may inhibit
  muscle growth

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TGF-b signalling
The TGFb superfamily is a large group of proteins
that regulate growth, differentiation and
apoptosis. One subclass within the TGFb
superfamily are growth and differentiation
factors (GDFs). Myostatin is a GDF whose knockout
in mice resulted in skeletal muscle hypertrophy
and hyperplasia (McPherron et al., 1997). The
muscles of myostatin knockout mice are shown on
the next slide.
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Myostatin knockout mice
Wildtype
Myostatin knockout
McPherron et al. (1997) knocked out growth and
development factor 8 (gdf-8) which was later
termed myostatin. The absence of myostatin leads
to muscle hypertrophy (e) compared to the
wildtype (d).
Figure from Lee et al. (1999)
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Myostatin also affects fibre type
Myostatin knockout (MSTN-/-) also leads to a
reduction in type I fibres when compared to the
wildtype Girgenrath et al. (2005).
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Myostatin

• Discovery of myostatin (GDF-8)
• Myostatin is almost only expressed in skeletal
  muscle.
• Myostatin could first be detected at day 9.5
  post-coitum.
• Myostatin knock-out mice muscles weigh 2-3 times
  more than those of wild type animals.
• Hyperplasia (more fibres) and hypertrophy (larger
  fibres) more DNA.
• (McPherron et al. 1997)

Myostatin is a negative growth factor. If you
have lots of it, youre small. If you dont have
it, youre musculous.
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Natural myostatin mutations occur
Natural myostatin mutations can occur as well
McPherron et al. (1997) found in Belgian Blue and
Piedmontese cattle a natural myostatin mutation
resulting in non-functional myostatin and
increased muscle size. An article reporting a
toddler with a natural myostatin mutation is on
the website.
Figure A fullblood Belgian Blue bull showing the
double muscling phenotype (McPherron et al.
1997).
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Natural myostatin mutation in cattle
Piedmontese G?A mutation
Belgian blue deletion
Myostatin protein structure and effect of mutation
Myostatin mutations (mut) in Belgian Blue and
Piedmontese cattle compared with wild-type (wt)
Holstein cattle. These mutations lead to a large
increase in muscle mass, known as double-muscling
(McPherron et al. 1997)
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Flex Wheeler a mutant?
Flex Wheeler has a website where someone states
that Flex has a special genotype linked to his
muscle grwoth Flex was one of only nine extreme
responders that had the very rare "myostatin
mutation." Myostatin is the gene that "limits
muscle growth." Specifically, Flex had the rarest
form of myostatin mutation at the "exon 2"
position on the gene. This simply means Flex has
a much larger number of muscle fibers compared to
the other subjects or the normal population.
Flex Wheeler Its all natural!?
Task Search for scientific literature on this
SNP/mutation. Does it exist?
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Flex Wheeler a mutant hang on
The gentleman who has written the letter for Flex
is Victor Conte. This is what the BBC says
(23.10.2004) Victor Conte is founder and
president of Bay Area Laboratory Co-operative
(Balco), the San Francisco-based company which
the United States Anti-Doping Agency says
developed the banned steroid THG
(tetrahydrogestrinone). So, Flex is all natural?
Victor Conte
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How is myostatin expression regulated?
Myostatin expression is regulated via DNA binding
elements for glucocorticoid (GC), androgen (Tes),
thyroid hormone (Thy), myogenic regulatory
factors (MRFs), MEF2, PPARg (Ma et al., 2001).
Only the glucocorticoid binding site has been
experimentally verified (Ma et al., 2003).
Myostatin
Resistance training
Other regulators
Tes
GC
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Does myostatin regulate the response to exercise?
If myostatin has such a dramatic effect on muscle
mass, is it involved in mediating hypertrophy in
response to resistance training? The story is
not clear at the moment Some reports suggest
that myostatin mRNA or circulating myostatin does
decrease after resistance training (Roth et al.,
2003Zambon et al., 2003Walker et al., 2004)
whereas others suggest it does not (Willoughby,
2004). The jury is still out and a
high-resolution time course needs to be obtained
after human resistance training in order to see
whether there is a regulatory myostatin
expression change in response to resistance
training. Some data are shown on the following
slide.
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Myostatin response to resistance training
Figure. Myostatin expression levels (arbitrary
units) for before ST (n 15) versus after ST (n
15) for each individual. , males , females.
A significant decrease in myostatin expression
was observed in response to ST, P lt 0.05, with no
group differences. (Roth et al. 2003)
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How does myostatin inhibit growth?
Myostatin is like IGF-1 a protein that is
secreted by muscle cells and myostatin dimers
bind so-called activin receptors. These receptors
then phosphorylate and activate the transcription
factor Smad2/3. A simplified version of the
events is shown below.
Myostatin dimer
Activin type I and II receptors
P
SMAD2,3
P
Growth inhibition
SMAD2,3
P
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Myostatin inhibitors (or not?)
Obviously, bodybuilders are very keen to inhibit
myostatin in order to promote muscle growth
further. Champion Nutrition already managed to
develop a myostatin inhibitor (or not?) After
extensive research, Champion Nutrition found a
scientist that had isolated a naturally occurring
fraction of a sea vegetable. It binds strongly to
myostatin, thereby deactivating it. This
discovery made it possible for Champion to begin
work on the first product ever specifically
designed to bind myostatin in humans. See
www.bodybuilding.com/store/cn/myostim.html
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Myostatin inhibitors
Researchers have identified natural myostatin
inhibitors that regulate the receptor binding of
myostatin in the organism. These binding-proteins
form so-called myostatin heterodimers. Inhibitor
proteins are myostatin propeptide, follistatin
and GASP-1.
Myostatin-gasp1 or myostatin-follistatin
heterodimers can not bind receptor
Myostatin homodimer can bind receptor
Activin type I and II receptors
P
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Myostatin inhibition can improve muscular
dystrophy
Bogdanovich et al. (2003) treated mdx mice (model
for muscular dystrophy) with a monoclonal
antibody against myostatin which inhibited
myostatin. Control mdx mice (a) had significantly
greater pathological changes (arrowheads) in the
diaphragm than treated mice (b), as evidenced by
a lack of cellular infiltration and fibrosis.
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Myostatin is an ideal treatment target
Myostatin is probably a very good target for
improving muscle mass in various conditions for
the following reasons a) Myostatin is secreted
and thus treatments do not need to cross
membranes b) Myostatin needs to be inhibited
which usually is easier than activation c)
Natural inhibitors (follistatin, gasp-1) exist
which can be copied d) Myostatin expression is
largely limited to muscle and thus side effects
should be limited. A myostatin-related therapy
via antibodies, peptides or other drugs seems a
realistic prospect.
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Summary
Myostatin is a muscle-specific muscle growth
inhibitor whose expression is probably controlled
by glucocorticoids and anabolic steroids among
other. Myostatin binds activin receptors as a
dimer. Myostatin binding to specific inhibitory
proteins (follistatin, gasp-1) can prevent
myostatin binding to its receptor. Myostatin
binding to the receptor activates the kinase bit
of the receptor which causes phosphorylation of
Smad2/3. Smad2/3 transport to the nucleus and DNA
binding can be modulated by inhibitory and
co-Smads. Smad2/3 probably regulates genes that
inhibit muscle growth. The events mentioned
above are shown on the next slide.
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Summary
Myostatin-gasp1 or myostatin-follistatin
heterodimer
Myostatin homodimer
Activin type I and II receptors
P
Myostatin modification
I-SMAD
Androgens, Glucocorticoids, thyroid
hormones, MRFs, MEF2
SMAD2,3
P
Co-SMAD
SMAD2,3
P
Growth inhibition
Co-SMAD
Enhancers and silencers
Nucleus
Skeletal muscle fibre
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The End