Christophe Chanoine

Regular Exercise Prolongs Survival in a Type 2 Spinal Muscular Atrophy Model Mouse

Several studies indicate that physical exercise is likely to be neuroprotective, even in the case of neuromuscular disease. In the present work, we evaluated the efficiency of running-based training on type 2 spinal muscular atrophy (SMA)-like mice. The model used in this study is an SMN (survival motor neuron)-null mouse carrying one copy of a transgene of human SMN2. The running-induced benefits sustained the motor function and the life span of the type 2 SMA-like mice by 57.3%. We showed that the extent of neuronal death is reduced in the lumbar anterior horn of the spinal cord of running-trained mice in comparison with untrained animals. Notably, exercise enhanced motoneuron survival. We showed that the running-mediated neuroprotection is related to a change of the alternative splicing pattern of exon 7 in the SMN2 gene, leading to increased amounts of exon 7-containing transcripts in the spinal cord of trained mice. In addition, analysis at the level of two muscles from the calf, the slow-twitch soleus and the fast-twitch plantaris, showed an overall conserved muscle phenotype in running-trained animals. These data provide the first evidence for the beneficial effect of exercise in SMA and might lead to important therapeutic developments for human SMA patients.

Nfat and miR-25 cooperate to reactivate the transcription factor Hand2 in heart failure

Although aberrant reactivation of embryonic gene programs is intricately linked to pathological heart disease, the transcription factors driving these gene programs remain ill-defined. Here we report that increased calcineurin/Nfat signalling and decreased miR-25 expression integrate to re-express the basic helix-loop-helix (bHLH) transcription factor dHAND (also known as Hand2) in the diseased human and mouse myocardium. In line, mutant mice overexpressing Hand2 in otherwise healthy heart muscle cells developed a phenotype of pathological hypertrophy. Conversely, conditional gene-targeted Hand2 mice demonstrated a marked resistance to pressure-overload-induced hypertrophy, fibrosis, ventricular dysfunction and induction of a fetal gene program. Furthermore, in vivo inhibition of miR-25 by a specific antagomir evoked spontaneous cardiac dysfunction and sensitized the murine myocardium to heart failure in a Hand2-dependent manner. Our results reveal that signalling cascades integrate with microRNAs to induce the expression of the bHLH transcription factor Hand2 in the postnatal mammalian myocardium with impact on embryonic gene programs in heart failure.

Motoneuron survival is promoted by specific exercise in a mouse model of amyotrophic lateral sclerosis

Several studies using transgenic mouse models of familial amyotrophic lateral sclerosis (ALS) have reported a life span increase in exercised animals, as long as animals are submitted to a moderate‐intensity training protocol. However, the neuroprotective potential of exercise is still questionable. To gain further insight into the cellular basis of the exercise‐induced effects in neuroprotection, we compared the efficiency of a swimming‐based training, a high‐frequency and ‐amplitude exercise that preferentially recruits the fast motor units, and of a moderate running‐based training, that preferentially triggers the slow motor units, in an ALS mouse model. Surprisingly, we found that the swimming‐induced benefits sustained the motor function and increased the ALS mouse life span by about 25 days. The magnitude of this beneficial effect is one of the highest among those induced by any therapeutic strategy in this disease. We have shown that, unlike running, swimming significantly delays spinal motoneuron death and, more specifically, the motoneurons of large soma area. Analysis of the muscular phenotype revealed a swimming‐induced relative maintenance of the fast phenotype in fast‐twitch muscles. Furthermore, the swimming programme preserved astrocyte and oligodendrocyte populations in ALS spinal cord. As a whole, these data are highly suggestive of a causal relationship not only linking motoneuron activation and protection, but also motoneuron protection and the maintenance of the motoneuron surrounding environment. Basically, exercise‐induced neuroprotective mechanisms provide an example of the molecular adaptation of activated motoneurons.

Regulation by thyroid hormones of terminal differentiation in the skeletal dorsal muscle. I: Neonate mouse

Changes both in the ATPase myofibrillar profile and in the electrophoretic pattern of myosin isoforms were examined in the mouse dorsal skeletal muscle (longissimus) during postnatal development. In the newborn, only type II C and a few type I fibers were present; differentiation into type II A and II B fibers took place during the 3 weeks following birth. During the same period, a transition from three neonatal isomyosins to four adult isoforms was observed. The two phenomena were related to a marked increase in the serum thyroid hormones levels. Hypothyroidism and hyperthyroidism experiments were performed. Hypothyroidism produced by propylthiouracil treatment of pregnant females and thiourea injections of the litters was shown to induce a complete inhibition of postnatal muscular differentiation. Hyperthyroidism produced by triiodothyronine treatment of the neonate mice significantly accelerated the myosin transition and the switch in the myofibrillar pattern. Our results suggest a primordial role for thyroid hormones in directly regulating the appearance of myosin and fiber adult types and in modulating directly or indirectly the disappearance of the neonatal types.

Expression and Neural Control of Myogenic Regulatory Factor Genes During Regeneration of Mouse Soleus

Given the importance of the myogenic regulatory factors (MRFs) for myoblast differentiation during development, the aims of this work were to clarify the spatial and temporal expression pattern of the four MRF mRNAs during soleus regeneration in mouse after cardiotoxin injury, using in situ hybridization, and to investigate the influence of innervation on the expression of each MRF during a complete degeneration/regeneration process. For this, we performed cardiotoxin injury-induced regeneration experiments on denervated soleus muscle. Myf-5, MyoD, and MRF4 mRNAs were detected in satellite cell-derived myoblasts in the first stages of muscle regeneration analyzed (2–3 days P-I). The Myf-5 transcript level dramatically decreased in young multinucleated myotubes, whereas MyoD and MRF4 transcripts were expressed persistently throughout the regeneration process. Myogenin mRNA was transiently expressed in forming myotubes. These results are discussed with regard to the potential relationships between MyoD and MRF4 in the satellite cell differentiation pathway. Muscle denervation precociously (at 8 days P-I) upregulated both the Myf-5 and the MRF4 mRNA levels, whereas the increase of both MyoD and myogenin mRNA levels was observed later, in the late stages of regeneration (30 days P-I). This significant accumulation of each differentially upregulated MRF during soleus regeneration after denervation suggests that each myogenic factor might have a distinct role in the regulatory control of muscle gene expression. This role is discussed in relation to the expression of the nerve-regulated genes, such as the nAChR subunit gene family.

Exercise-Induced Activation of NMDA Receptor Promotes Motor Unit Development and Survival in a Type 2 Spinal Muscular Atrophy Model Mouse

Spinal muscular atrophy (SMA) is an inborn neuromuscular disorder caused by low levels of survival motor neuron protein, and for which no efficient therapy exists. Here, we show that the slower rate of postnatal motor-unit maturation observed in type 2 SMA-like mice is correlated with the motor neuron death. Physical exercise delays motor neuron death and leads to an increase in the postnatal maturation rate of the motor-units. Furthermore, exercise is capable of specifically enhancing the expression of the gene encoding the major activating subunit of the NMDA receptor in motor neurons, namely the NR2A subunit, which is dramatically downregulated in the spinal cord of type 2 SMA-like mice. Accordingly, inhibiting NMDA-receptor activity abolishes the exercise-induced effects on muscle development, motor neuron protection and life span gain. Thus, restoring NMDA-receptor function could be a promising therapeutic approach to SMA treatment.

Xenopus muscle development: from primary to secondary myogenesis.

Xenopus myogenesis is characterized by specific features, different from those of mammalian and avian systems both at the cellular level and in gene expression patterns. During early embryogenesis, after the initial molecular signals inducing mesoderm, the myogenic determination factors XMyoD and XMyf‐5 are activated in presomitic mesoderm in response to mesoderm‐inducing factors. After these first inductions of the myogenic program, forming muscles in Xenopus can have different destinies, some of these resulting in cell death before adulthood. In particular, it is quite characteristic of this species that, during metamorphosis, the primary myotomal myofibers completely die and are progressively replaced by secondary “adult” multinucleated myofibers. This feature offers the unique opportunity to totally separate the molecular analysis of these two distinct types of myogenesis. The aim of this review is to summarize our knowledge on the cellular and molecular events as well as the epigenetic regulations involved in the construction of Xenopus muscles during development.

In Vivo NMDA Receptor Activation Accelerates Motor Unit Maturation, Protects Spinal Motor Neurons, and Enhances SMN2 Gene Expression in Severe Spinal Muscular Atrophy Mice

Spinal muscular atrophy (SMA), a lethal neurodegenerative disease that occurs in childhood, is caused by the misexpression of the survival of motor neuron (SMN) protein in motor neurons. It is still unclear whether activating motor units in SMA corrects the delay in the postnatal maturation of the motor unit resulting in an enhanced neuroprotection. In the present work, we demonstrate that an adequate NMDA receptor activation in a type 2 SMA mouse model significantly accelerated motor unit postnatal maturation, counteracted apoptosis in the spinal cord, and induced a marked increase of SMN expression resulting from a modification of SMN2 gene transcription pattern. These beneficial effects were dependent on the level of NMDA receptor activation since a treatment with high doses of NMDA led to an acceleration of the motor unit maturation but favored the apoptotic process and decreased SMN expression. In addition, these results suggest that the NMDA-induced acceleration of motor unit postnatal maturation occurred independently of SMN. The NMDA receptor activating treatment strongly extended the life span in two different mouse models of severe SMA. The analysis of the intracellular signaling cascade that lay downstream the activated NMDA receptor revealed an unexpected reactivation of the CaMKII/AKT/CREB (cAMP response element-binding protein) pathway that induced an enhanced SMN expression. Therefore, pharmacological activation of spinal NMDA receptors could constitute a useful strategy for both increasing SMN expression and limiting motor neuron death in SMA spinal cord.

Expression and neural control of follistatin versus myostatin genes during regeneration of mouse soleus

Follistatin and myostatin are two secreted proteins involved in the control of muscle mass during development. These two proteins have opposite effects on muscle growth, as documented by genetic models. The aims of this work were to analyze in mouse, by using in situ hybridization, the spatial and temporal expression patterns of follistatin and myostatin mRNAs during soleus regeneration after cardiotoxin injury, and to investigate the influence of innervation on the accumulation of these two transcripts. Follistatin transcripts could be detected in activated satellite cells as early as the first stages of regeneration and were transiently expressed in forming myotubes. In contrast, myostatin mRNAs accumulated persistently throughout the regeneration process as well as in adult control soleus. Denervation significantly affected both follistatin and myostatin transcript accumulation, but in opposite ways. Muscle denervation persistently reduced the levels of myostatin transcripts as early as the young myotube stage, whereas the levels of follistatin mRNA were strongly increased in the small myotubes in the late stages of regeneration. These results are discussed with regard to the potential functions of both follistatin, as a positive regulator of muscle differentiation, and myostatin, as a negative regulator of skeletal muscle growth. We suggest that the belated up‐regulation of the follistatin mRNA level in the small myotubes of the regenerating soleus as well as the down‐regulation of the myostatin transcript level after denervation contribute to the differentiation process in denervated regenerating muscle. Developmental Dynamics 227:256–265, 2003.

Expression of myogenic regulatory factors during muscle development of Xenopus: Myogenin mRNA accumulation is limited strictly to secondary myogenesis

To clarify the acquisition of the adult muscle pattern in Xenopus laevis, in situ hybridization and reverse transcriptase‐polymerase chain reaction were used to correlate the time course of gene expression for myogenic regulatory factors (Myf‐5, MyoD, and myogenin) with the expression of contractile protein (myosin heavy chain; MHC) genes during hindlimb formation compared with their expression in dorsal body muscles. After the precocious expression of Myf‐5 and MyoD mRNA in limb bud (stage 50), myogenin mRNA strongly accumulated later at paddle stages (stages 52/ 53) concomitantly with the accumulation of both the larval and the adult MHC mRNAs. In dorsal body muscles, as early as stage 52, myogenin transcripts accumulated in a few small, secondary myofibers expressing the adult MHC mRNA that were located along the dorsomedial edge, but they were never detected in the large, primary myofibers of the body expressing the larval MHC mRNA. During metamorphosis, the areas expressing both the adult MHC and the myogenin transcripts gradually expanded from the dorsomedial edge to the ventral side of the dorsal body muscles, accounting for the progression of the secondary “adult” myogenesis described previously (Nishikawa and Hayashi [1994] Dev. Biol. 165:86–94). This work shows that, in Xenopus, the accumulation of myogenin mRNA is restricted to secondary myogenesis, including the formation of new muscles in developing limbs as well as in dorsal muscles during body remodeling. This shows that myogenin is not required for primary myogenesis, and it suggests a crucial role for myogenin in the terminal differentiation program, including myoblast fusion and the activation of adult‐type muscle genes. Dev. Dyn. 1998;213:309–321.