download report - Istituto Pasteur

P a r t i c i p a n t s :
Gabriella Dobrowolny, Emanuele Rizzuto, post-doc fellows;
Michela Aucello, PhD student; Carmine Nicoletti, technician.
C o l l a b o r a t i o n s :
Ce.S.I. Centro Scienze dell’Invecchiamento, IIM, Università degli
Studi G. d’Annunzio, Chieti (Prof. Giorgio Fanò, Prof. Feliciano
Protasi); Dipartimento di Scienze Biomediche, Università di
Padova (Dr. Marco Sandri); EMBL Mouse Biology Program,
Monterotondo, Rome (Dr. Nadia Rosenthal).
Report of activity
The aim of the project is to define the molecular
mechanisms that modulate muscle atrophy, associated
with several pathological conditions. Oxidative
status has profound consequences on the function of
skeletal muscle, where the oxidative level is the highest
in the whole body. The anti-oxidant status of
muscle decreases with age, and is affected in several
other pathological conditions, such as sarcopenia,
chronic fatigue syndrome, liver and kidney diseases,
cancer, muscular dystrophy and amyotrophic lateral
sclerosis (ALS), in which the delicate balance
between oxidant production and antioxidant defense
is severely compromised, leading to muscle wasting.
The combined facts that mice lacking the major
antioxidant enzyme superoxide dismutase 1 (SOD1)
display a dramatic acceleration in age-related loss of
skeletal muscle mass, that elevated levels of reactive
oxygen species (ROS) may contribute to chronic diseases,
and that mutation in SOD1 is associated with
one fifth of familial ALS, have implicated oxidative
stress as a key mechanism underlying the pathogenesis
of aging and neuromuscular diseases. However,
how such an oxidative insult plays a role in the diseases-related
decrease of muscle performance and
mass and whether skeletal muscle is a direct target
of SOD1 mutation remains largely unknown.
In this study we undertook a transgenic approach
Principal investigator: Antonio Musarò
Professor of Biotechnology
Dipartimento di Istologia ed Embriologia Medica
Tel: (+39) 06 49766956; Fax: (+39) 06 4462854
antonio.musaro@uniroma1.it
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Molecular genetics of eukaryotes - AREA 3
Study of the molecular and cellular mechanisms of sarcopenia:
role of mIGF-1 and oxidative stress
to define the direct role of oxidative stress on muscle
homeostasis and function. To this purpose and
to verify whether skeletal muscle is a direct target
of SOD1 mutation we generated transgenic mice
with the mutated isoform of human superoxide
dismutase 1 (SOD1G93A) cDNA driven by skeletal
muscle specific regulatory elements from the rat
myosin light chain (MLC)-1/3 locus. Expression of
the MLC/SOD1G93A transgene in adult mice was
restricted to skeletal muscle, predominated in muscles
enriched in fast fibers and reduced in slow
muscles such as the soleus, where the MLC regulatory
cassette is characteristically expressed at very
low levels. Notably, no expression of human SOD1
protein and transcript were found in heart, brain,
liver, spleen, or spinal cord of transgenic mice.
Muscle-restricted expression of mutant SOD1G93A
was sufficient to induces severe muscle atrophy associated
with significant reduction in muscle strength, sarcomere
disorganization, significant changes of mitochondria
morphology and of their sarcomeric disposition,
and disorganization of the sarcotubular system.
In this study, we also disclosed the potential molecular
mechanisms associated with muscle atrophy
induced by selective accumulation of oxidative stress.
We report that accumulation of ROS serves as signalling
to initiate autophagy, one of the major intracellular
degradation mechanisms that we demonstrated
to be a key determinant for the induction of
muscle atrophy associated with oxidative stress.
In particular, the critical role of autophagy in the
promotion of muscle atrophy was disclosed by
genetic manipulation of LC3 expression, a molecular
marker of autophagy. In vivo electroporation of
siRNA against LC3 gene rescued the atrophic phenotype
in MLC/SOD1G93A mice, suggesting that
autophagy is the dominant pathway that mediates
the atrophic stimulus of oxidative stress and that the
modulation of the autophagy pathway can be a
potential therapeutic mechanism to counteract muscle
atrophy associated with oxidative stress.