Glucose Restriction: Longevity SIRTainly,but without Building Muscle?

The two metabolic sensors AMPK and SIRT1 take center stage as Fulco et al. reveal, in this issue of Developmental Cell, the signaling mechanism by which low glucose prevents the correct development of the myogenic program. These observations may hold some therapeutic promise against muscle wasting.     

Five Alternative Myosin Converter Domains Influence Muscle Power,Stretch Activation,and Kinetics

Muscles have evolved to power a wide variety of movements. A protein component critical to varying power generation is the myosin isoform present in the muscle. However, how functional variation in muscle arises from myosin structure is not well understood. We studied the influence of the converter, a myosin structural region at the junction of the lever arm and catalytic domain, using Drosophila because its single myosin heavy chain gene expresses five alternative converter versions (11a–e). We created five transgenic fly lines, each forced to express one of the converter versions in their indirect flight muscle (IFM) fibers. Electron microscopy showed that the converter exchanges did not alter muscle ultrastructure. The four lines expressing converter versions (11b–e) other than the native IFM 11a converter displayed decreased flight ability. IFM fibers expressing converters normally found in the adult stage muscles generated up to 2.8-fold more power and displayed up to 2.2-fold faster muscle kinetics than fibers with converters found in the embryonic and larval stage muscles. Small changes to stretch-activated force generation only played a minor role in altering power output of IFM. Muscle apparent rate constants, derived from sinusoidal analysis of the chimeric converter fibers, showed a strong positive correlation between optimal muscle oscillation frequency and myosin attachment kinetics to actin, and an inverse correlation with detachment related cross-bridge kinetics. This suggests the myosin converter alters at least two rate constants of the cross-bridge cycle with changes to attachment and power stroke related kinetics having the most influence on setting muscle oscillatory power kinetics.     

Nonlinear Actomyosin Elasticity in Muscle?

Cyclic interactions between myosin II motor domains and actin filaments that are powered by turnover of ATP underlie muscle contraction and have key roles in motility of nonmuscle cells. The elastic characteristics of actin-myosin cross-bridges are central in the force-generating process, and disturbances in these properties may lead to disease. Although the prevailing paradigm is that the cross-bridge elasticity is linear (Hookean), recent single-molecule studies suggest otherwise. Despite convincing evidence for substantial nonlinearity of the cross-bridge elasticity in the single-molecule work, this finding has had limited influence on muscle physiology and physiology of other ordered cellular actin-myosin ensembles. Here, we use a biophysical modeling approach to close the gap between single molecules and physiology. The model is used for analysis of available experimental results in the light of possible nonlinearity of the cross-bridge elasticity. We consider results obtained both under rigor conditions (in the absence of ATP) and during active muscle contraction. Our results suggest that a wide range of experimental findings from mechanical experiments on muscle cells are consistent with nonlinear actin-myosin elasticity similar to that previously found in single molecules. Indeed, the introduction of nonlinear cross-bridge elasticity into the model improves the reproduction of key experimental results and eliminates the need for force dependence of the ATP-induced detachment rate, consistent with observations in other single-molecule studies. The findings have significant implications for the understanding of key features of actin-myosin-based production of force and motion in living cells, particularly in muscle, and for the interpretation of experimental results that rely on stiffness measurements on cells or myofibrils.     

Diurnal Variations of Plasma Homocysteine,Total Antioxidant Status,and Biological Markers of Muscle Injury During Repeated Sprint: Effect on Performance and Muscle Fatigue—A Pilot Study

The aim of this study was (i) to evaluate whether homocysteine (Hcy), total antioxidant status (TAS), and biological markers of muscle injury would be affected by time of day (TOD) in football players and (ii) to establish a relationship between diurnal variation of these biomarkers and the daytime rhythm of power and muscle fatigue during repeated sprint ability (RSA) exercise. In counterbalanced order, 12 football (soccer) players performed an RSA test (5?×?[6 s of maximal cycling sprint?+?24 s of rest]) on two different occasions: 07:00–08:30?h and 17:00–18:30?h. Fasting blood samples were collected from a forearm vein before and 3–5?min after each RSA test. Core temperature, rating of perceived exertion, and performances (i.e., Sprint 1, Sprint 2, and power decrease) during the RSA test were significantly higher at 17:00 than 07:00?h (p?<?.001, p?<?.05, and p?<?.05, respectively). The results also showed significant diurnal variation of resting Hcy levels and all biological markers of muscle injury with acrophases (peak times) observed at 17:00?h. These fluctuations persisted after the RSA test. However, biomarkers of antioxidant status' resting levels (i.e., total antioxidant status, uric acid, and total bilirubin) were higher in the morning. This TOD effect was suppressed after exercise for TAS and uric acid. In conclusion, the present study confirms diurnal variation of Hcy, selected biological markers of cellular damage, and antioxidant status in young football players. Also, the higher performances and muscle fatigue showed in the evening during RSA exercise might be due to higher levels of biological markers of muscle injury and lower antioxidant status at this TOD. (Author correspondence: n_souissi@yahoo.fr)     

Increased Skeletal Muscle 11βHSD1 mRNA Is Associated with Lower Muscle Strength in Ageing

Background

Sarcopenia, the loss of muscle mass and function with age, is associated with increased morbidity and mortality. Current understanding of the underlying mechanisms is limited. Glucocorticoids (GC) in excess cause muscle weakness and atrophy. We hypothesized that GC may contribute to sarcopenia through elevated circulating levels or increased glucocorticoid receptor (GR) signaling by increased expression of either GR or the GC-amplifying enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11βHSD1) in muscle.

Methods

There were 82 participants; group 1 comprised 33 older men (mean age 70.2years, SD 4.4) and 19 younger men (22.2years, 1.7) and group 2 comprised 16 older men (79.1years, 3.4) and 14 older women (80.1years, 3.7). We measured muscle strength, mid-thigh cross-sectional area, fasting morning plasma cortisol, quadriceps muscle GR and 11βHSD1 mRNA, and urinary glucocorticoid metabolites. Data were analysed using multiple linear regression adjusting for age, gender and body size.

Results

Muscle strength and size were not associated with plasma cortisol, total urinary glucocorticoids or the ratio of urinary 5β-tetrahydrocortisol +5α-tetrahydrocortisol to tetrahydrocortisone (an index of systemic 11βHSD activity). Muscle strength was associated with 11βHSD1 mRNA levels (β -0.35, p = 0.04), but GR mRNA levels were not significantly associated with muscle strength or size.

Conclusion

Although circulating levels of GC are not associated with muscle strength or size in either gender, increased cortisol generation within muscle by 11βHSD1 may contribute to loss of muscle strength with age, a key component of sarcopenia. Inhibition of 11βHSD1 may have therapeutic potential in sarcopenia.     

Muscle wasting in the presence of disease,why is it so variable?

Skeletal muscle wasting is a common clinical feature of many chronic diseases and also occurs in response to single acute events. The accompanying loss of strength can lead to significant disability, increased care needs and have profound negative effects on quality of life. As muscle is the most abundant source of amino acids in the body, it appears to function as a buffer for fuel and substrates that can be used to repair damage elsewhere and to feed the immune system. In essence, the fundamentals of muscle wasting are simple: less muscle is made than is broken down. However, although well‐described mechanisms modulate muscle protein turnover, significant individual differences in the amount of muscle lost in the presence of a given severity of disease complicate the understanding of underlying mechanisms and suggest that individuals have different sensitivities to signals for muscle loss. Furthermore, the rate at which muscle protein is turned over under normal conditions means that clinically significant muscle loss can occur with changes in the rate of protein synthesis and/or breakdown that are too small to be measurable. Consequently, the changes in expression of factors regulating muscle turnover required to cause a decline in muscle mass are small and, except in cases of rapid wasting, there is no consistent pattern of change in the expression of factors that regulate muscle mass. MicroRNAs are fine tuners of cell phenotype and are therefore ideally suited to cause the subtle changes in proteome required to tilt the balance between synthesis and degradation in a way that causes clinically significant wasting. Herein we present a model in which muscle loss as a consequence of disease in non‐muscle tissue is modulated by a set of microRNAs, the muscle expression of which is associated with severity of disease in the non‐muscle tissue. These microRNAs alter fundamental biological processes including the synthesis of ribosomes and mitochondria leading to reduced protein synthesis and increased protein breakdown, thereby freeing amino acids from the muscle. We argue that the variability in muscle loss observed in the human population arises from at least two sources. The first is from pre‐existing or disease‐induced variation in the expression of microRNAs controlling the sensitivity of muscle to the atrophic signal and the second is from the expression of microRNAs from imprinted loci (i.e. only expressed from the maternally or paternally inherited allele) and may control the rate of myonuclear recruitment. In the absence of disease, these factors do not correlate with muscle mass, since there is no challenge to the established balance. However, in the presence of such a challenge, these microRNAs determine the rate of decline for a given disease severity. Together these mechanisms provide novel insight into the loss of muscle mass and its variation in the human population. The involvement of imprinted loci also suggests that genes that regulate early development also contribute to the ability of individuals to resist muscle loss in response to disease.     

Efficient Single Muscle Fiber Isolation from Alcohol-Fixed Adult Muscle following ��-Galactosidase Staining for Satellite Cell Detection

Staining for β-galactosidase activity for whole tissues, sections, and cells is a common method to detect expression of β-galactosidase reporter transgene as well as senescence-dependent β-galactosidase activity. Choice of fixatives is a critical step for detection of β-galactosidase activity, subsequent immunostaining, and enzymatic digestion of tissue to dissociate cells. In this report, the authors examined several aldehyde and alcohol fixatives in mouse skeletal muscle tissues for their efficiency at improving detection of β-galactosidase activity as well as detection by immunostaining. In addition, fixatives were also analyzed for their efficiency for collagenase digestion to isolate single muscle fibers on postfixed β-galactosidase-stained whole skeletal muscle tissues. The results show that fixing cells with isopropanol yields the greatest reliability and intensity in both β-galactosidase staining as well as double staining for β-galactosidase activity and antibodies. In addition, isopropanol and ethanol, but not glutaraldehyde or paraformaldehyde, allow for the isolation of single muscle fibers from the diaphragm and tibialis anterior muscles following postfixed β-galactosidase staining. Using this method, it is possible to identify the amount of cells that occupy the satellite cell compartment in single muscle fibers prepared from any muscle tissues, including tibialis anterior muscle and diaphragm.     

Muscle assembly: a titanic achievement?

The formation of perfectly aligned myofibrils in striated muscle represents a dramatic example of supramolecular assembly in eukaryotic cells. Recently, considerable progress has been made in deciphering the roles that titin, the third most abundant protein in muscle, has in this process. An increasing number of sarcomeric proteins (ligands) are being identified that bind to specific titin domains. Titin may serve as a molecular blueprint for sarcomere assembly and turnover by specifying the precise position of its ligands within each half-sarcomere in addition to functioning as a molecular spring that maintains the structural integrity of the contracting myofibrils.     

ERRγ Promotes Angiogenesis,Mitochondrial Biogenesis,and Oxidative Remodeling in PGC1α/β-Deficient Muscle

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Na,K-ATPase Activity in Mouse Muscle is Regulated by AMPK and PGC-1α

Methyl-β-cyclodextrins (MβCDs) are molecules that are extensively used to remove and to load cholesterol (Chol) from artificial and natural membranes; however, the mechanism of Chol extraction by MβCD from pure lipids or from complex mixtures is not fully understood. One of the outstanding questions in this field is the capability of MβCD to remove Chol from lipid domains having different packing. Here, we investigated the specificity of MβCD to remove Chol from coexisting macrodomains with different lipid packing. We used giant unilamellar vesicles (GUVs) made of 1,2-dioleoylphosphatidylcholine:1,2-dipalmitoylphatidylcholine:free cholesterol, 1:1:1 molar ratio at 27°C. Under these conditions, individual GUVs present Chol distributed into l _o and l _d phases. The two phases can be distinguished and visualized using Laurdan generalized polarization and two-photon excitation fluorescence microscopy. Our data indicate that MβCD removes Chol preferentially from the more disordered phase. The process of selective Chol removal is dependent on the MβCD concentration. At high concentrations, MβCD also removes phospholipids.     

Muscle fatigue: conduction or mechanical failure?

It is well documented that repeated voluntary activity or electrical stimulation of skeletal muscle results in a decline in force production or power output. However, the precise physiological causes of "muscle fatigue" are not yet well understood. It is conceivable that the mechanism(s) may lie either in the conduction of action potentials in the central and peripheral nervous systems or in the transformation of the electrical event into mechanical force production by the muscle itself. In fact, none of the components of the electrical pathway from generation of impulses in the brain to their conduction over the neuron and the excitable membranes of the muscle can as yet be ruled out as potential contributors to the fatigue process. Relative to that on conduction failure, more information exists concerning the possibility that a defect in the excitation contraction coupling process in skeletal muscle, e.g., intracellular acidosis, inadequate supply of energy for contraction, or a disruption in Ca2+ homeostasis may also be significant in compromising force production following sustained activity. Despite this, the amount of conflicting data derived from these experiments has hindered the resolution of this question. In the future more attention must be given to such issues as the type of activity used to elicit fatigue and the fiber composition of the muscles studied. This is imperative as these factors clearly impact the nature of correlations between the biochemical and physiological events in muscle that are required to support prospective fatigue mechanisms.     

Muscle development in the marbled crayfish—insights from an emerging model organism (Crustacea, Malacostraca, Decapoda)

The development of the crustacean muscular system is still poorly understood. We present a structural analysis of muscle development in an emerging model organism, the marbled crayfish—a representative of the Cambaridae. The development and differentiation of muscle tissue and its relation to the mesoderm-forming cells are described using fluorescent and non-fluorescent imaging tools. We combined immunohistochemical staining for early isoforms of myosin heavy chain with phallotoxin staining of F-actin, which distinguishes early and more differentiated myocytes. We were thus able to identify single muscle precursor cells that serve as starting points for developing muscular units. Our investigations show a significant developmental advance in head appendage muscles and in the posterior end of the longitudinal trunk muscle strands compared to other forming muscle tissues. These findings are considered evolutionary relics of larval developmental features. Furthermore, we document the development of the muscular heart tissue from myogenic precursors and the formation and differentiation of visceral musculature.     

Gαi2 Signaling Is Required for Skeletal Muscle Growth,Regeneration, and Satellite Cell Proliferation and Differentiation

We have previously shown that activation of Gαi2, an α subunit of the heterotrimeric G protein complex, induces skeletal muscle hypertrophy and myoblast differentiation. To determine whether Gαi2 is required for skeletal muscle growth or regeneration, Gαi2-null mice were analyzed. Gαi2 knockout mice display decreased lean body mass, reduced muscle size, and impaired skeletal muscle regeneration after cardiotoxin-induced injury. Short hairpin RNA (shRNA)-mediated knockdown of Gαi2 in satellite cells (SCs) leads to defective satellite cell proliferation, fusion, and differentiation ex vivo. The impaired differentiation is consistent with the observation that the myogenic regulatory factors MyoD and Myf5 are downregulated upon knockdown of Gαi2. Interestingly, the expression of microRNA 1 (miR-1), miR-27b, and miR-206, three microRNAs that have been shown to regulate SC proliferation and differentiation, is increased by a constitutively active mutant of Gαi2 [Gαi2(Q205L)] and counterregulated by Gαi2 knockdown. As for the mechanism, this study demonstrates that Gαi2(Q205L) regulates satellite cell differentiation into myotubes in a protein kinase C (PKC)- and histone deacetylase (HDAC)-dependent manner.     

Muscle gene expression of CGRP-α, CGRP receptor,nAchR-β, and GDNF in response to different endurance training protocols of Wistar rats

Molecular Biology Reports - The neuromuscular junction underwent adaptations to meet the demands of muscles following increased muscle activity. This study aimed to investigate the effects of...     

Rat Fibroblasts Cultured from Various Organs Exhibit Differences in α-Smooth Muscle Actin Expression, Cytoskeletal Pattern, and Adhesive Structure Organization

In vivo,α-smooth muscle actin (SMA) is expressedde novoand temporarily by fibroblastic cells during wound healing and correlates particularly with wound contraction. In culture, the presence of varying proportions of cells expressing and not expressing this actin isoform (α-SMA-positive and α-SMA-negative cells) is characteristic of fibroblastic populations from different tissues. It is possible that mechanisms controlling the expression of actin isoforms, and thus modulating cytoskeleton-related functions, play a major role in the organization of cell shape and motility. We have compared the cell shape as well as the cytoskeleton and focal contact organization in α-SMA-positive and α-SMA-negative rat fibroblasts from various organs (i.e., skeletal muscle, dermis, subcutaneous tissue, and lung). Within each category, i.e., α-SMA-positive or α-SMA-negative fibroblasts, no significant morphological differences were seen among populations derived from different tissues. In contrast, α-SMA-positive and α-SMA-negative fibroblasts were significantly different, independently of their origin: α-SMA-positive cells had larger average areas, higher numbers of narrow extensions at the edges, larger focal adhesions with the substratum, and a more important network of cellular fibronectin than α-SMA-negative cells. Thus, α-SMA-positive and α-SMA-negative variants naturally present in fibroblastic populations exhibit important phenotypic differences probably associated with distinct functional activities.     

Does Magnesium Supplementation Improve Body Composition and Muscle Strength in Middle-Aged Overweight Women? A Double-Blind, Placebo-Controlled, Randomized Clinical Trial

Muscle strength, an independent predictor of metabolic disorders, disability, and mortality, reduces gradually with advancing age. Little is known about the influence of nutritional intervention on muscle strength in middle-aged. The aim of the present study is to examine whether magnesium could improve body composition and muscle strength in middle-aged overweight women. In this double-blinded, placebo-controlled, randomized trial, a total of 74 healthy middle-aged overweight women (25?≤?BMI?≤?30 kg/m²) received either 250 mg magnesium in the form of magnesium oxide or placebo daily for 8 weeks. Body composition was assessed using Bioelectrical Impedance Analysis (BIA). Handgrip strength and knee extension strength were measured with isometric dynamometry. Functional mobility was assessed using Time Get Up and Go Test (TGUG). A significant increase in mean lean body mass was observed (P?=?0.05) accompanied with a significant decrease in fat mass (P?=?0.02) solely in the magnesium group at the end of 8 weeks compared to baseline values but the changes did not reach significant as compared to placebo group. Handgrip strength and TGUG improved in the magnesium group compared to baseline but they were not significant compared to placebo. There were no significant differences in increasing knee extension strength in the magnesium group as compared with placebo. Baseline values of serum magnesium and muscle strength of participants did not indicate any influences on response to magnesium supplementation. Our findings indicate that magnesium as magnesium oxide, 250 mg/day, for 8 weeks do not lead to a significant greater gain in muscle strength and function compared to placebo.     

Kakapo,a Novel Cytoskeletal-associated Protein Is Essential for the Restricted Localization of the Neuregulin-like Factor,Vein, at the Muscle–Tendon Junction Site

In the Drosophila embryo, the correct association of muscles with their specific tendon cells is achieved through reciprocal interactions between these two distinct cell types. Tendon cell differentiation is initiated by activation of the EGF-receptor signaling pathway within these cells by Vein, a neuregulin-like factor secreted by the approaching myotube. Here, we describe the cloning and the molecular and genetic analyses of kakapo, a Drosophila gene, expressed in the tendons, that is essential for muscle-dependent tendon cell differentiation. Kakapo is a large intracellular protein and contains structural domains also found in cytoskeletal-related vertebrate proteins (including plakin, dystrophin, and Gas2 family members). kakapo mutant embryos exhibit abnormal muscle-dependent tendon cell differentiation. A major defect in the kakapo mutant tendon cells is the failure of Vein to be localized at the muscle–tendon junctional site; instead, Vein is dispersed and its levels are reduced. This may lead to aberrant differentiation of tendon cells and consequently to the kakapo mutant deranged somatic muscle phenotype.