Cell signalling in muscle wasting

Confocal microscopy analysis shows that autophagosome vesiscles (green) are induced by FoxO3 in isolated muscle fibres and are distributed throughout the fibres. Single fibres from the FDB muscle were transfected with LC3-GFP (an autophagosome marker) mitoRed (mitochondria marker) and FoxO3 and examined by confocal microscopy. Co-transfection of skeletal muscles with FoxO3 and LC3-GFP was found to induce a large number of green fluorescent dots in myofibres and concomitantly a reduction in number and size of mitochondria (red). Furthemore some mitochondria are inside autophagosome (yellow stain in merge picture) suggesting that FoxO3 induces reduction in mitochondria via mitophagy.

Confocal microscopy analysis shows that autophagosome vesiscles (green) are induced by FoxO3 in isolated muscle fibres and are distributed throughout the fibres. Single fibres from the FDB muscle were transfected with LC3-GFP (an autophagosome marker) mitoRed (mitochondria marker) and FoxO3 and examined by confocal microscopy. Co-transfection of skeletal muscles with FoxO3 and LC3-GFP was found to induce a large number of green fluorescent dots in myofibres and concomitantly a reduction in number and size of mitochondria (red). Furthemore some mitochondria are inside autophagosome (yellow stain in merge picture) suggesting that FoxO3 induces reduction in mitochondria via mitophagy.
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The mass and functional capacity of skeletal muscle are tightly regulated by contractile activity, nutrient supply and hormones. Contractile activity is necessary for postnatal muscle growth and for the maintenance of muscle mass in adults, and increased work can cause fiber hypertrophy. Conversely, disuse or denervation causes rapid atrophy. Skeletal muscle also serves as the organisms major protein reservoir from which amino acids can be mobilized for gluconeogenesis, new protein synthesis or as an energy store. Consequently, upon food deprivation and in many systemic disease states, including sepsis, cancer, burn injury, diabetes, cardiac and renal failure, there is a generalized muscle wasting, which results primarily from increased breakdown of muscle proteins, although protein synthesis also falls in most of these conditions. In all these systemic catabolic states, the loss of muscle mass involves a common pattern of transcriptional changes, including induction of genes for protein degradation and decreased expression of various genes for growth-related and energy-yielding processes. We have termed this group of co-ordinately regulated genes, "atrogenes". Recent work indicates that the same transcriptional program occurs during atrophy induced by denervation and disuse as occurs in these catabolic states. In the atrophying muscles, the ubiquitin-proteasome system is activated and catalyzes the degradation of the bulk of muscle proteins, especially myofibrillar components. Atrophy is accompanied by a dramatic (8-40 fold) induction of two muscle-specific ubiquitin ligases, atrogin-1/MAFbx and MuRF-1, whose induction occurs prior to the onset of muscle weight loss and is necessary for rapid atrophy. On the other hand, the expression of these ubiquitin ligases and the enhancement of overall protein breakdown are blocked by the IGF-1/insulin/PI3K/AKT signaling pathway (Sandri, 2004), which also activates protein synthesis and net growth of these muscles. The key mediators of this catabolic response during atrophy are the FoxO family of transcription factors, whose activity is suppressed during growth by phosphorylation by AKT, but whose expression and dephosphorylation rises in these catabolic states. Activation of FoxO3 promotes the expression of atrogin-1 and other atrogenes, leading to a dramatic loss of muscle mass. On the other hand, when FoxO3 function is blocked, atrogin-1 expression and the muscle atrophy induced by fasting or glucocorticoids are prevented (Sandri, 2004).

Role of physical activity, PGC1a, mitochondrial biogenesis and energy balance in muscle atrophy

The mechanisms by which contractile activity preserves muscle mass, even in the face of catabolic signals, has long been a mystery. Muscle wasting does not occur similarly in all types of muscle fibers. Upon fasting, exposure to glucocorticoids, sepsis, and cancer cachexia, type II glycolytic muscle fibers show greater atrophy than the type I oxidative fibers. On the other hand, upon unloading or denervation, the fatigue-resistant, slow-contracting, dark muscles show more pronounced atrophy than fast-contracting, glycolytic, pale ones. We recently clarify how exercise can retard muscle wasting, and why fiber types differ in their susceptibility to atrophy (Sandri, 2006). We demonstrate that PGC 1a and FoxO3 plan an opposing role in determining muscle mass. Elevated levels of PGC 1a through transgenic expression reduces the atrophy-promoting effects of FoxO3 and suppresses the associated changes in transcription of key atrogenes. Moreover, we show that PGC 1a expression falls dramatically after denervation and in various other types of muscle wasting, enhancing the FoxO-induced loss of muscle mass.

Regulation of the major proteolytic systems

The transcription factors FoxO are involved in muscle atrophy. In previous studies we found that FoxO3 is the transcription factor which regulates the expression of the muscle-specififc atrophy-related ubiquitin ligases atrogin1 and MuRF1. Very little is known on the transcriptional regulation of autophagy genes. Now we have found that FoxO transcription factors are required for the induction of autophagy in skeletal muscle in vivo. Constitutively active FoxO3 is able to induce autophagosome formation and up-regulation of the autophagy genes in skeletal muscle. In contrast, suppression of FoxO3 activity prevents autophagosome formation induced by starvation. Thus FoxO3 controls independently the two major pathways of protein breakdown in skeletal muscle, the ubiquitin-proteasome and autophagy-lysosome pathway.

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Marco Sandri Venetian Institute of Molecular Medicine Via Orus 2 35129 Padua — Italy

Tel.:  (+39) 049 7923 258 Fax:  (+39) 049 7923 250

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