Diminished overload-induced hypertrophy in aged fast-twitch skeletal muscle is associated with AMPK hyperphosphorylation.

Skeletal muscle mass declines with age, as does the potential for overload-induced fast-twitch skeletal muscle hypertrophy. Because 5'-AMP-activated protein kinase (AMPK) activity is thought to inhibit skeletal muscle protein synthesis and may therefore modulate muscle mass and hypertrophy, the purpose of this investigation was to examine AMPK phosphorylation status (a marker of AMPK activity) and its potential association with the attenuated overload-induced hypertrophy observed in aged skeletal muscle. One-week overload of fast-twitch plantaris and slow-twitch soleus muscles was achieved in young adult (8 mo; n = 7) and old (30 mo; n = 7) Fischer344 x Brown Norway male rats via unilateral gastrocnemius ablation. Significant (P < or = 0.05) age-related atrophy (as measured by total protein content) was noted in plantaris and soleus control (sham-operated) muscles. In fast-twitch plantaris muscles, percent hypertrophy with overload was significantly attenuated with age, whereas AMPK phosphorylation status as determined by Western blotting [phospho-AMPK (Thr172)/total AMPK] was significantly elevated with age (regardless of loading status). There was also a main effect of loading on AMPK phosphorylation status in plantaris muscles (overload > control). Moreover, a strong and significant negative correlation (r = -0.82) was observed between AMPK phosphorylation status and percent hypertrophy in the overloaded plantaris muscles of all animals. In contrast to the plantaris, overload-induced hypertrophy of the slow-twitch soleus muscle was similar between ages, and AMPK phosphorylation in this muscle was also unaffected by age or overload. These data support the possibility that an age-related elevation in AMPK phosphorylation may partly contribute to the attenuated hypertrophic response observed with age in overloaded fast-twitch plantaris muscle.     

Aging influences cellular and molecular responses of apoptosis to skeletal muscle unloading

The influence of aging on skeletal myocyte apoptosis is not well understood. In this study we examined apoptosis and apoptotic regulatory factor responses to muscle atrophy induced via limb unloading following loading-induced hypertrophy. Muscle hypertrophy was induced by attaching a weight to one wing of young and aged Japanese quails for 14 days. Removing the weight for 7 or 14 days after the initial 14 days of loading induced muscle atrophy. The contralateral wing served as the intra-animal control. A time-released bromodeoxyuridine (BrdU) pellet was implanted subcutaneously with wing weighting to identify activated satellite cells/muscle precursor cells throughout the experimental period. Bcl-2 mRNA and protein levels decreased after 7 days of unloading, but they were unchanged after 14 days of unloading in young muscles. Bcl-2 protein level but not mRNA level decreased after 7 days of unloading in muscles of aged birds. Seven days of unloading increased the mRNA level of Bax in muscles from both young and aged birds. Fourteen days of unloading increased mRNA and protein levels of Bcl-2, decreased protein levels of Bax, and decreased nuclear apoptosis-inducing factor (AIF) protein level in muscles of aged birds. BrdU-positive nuclei were found in all unloaded muscles from both age groups, but the number of BrdU-positive nuclei relative to the total nuclei decreased after 14 days of unloading compared with 7 days of unloading. The TdT-mediated dUTP nick end labeling (TUNEL) index was higher after 7 days of unloading in both young and aged muscles and after 14 days of unloading in aged muscles. Immunofluorescent staining revealed that almost all of the TUNEL-positive nuclei were also BrdU immunopositive, suggesting that activated satellite cell nuclei (both fused and nonfused) underwent nuclear apoptosis during unloading. There were significant correlations among levels of Bcl-2, Bax, and AIF and TUNEL index. Our data are consistent with the hypothesis that apoptosis regulates, at least in part, unloading-induced muscle atrophy and loss of activated satellite cell nuclei in previously loaded muscles. Moreover, these data suggest that aging influences the apoptotic responses to prolonged unloading following hypertrophy in skeletal myocytes. satellite cells; Bcl-2 protein family     

Vasodilatory mechanisms in contracting skeletal muscle.

Skeletal muscle blood flow is closely coupled to metabolic demand, and its regulation is believed to be mainly the result of the interplay of neural vasoconstrictor activity and locally derived vasoactive substances. Muscle blood flow is increased within the first second after a single contraction and stabilizes within approximately 30 s during dynamic exercise under normal conditions. Vasodilator substances may be released from contracting skeletal muscle, vascular endothelium, or red blood cells. The importance of specific vasodilators is likely to vary over the time course of flow, from the initial rapid rise to the sustained elevation during steady-state exercise. Exercise hyperemia is therefore thought to be the result of an integrated response of more than one vasodilator mechanism. To date, the identity of vasoactive substances involved in the regulation of exercise hyperemia remains uncertain. Numerous vasodilators such as adenosine, ATP, potassium, hypoxia, hydrogen ion, nitric oxide, prostanoids, and endothelium-derived hyperpolarizing factor have been proposed to be of importance; however, there is little support for any single vasodilator being essential for exercise hyperemia. Because elevated blood flow cannot be explained by the failure of any single vasodilator, a consensus is beginning to emerge for redundancy among vasodilators, where one vasoactive compound may take over when the formation of another is compromised. Conducted vasodilation or flow-mediated vasodilation may explain dilation in vessels (i.e., feed arteries) not directly exposed to vasodilator substances in the interstitium. Future investigations should focus on identifying novel vasodilators and the interaction between vasodilators by simultaneous inhibition of multiple vasodilator pathways.     

Mechanotransduction pathways in skeletal muscle hypertrophy

In the last decade, molecular biology has contributed to define some of the cellular events that trigger skeletal muscle hypertrophy. Recent evidence shows that insulin like growth factor 1/phosphatidyl inositol 3-kinase/protein kinase B (IGF-1/PI3K/Akt) signaling is not the main pathway towards load-induced skeletal muscle hypertrophy. During load-induced skeletal muscle hypertrophy process, activation of mTORC1 does not require classical growth factor signaling. One potential mechanism that would activate mTORC1 is increased synthesis of phosphatidic acid (PA). Despite the huge progress in this field, it is still early to affirm which molecular event induces hypertrophy in response to mechanical overload. Until now, it seems that mTORC1 is the key regulator of load-induced skeletal muscle hypertrophy. On the other hand, how mTORC1 is activated by PA is unclear, and therefore these mechanisms have to be determined in the following years. The understanding of these molecular events may result in promising therapies for the treatment of muscle-wasting diseases. For now, the best approach is a good regime of resistance exercise training. The objective of this point-of-view paper is to highlight mechanotransduction events, with focus on the mechanisms of mTORC1 and PA activation, and the role of IGF-1 on hypertrophy process.     

Age effects on myosin subunit and biochemical alterations with skeletal muscle hypertrophy.

The purpose of this study was to determine whether skeletal muscle mass, myofibrillar adenosinetriphosphatase activity, and the expression of myosin heavy (MHC) and light chain subunits are differentially affected in juvenile (4 wk) and young adult (12 wk) rats by a hypertrophic growth stimulus. Hypertrophy of the plantaris or soleus was studied 4 wk after ablation of either two [gastrocnemius (GTN) and soleus or plantaris] or one (GTN) synergistic muscle(s). There was no difference in the relative magnitude of hypertrophy because of age. Plantaris myofibrillar adenosinetriphosphatase activity was decreased 21 and 12% in juvenile and adult rats, respectively, as a result of ablation of both the GTN and soleus. Slow myosin light chain isoforms (1s and 2s) were expressed to a greater extent in hypertrophied plantaris muscles of both ages, but the increase in 1s was greater in juvenile rats. The relative expression of slow beta-MHC in hypertrophied plantaris muscles increased by 470 and 350%, whereas MHC IIb decreased by 70 and 33% in juvenile and adult rats, respectively. The relative expression of MHC IIa increased (56%) in the plantaris after ablation in juvenile rats only. These shifts in myosin subunit expression and the increases in mass were generally about one-half the magnitude when only the GTN was removed. There were no detectable myosin shifts in hypertrophied soleus muscles. Although the extent of muscle hypertrophy is similar, the shifts in myosin subunits were greater in juvenile than in young adult rats.     

Heat stress inhibits skeletal muscle hypertrophy

Heat shock proteins (Hsps) are molecular chaperones that aid in protein synthesis and trafficking and have been shown to protect cells/tissues from various protein damaging stressors. To determine the extent to which a single heat stress and the concurrent accumulation of Hsps influences the early events of skeletal muscle hypertrophy, Sprague-Dawley rats were heat stressed (42 degrees C, 15 minutes) 24 hours prior to overloading 1 plantaris muscle by surgical removal of the gastrocnemius muscle. The contralateral plantaris muscles served as controls. Heat-stressed and/or overloaded plantaris muscles were assessed for muscle mass, total muscle protein, muscle protein concentration, Type I myosin heavy chain (Type I MHC) content, as well as Hsp72 and Hsp25 content over the course of 7 days following removal of the gastrocnemius muscle. As expected, in non-heat-stressed animals, muscle mass, total muscle protein and MHC I content were significantly increased (P < 0.05) following overload. In addition, Hsp25 and Hsp72 increased significantly after 2 and 3 days of overload, respectively. A prior heat stress-elevated Hsp25 content to levels similar to those measured following overload alone, but heat stress-induced Hsp72 content was increased significantly greater than was elicited by overload alone. Moreover, overloaded muscles from animals that experienced a prior heat stress showed a lower muscle mass increase at 5 and 7 days; a reduced total muscle protein elevation at 3, 5, and 7 days; reduced protein concentration; and a diminished Type I MHC content accumulation at 3, 5, and 7 days relative to nonheat-stressed animals. These data suggest that a prior heat stress and/or the consequent accumulation of Hsps may inhibit increases in muscle mass, total muscle protein content, and Type I MHC in muscles undergoing hypertrophy.     

Mechanotransduction pathways in skeletal muscle hypertrophy

In the last decade, molecular biology has contributed to define some of the cellular events that trigger skeletal muscle hypertrophy. Recent evidence shows that insulin like growth factor 1/phosphatidyl inositol 3-kinase/protein kinase B (IGF-1/PI3K/Akt) signaling is not the main pathway towards load-induced skeletal muscle hypertrophy. During load-induced skeletal muscle hypertrophy process, activation of mTORC1 does not require classical growth factor signaling. One potential mechanism that would activate mTORC1 is increased synthesis of phosphatidic acid (PA). Despite the huge progress in this field, it is still early to affirm which molecular event induces hypertrophy in response to mechanical overload. Until now, it seems that mTORC1 is the key regulator of load-induced skeletal muscle hypertrophy. On the other hand, how mTORC1 is activated by PA is unclear, and therefore these mechanisms have to be determined in the following years. The understanding of these molecular events may result in promising therapies for the treatment of muscle-wasting diseases. For now, the best approach is a good regime of resistance exercise training. The objective of this point-of-view paper is to highlight mechanotransduction events, with focus on the mechanisms of mTORC1 and PA activation, and the role of IGF-1 on hypertrophy process.     

NAD+ centric mechanisms and molecular determinants of skeletal muscle disease and aging

Molecular and Cellular Biochemistry - The nicotinamide adenine dinucleotide (NAD+) is an essential redox cofactor, involved in various physiological and molecular processes, including energy...     

Polyamines, androgens, and skeletal muscle hypertrophy

The naturally occurring polyamines, spermidine, spermine, and their precursor putrescine, play indispensible roles in both prokaryotic and eukaryotic cells, from basic DNA synthesis to regulation of cell proliferation and differentiation. The rate-limiting polyamine biosynthetic enzymes, ornithine decarboxylase (ODC) and S-adenosylmethionine decarboxylase, are essential for mammalian development, with knockout of the genes encoding these enzymes, Odc1 and Amd1, causing early embryonic lethality in mice. In muscle, the involvement of polyamines in muscle hypertrophy is suggested by the concomitant increase in cardiac and skeletal muscle mass and polyamine levels in response to anabolic agents including β-agonists. In addition to β-agonists, androgens, which increase skeletal mass and strength, have also been shown to stimulate polyamine accumulation in a number of tissues. In muscle, androgens act via the androgen receptor to regulate expression of polyamine biosynthetic enzyme genes, including Odc1 and Amd1, which may be one mechanism via which androgens promote muscle growth. This review outlines the role of polyamines in proliferation and hypertrophy, and explores their possible actions in mediating the anabolic actions of androgens in muscle.     

Proteomic analysis of bovine skeletal muscle hypertrophy

Myostatin plays a major role in muscle growth and development and animals with disruption of this gene display marked increases in muscle mass. Little is known about muscle physiological adaptations in relation to this muscle hypertrophy. To provide a more comprehensive view, we analyzed bovine muscles from control, heterozygote and homozygote young Belgian blue bulls for myostatin deletion, which results in a normal level of inactive myostatin. Heterozygote and homozygote animals were characterized by a higher proportion of fast-twitch glycolytic fibers in Semitendinosus muscle. Differential proteomic analysis of this muscle was performed using two-dimensional gel electrophoresis followed by mass spectrometry. Thirteen proteins, corresponding to 28 protein spots, were significantly altered in response to the myostatin deletion. The observed changes in protein expression are consistent with an increased fast muscle phenotype, suggesting that myostatin negatively controls mainly fast-twitch glycolytic fiber number. Finally, we demonstrated that differential mRNA splicing of fast troponin T is altered by the loss of myostatin function. The structure of mutually exclusive exon 16 appears predominantly expressed in muscles from heterozygote and homozygote animals. This suggests a role for exon 16 of fast troponin T in the physiological adaptation of the fast muscle phenotype.     

Ionic and molecular mechanisms of beta-amyloid-induced depolarization of the mouse skeletal muscle fibres

Excess production and accumulation of beta-amyloid peptide (betaAP) are central for pathogenesis of Alzheimer's disease. Numerous studies showed that betaAP possessed wide range of toxic effects on neurons, however the mechanism of betaAP influence on another types of excitable cells, for example, skeletal muscle fibres, is unknown. In electrophysiological experiments on the mouse diaphragm, we found for the first time that betaAP (25-35 fragment, 10-6 M) disturbs the processes of the resting membrane potential generation in muscle fibres, leading to depolarization by two mechanisms: 1) inhibition of Na+,K(+)-ATPase, which leads to loss of impact of this pump to the resting membrane potential; 2) increase of membrane cationic permeability due to formation of "amyloid" channels blocked with Zn2+ ions. Our results significantly broaden current understanding of mechanisms of motor disturbances and skeletal muscle pathology in Alzheimer's disease, inclusion body myositis and other betaAP-related disorders.     

Stem cells to tissue: molecular,cellular and anatomical heterogeneity in skeletal muscle

Multiple waves of muscle precursors are released from skeletal muscle progenitor cells throughout developmental life, and this process is initiated in precise locations in the embryo. Skeletal muscle diversifies not only after the acquisition of muscle identity, but curiously heterogeneity is observed even in the stem cell population. Recent studies on cell lineage, cell fusion and the nature of post-natal satellite cells have expanded on our fundamental knowledge of the formation of this tissue, and how this tissue is replenished by resident and circulating regenerative stem cells during adult life.     

Intensity-controlled treadmill running in mice: cardiac and skeletal muscle hypertrophy.

Whereas novel pathways of pathological heart enlargement have been unveiled by thoracic aorta constriction in genetically modified mice, the molecular mechanisms of adaptive cardiac hypertrophy remain virtually unexplored and call for an effective and well-characterized model of physiological mechanical loading. Experimental procedures of maximal oxygen consumption (VO(2 max)) and intensity-controlled treadmill running were established in 40 female and 36 male C57BL/6J mice. An inclination-dependent VO(2 max) with 0.98 test-retest correlation was found at 25 degrees treadmill grade. Running for 2 h/day, 5 days/wk, in intervals of 8 min at 85-90% of VO(2 max) and 2 min at 50% (adjusted to weekly VO(2 max) testing) increased VO(2 max) to a plateau 49% above sedentary females and 29% in males. Running economy improved in both sexes, and echocardiography indicated significantly increased left ventricle posterior wall thickness. Ventricular weights increased by 19-29 and 12-17% in females and males, respectively, whereas cardiomyocyte dimensions increased by 20-32, and 17-23% in females and males, respectively; skeletal muscle mass increased by 12-18%. Thus the model mimics human responses to exercise and can be used in future studies of molecular mechanisms underlying these adaptations.     

Signalling pathways that mediate skeletal muscle hypertrophy and atrophy

Atrophy of skeletal muscle is a serious consequence of numerous diseases, including cancer and AIDS. Successful treatments for skeletal muscle atrophy could either block protein degradation pathways activated during atrophy or stimulate protein synthesis pathways induced during skeletal muscle hypertrophy. This perspective will focus on the signalling pathways that control skeletal muscle atrophy and hypertrophy, including the recently identified ubiquitin ligases muscle RING finger 1 (MuRF1) and muscle atrophy F-box (MAFbx), as a basis to develop targets for pharmacologic intervention in muscle disease.