Skeletal Muscle Fiber Type Composition Research Articles

Diversity and Differential Expression of MicroRNAs in the Human Skeletal Muscle with Distinct Fiber Type Composition.

The ratio of fast- and slow-twitch fibers in human skeletal muscle is variable and largely determined by genetic factors. In this study, we investigated the contribution of microRNA (miRNA) in skeletal muscle fiber type composition. The study involved biopsy samples of the vastus lateralis muscle from 24 male participants with distinct fiber type ratios. The miRNA study included samples from five endurance athletes and five power athletes with the predominance of slow-twitch (61.6-72.8%) and fast-twitch (69.3-80.7%) fibers, respectively. Total and small RNA were extracted from tissue samples. Total RNA sequencing (N = 24) revealed 352 differentially expressed genes between the groups with the predominance of fast- and slow-twitch muscle fibers. Small RNA sequencing showed upregulation of miR-206, miR-501-3p and miR-185-5p, and downregulation of miR-499a-5p and miR-208-5p in the group of power athletes with fast-twitch fiber predominance. Two miRtronic miRNAs, miR-208b-3p and miR-499a-5p, had strong correlations in expression with their host genes (MYH7 and MYH7B, respectively). Correlations between the expression of miRNAs and their experimentally validated messenger RNA (mRNA) targets were calculated, and 11 miRNA-mRNA interactions with strong negative correlations were identified. Two of them belonged to miR-208b-3p and miR-499a-5p, indicating their regulatory links with the expression of CDKN1A and FOXO4, respectively.

Skeletal Muscle Fiber Type Composition and Citrate Synthase Activity in Fit and Unfit Warmbloods and Quarter Horses

Selective breeding and discipline specific training has led to equine breeds adept at various athletic disciplines. Breed-specific skeletal muscle adaptations have been studied in many breeds but not Warmbloods (WB). We evaluated gluteal muscle contractile muscle fiber types and citrate synthase activity (CS), a marker for mitochondrial volume density, in WB trained for dressage (second level-Grand Prix) contrasted with Quarter Horses (QH). Gluteus medius muscle biopsies from 14 unfit/18 fit dressage-trained WB and 20 unfit/16 fit reining/working cow QH were analyzed fluorometrically and fiber types determined by ATPase activity. Comparisons were made by one-way ANOVA. Unfit and fit WB had significantly higher % type 1 and lower % type 2X fibers than QH. Unfit WB had significantly higher CS than unfit QH but CS did not differ between fit WB and fit QH. CS was only significantly higher in fit versus unfit QH, not fit versus unfit WB. In conclusion, WB gluteal muscle has an inherently high % type 1/low % type 2X fibers and high mitochondrial content whether unfit or trained for dressage, contrasting QH with an inherently low % type 1/high % type 2X and low mitochondrial content, that was enhanced in fit QH. Similar CS activity in fit WB versus QH despite a two-fold difference in % type 2X fibers indicates that mitochondrial volume density cannot accurately be predicted from contractile fiber type composition.

The ROS scavenger PDTC affects adaptation to treadmill running in mice: distinct effects on murine body mass, resting heart rate and skeletal muscle fiber type composition.

Regular exercise induces a broad spectrum of adaptation reactions in a variety of tissues and organs. However, the respective mechanisms are incompletely understood. In the context of their analysis, animal model systems, specifically rodent treadmill running protocols, play an important role. However, few researchers have studied different aspects of adaptation, such as cardiorespiratory and skeletal muscle training effects, within one set of experiments. Here, we analyzed physiological adaptation to 10weeks of regular, moderate-intensity, uphill treadmill running in mice, a widely used model for endurance exercise training. To study the effects of reactive oxygen species (ROS), which have been suggested to be major regulators of training adaptation, a subgroup of mice was treated with the ROS scavenger PDTC (pyrrolidine dithiocarbamate). We found that mass gain in mice that exercised under PDTC treatment lagged behind that of all other experimental groups. In addition, both exercise and PDTC significantly and additively decreased resting heart rate. Furthermore, there was a trend towards an enhanced proportion of type 2A skeletal muscle fibers and differential expression of metabolism-associated genes, indicating metabolic and functional adaptation of skeletal muscle fibers. By contrast, there were no effects on grip strength and relative mass of individual muscles, suggesting that our protocol of uphill running did not increase skeletal muscle hypertrophy and strength. Taken together, our data suggest that a standard protocol of moderate-intensity uphill running induces adaptation reactions at multiple levels, part of which might be modulated by ROS, but does not enhance skeletal muscle hypertrophy and force.

Skeletal Muscle Fiber Type in Hypoxia: Adaptation to High-Altitude Exposure and Under Conditions of Pathological Hypoxia.

Skeletal muscle is able to modify its size, and its metabolic/contractile properties in response to a variety of stimuli, such as mechanical stress, neuronal activity, metabolic and hormonal influences, and environmental factors. A reduced oxygen availability, called hypoxia, has been proposed to induce metabolic adaptations and loss of mass in skeletal muscle. In addition, several evidences indicate that muscle fiber-type composition could be affected by hypoxia. The main purpose of this review is to explore the adaptation of skeletal muscle fiber-type composition to exposure to high altitude (ambient hypoxia) and under conditions of pathological hypoxia, including chronic obstructive pulmonary disease (COPD), chronic heart failure (CHF) and obstructive sleep apnea syndrome (OSAS). The muscle fiber-type composition of both adult animals and humans is not markedly altered during chronic exposure to high altitude. However, the fast-to-slow fiber-type transition observed in hind limb muscles during post-natal development is impaired in growing rats exposed to severe altitude. A slow-to-fast transition in fiber type is commonly found in lower limb muscles from patients with COPD and CHF, whereas a transition toward a slower fiber-type profile is often found in the diaphragm muscle in these two pathologies. A slow-to-fast transformation in fiber type is generally observed in the upper airway muscles in rodent models of OSAS. The factors potentially responsible for the adaptation of fiber type under these hypoxic conditions are also discussed in this review. The impaired locomotor activity most likely explains the changes in fiber type composition in growing rats exposed to severe altitude. Furthermore, chronic inactivity and muscle deconditioning could result in the slow-to-fast fiber-type conversion in lower limb muscles during COPD and CHF, while the factors responsible for the adaptation of muscle fiber type during OSAS remain hypothetical. Finally, the role played by cellular hypoxia, hypoxia-inducible factor-1 alpha (HIF-1α), and other molecular regulators in the adaptation of muscle fiber-type composition is described in response to high altitude exposure and conditions of pathological hypoxia.

The regulation of skeletal muscle fiber-type composition by betaine is associated with NFATc1/MyoD.

Increasing evidence indicates that muscular dysfunction or alterations in skeletal muscle fiber-type composition not only are involved in muscle metabolism and function but also can limit functional capacity. Therefore, understanding the mechanisms regulating key events during skeletal myogenesis is necessary. Betaine is a naturally occurring component of commonly eaten foods. Here, we showed that 10mM betaine supplementation in vitro significantly repressed myoblast proliferation and enhanced myoblast differentiation. This effect can be mediated by regulation of miR-29b-3p. Further analysis showed that betaine supplementation in vitro regulated skeletal muscle fiber-type composition through the induction of NFATc1 and the negative regulation of MyoD expression. Furthermore, mice fed with 10mM betaine in water for 133days showed no impairment in overall health. Consistently, betaine supplementation increased muscle mass, promoted muscle formation, and modulated the ratio of fiber types in skeletal muscle in vivo. These findings shed light on the diverse biological functions of betaine and indicate that betaine supplementation may lead to new therapies for diseases such as muscular dystrophy or other diseases related to muscle dysfunction. KEY MESSAGES: Betaine supplementation inhibits proliferation and promotes differentiation of C2C12 myoblasts. Betaine supplementation regulates fast to slow muscle fiber-type conversion and is associated with NFATc1/MyoD. Betaine supplementation enhances skeletal myogenesis in vivo. Betaine supplementation does not impair health of mice.

MicroRNA-139-5p suppresses myosin heavy chain I and IIa expression via inhibition of the calcineurin/NFAT signaling pathway

MicroRNAs (miRNAs) are a class of small non-coding RNAs that are widely involved in a variety of biological processes. Different skeletal muscle fiber type composition exhibits characteristic differences in functional properties and energy metabolism of skeletal muscle. However, the molecular mechanism by which miRNAs control the different type of muscle fiber formation is still not fully understood. In the present study, we characterized the role of microRNA-139-5p (miR-139-5p) in the regulation of myosin heavy chain (MyHC) isoform expression and its underlying mechanisms. Here we found that the expression of miR-139-5p was significantly higher in mouse slow-twitch muscle than in fast-twitch muscle. Overexpression of miR-139-5p downregulated the expression of MyHC I and MyHC IIa, whereas inhibition of miR-139-5p upregulated them. We also found that the levels of calcineurin (CaN), NFATc1, MEF2C and MCIP1.4, which are the components of CaN/NFAT signaling pathway that has shown to positively regulate slow fiber-selective gene expression, were notably inhibited by miR-139-5p overexpression. Furthermore, treatment of phenylephrine (PE), a α1-adrenoceptor agonist, abolished the inhibitory effect of miR-139-5p on MyHC I and MyHC IIa expression. Together, our findings indicated that the role of miR-139-5p in regulating the MyHC isoforms, especially MyHC I and MyHC IIa, may be achieved through inhibiting CaN/NFAT signaling pathway.

Tensiomyographic Assessment of Muscle Contractile Properties in 9- to 14-Year Old Children.

While there are numerous data on the skeletal muscle fiber type composition in adults, little is known about the changes in fiber type composition and contractile properties during maturational growth in children. Using noninvasive tensiomyography, we measured contraction time (Tc), an indirect estimate of the myosin heavy chain I (MHC-I) proportion, to assess the longitudinal changes of the biceps brachii (BB), biceps femois (BF), vastus lateralis (VL), and erector spinae (ES) muscles in 53 boys and 54 girls. The children were 9 years at the start of the study and returned for 5 follow-up measurements until the age of 14 years. The ES has the shortest and the BF has the longest Tc. The VL and ES of boys have shorter Tc than those of girls. When applying the relationship between proportion of MHC-I and Tc established in adults to children's TMG data, we found a slow-to-fast transition in the VL between, at least, the ages of 6 to 10 years, when it stabilized to adult proportions. Regular participation in sports was associated with a faster BF, but not in the VL. Our data represents a first non-invasive indication of the developmental changes in muscle fiber type composition in children.

1,25‐Dihydroxyvitamin D3 Mediates L6 Myoblast Differentiation via Vitamin D Receptor (VDR)

In our previous study, it was demonstrated that skeletal muscle fiber type composition was changed by dietary supplementation of 25‐hydroxyvitamin D3 in growing rats. Potential mechanisms of 25‐hydroxyvitamin D3, a precursor of 1,25‐dihydroxyvitamin D3 (active form), have been suggested in previous studies that VDR was involved with mouse C2C12 myoblasts. However, it has not been explained in rat L6 myoblasts. In this study, we examined the role of VDR in skeletal muscle differentiation and fiber type‐specific gene expression by using short hairpin RNA (shRNA) for VDR knockdown in rat L6 myoblast. L6 myoblasts transfected either with scrambled or shVDR vector were treated with vehicle (CON), 1 pM, 10 pM, or 100 pM 1,25‐dihydroxyvitamin D3 (VitD) for 8 days of differentiation. VDR knockdown was confirmed by Western blot. And myogenic regulatory markers and fiber type‐specific gene expression levels were determined by qPCR. VDR protein was increased by VitD at 10 pM and 100 pM compared to that of CON (1.46 and 1.89 folds, respectively, p<0.05). However, shVDR was effective in suppressing VDR up to 10 pM as VDR protein levels were decreased to 34 %, 24 %, and 39 % (CON, 1 pM, and 10 pM, respectively, p<0.05) compared to scrambled vector. And at 100 pM VitD, VDR was not repressed by shVDR compared to that of scrambled vector. Relative gene expression levels of both MyoD and Myogenin were downregulated by VDR knockdown in CON (0.56 and 0.36 folds, respectively, p<0.05), however, VitD treatment abolished the effects. Fiber type‐specific genes were also modulated by VDR knockdown. Gene expression level of Myh7, dominantly expressed gene in slow‐twitch fiber, was suppressed to 37 %, 29 %, and 43 % (CON, 1 pM, and 10 pM, respectively, p<0.05) by VDR knockdown compared to scrambled vector, but was restored at 100 pM. Myh2, which consists of fast‐twitch fiber, was also downregulated by VDR knockdown at CON (0.57 folds, p<0.05), but its level was restored by VitD treatment. The present study demonstrates that VDR affects mRNA levels of the genes involved in L6 myoblast differentiation and fiber type specificity. Also, the VDR knockdown effect was partially modulated by VitD. It could suggest that the effects of VitD on myogenic differentiation might be partially mediated through VDR.

Correlation between Plasma 25‐hydroxyvitamin D and Fiber Composition of Skeletal Muscle in Response to Dietary Vitamin D3 or 25‐hydroxyvitamin D3 Supplementation

An experiment was conducted to examine the effects of dietary vitamin D3 (D3) level and 25‐hydroxyvitamin D3 (25OHD3) on plasma 25‐hydroxyvitamin D and skeletal muscle. Growing rats (3‐week‐old) were fed either D3‐sufficient diet (1000 IU/kg diet) or D3‐deficient diet for 4 weeks (Phase I). During Phase II for another 4 weeks, rats fed D3‐deficient diet were allotted to 7 treatments. Experimental diets included D3 at 0, 400, 700, 1000, 1500, and 2000 IU/kg diet or 400 IU 25OHD3/kg diet. Positive and negative control groups were fed either D3‐containing (1000 IU/kg diet) or D3‐deficient diet throughout the experiment. Body weight, plasma 25‐hydroxyvitamin D, bone mineral density (BMD), lean percentage, and bicep femoris (representative of skeletal muscle) mass were determined. Also, fiber type composition and relative gene expression levels of myosin heavy chain (MHC) isoforms, PGC‐1α and Mitofusin 1 (Mfn1) in bicep femoris were determined. Dietary D3 levels or 25OHD3 did not affect body weight, BMD, lean percentage, or skeletal muscle mass in growing rats. However, plasma 25‐hydroxyvitamin D level was increased in a dose‐dependent manner upon dietary D3 level, and 25OHD3 group resulted in the highest level among treatments. Compared to the same dose of D3 (400 IU/kg diet), dietary 25OHD3 showed 2.3‐fold increase in plasma 25‐hydroxyvitamin D level (23.5±5.1 ng/mL vs. 53.3±1.5 ng/mL). Skeletal muscle fiber type composition was also altered by dietary D3 levels and 25OHD3. Vitamin D deficiency decreased type I fiber percentage and this decrease was diminished by dietary D3 in a dose‐dependent manner. Dietary 25OHD3, on the other hand, showed much more increase in type I percentage and decrease in type II percentage compared to same dose of D3 (400 IU/kg diet) (15.1±1.6 % vs. 5.4±1.7 %, and 63.4±1.6 % vs. 79.1±6.7 %), and showed the highest percentage of type I fiber than all the other treatments. However, relative gene expression levels of MHC isoforms did not coincided with the phenotypic difference. MHC I, PGC‐1α and Mfn1 transcript levels were numerically increased in response to dietary D3 level with the highest value resulted from 1500 IU/kg D3 supplementation. It showed 3‐fold increase in MHC I (3.4±2.4 vs. 1.0±0.7) and PGC‐1α (3.6±0.8 vs. 1.0±0.5), and 2‐fold increase in Mfn1 (2.4±0.6 vs. 1.0±0.8) transcript levels compared to positive control. In summary, 4 weeks of dietary D3 or 25OHD3 supplementation did not affect growth, lean percentage, skeletal muscle mass, and BMD in growing rats. However, plasma 25‐hydroxyvitamin D level, percentage of type I fiber and its gene expression levels in skeletal muscle responded to dietary D3 levels and 25OHD3. Also, it is notable that correlation between plasma 25‐hydroxyvitamin D level and percentage of type I fiber in bicep femoris was observed (r2=0.86). The present study demonstrates responses of plasma 25‐hydroxyvitamin D level and skeletal muscle fiber type composition upon dietary D3 and 25OHD3 with greater efficacy.

Physical activity-induced remodeling of vasculature in skeletal muscle: role in treatment of type 2 diabetes

This manuscript summarizes and discusses adaptations of skeletal muscle vasculature induced by physical activity and applies this understanding to benefits of exercise in prevention and treatment of type 2 diabetes (T2D). Arteriolar trees of skeletal muscle are heterogeneous. Exercise training increases capillary exchange and blood flow capacities. The distribution of vascular adaptation to different types of exercise training are influenced by muscle fiber type composition and fiber recruitment patterns that produce different modes of exercise. Thus training-induced adaptations in vascular structure and vascular control in skeletal muscle are not homogeneously distributed throughout skeletal muscle or along the arteriolar tree within a muscle. Results summarized indicate that similar principles apply to vascular adaptation in skeletal muscle in T2D. It is concluded that exercise training-induced changes in vascular gene expression differ along the arteriolar tree and by skeletal muscle fiber type composition. Results suggest that it is unlikely that hemodynamic forces are the only exercise-induced signals mediating the regulation of vascular gene expression. In patients with T2D, exercise training is perhaps the most effective treatment of the many related symptoms. Training-induced changes in the vasculature and in insulin signaling in the muscle fibers and vasculature augment glucose and insulin delivery as well as glucose uptake. If these adaptations occur in a sufficient amount of muscle mass, exposure to hyperglycemia and hyperinsulinemia will decrease along with the risk of microvascular complications throughout the body. It is postulated that exercise sessions in programs of sufficient duration, that engage as much skeletal muscle mass as possible, and that recruit as many muscle fibers within each muscle as possible will produce the greatest benefit. The added benefit of combined resistance and aerobic training programs and of high-intensity exercise programs is not simply "more exercise is better".

Muscle variables of importance for physiological performance in competitive football

To examine how match performance parameters in trained footballers relate to skeletal muscle parameters, sprint ability and intermittent exercise performance. 19 male elite football players completed an experimental game with physical performance determined by video analysis and exercise capacity assessed by intermittent Yo-Yo IR1 and IR2 tests, and a repeated sprint test (RST). Muscle tissue was obtained for analysis of metabolic enzyme maximal activity and key muscle protein expression. Total game distance, distance deficit from first to second half and high-intensity running in the final 15min of the game were all correlated to the players' Yo-Yo IR1 performance (r=0.55-0.87) and beta-hydroxyacyl-CoA-dehydrogenase (HAD) maximal activity (r=0.55-0.65). Furthermore, platelet/endothelial cell adhesion molecule-1 (PECAM1) protein expression was weakly (r=0.46) correlated to total game distance. Peak 5-min game distance faster than 21kmh(-1) was related to the Na(+)-K(+) ATPase subunit (α1, α2, β1 and FXYD1) protein levels (r=0.54-0.70), while Yo-Yo IR2 performance explained 40% of the variance in high-intensity game distance. Total and 1-min peak sprint distance correlated to myosin heavy chain II/I ratio (MHCII/I ratio) and sarcoendoplasmic reticulum Ca(2+) ATPase isoform-1 (SERCA1) protein (r=0.56-0.86), while phosphofructokinase (PFK) maximal activity also correlated to total sprint distance (r=0.46). The findings emphasize the complexity of parameters predicting physical football performance with Yo-Yo IR1 and HAD as the best predictors of total distance, while high expression of Na(+)-K(+) ATPase proteins and the Yo-Yo IR2 test are better predictors of high-intensity performance. Finally, sprint performance relates to skeletal muscle fiber-type composition.

Integrative and adaptive responses.

Darwin is considered the father of modern evolutionary biology by recognizing the importance of integrative and adaptive structural and physiological responses in living organisms. In his textbook Principles of Biology (1864), Herbert Spencer succinctly stated the essence of Darwin's theory of

Effects of myogenin on muscle fiber types and key metabolic enzymes in gene transfer mice and C2C12 myoblasts

Skeletal muscle fiber type composition is one of the important factors influencing muscle growth and meat quality. As a member of the myogenic transcription factors, myogenin (MyoG) is required for embryonic myoblast differentiation, but the expression of MyoG continues in mature muscle tissue of adult animals, especially in oxidative metabolic muscle, which suggests that MyoG may play a more extended role. Therefore, using MyoG gene transfer mice and C2C12 myoblasts as in vivo and in vitro models, respectively, we elected to study the role of MyoG in muscle fiber types and oxidative metabolism by using overexpression and siRNA suppression strategies. The overexpression of MyoG by DNA electroporation in mouse gastrocnemius muscle had no significant effect on fiber type composition but upregulated the mRNA expression (P<0.01) and enzyme activity (P<0.05) of oxidative succinic dehydrogenase (SDH). In addition, downregulation of the activity of the glycolytic enzymes lactate dehydrogenase (LDH, P<0.05) and pyruvate kinase (PK, P<0.05) was observed in MyoG gene transfer mice. In vitro experiments verified the results obtained in mice. Stable MyoG-transfected differentiating C2C12 cells showed higher mRNA expression levels of myosin heavy chain (MyHC) isoform IIX (P<0.01) and SDH (P<0.05), while the LDH mRNA was attenuated. The enzyme activities of SDH (P<0.01) and LDH (P<0.05) were similarly altered at the mRNA level. When MyoG was knocked down in C2C12 cells, MyHC IIX expression (P<0.05) was decreased, but the mRNA level (P<0.05) and the enzyme activity (P<0.05) of SDH were increased. Downregulating MyoG also increased the activity of the glycolytic enzymes PK (P<0.05) and hexokinase (HK, P<0.05). Based on those results, we concluded that MyoG barely changes the MyHC isoforms, except MyHC IIX, in differentiating myoblasts but probably influences the shift from glycolytic metabolism towards oxidative metabolism both in vivo and in vitro. These results contribute to further understand the role of MyoG in skeletal muscle energy metabolism and also help to explore the key genes that regulate meat quality.

Effects of Resveratrol in Patients with Type 2 Diabetes Mellitus on Skeletal Muscle SIRT1 Expression and Energy Expenditure

The primary aims of the study were to examine the effect of resveratrol on skeletal muscle SIRT1 expression and energy expenditure in subjects with Type 2 diabetes mellitus (T2DM). Animal and in vivo studies indicate that resveratrol increases SIRT1 expression that stimulates PGC1α activity. Subsequent upregulation of AMPK and GLUT4 expression are associated with improved insulin sensitivity in peripheral tissues. Ten subjects with T2DM were randomized in a double-blind fashion to receive 3g resveratrol or placebo daily for 12 weeks. Secondary outcomes include measures of AMPK, p-AMPK and GLUT4 expression levels, energy expenditure, physical activity levels, distribution of abdominal adipose tissue and skeletal muscle fiber type composition, body weight, HbA1c, plasma lipid subfraction, adiponectin levels, and insulin sensitivity. There was a significant increase in both SIRT1 expression (2.01 vs. 0.86 arbitrary units [AU], p = .016) and p-AMPK to AMPK expression ratio (2.04 vs. 0.79 AU, p = .032) in the resveratrol group compared with the placebo group. Although the percentage of absolute change (8.6 vs. -13.9%, p = .033) and percentage of predicted resting metabolic rate (RMR; 7.8 vs. -13.9%, p = .013) were increased following resveratrol, there was a significant reduction in average daily activity (-38 vs. 43.2%, p = .028) and step counts (-39.5 vs. 11.8%, p = .047) when compared with placebo. In patients with T2DM, treatment with resveratrol regulates energy expenditure through increased skeletal muscle SIRT1 and AMPK expression. These findings indicate that resveratrol may have beneficial exercise-mimetic effects in patients with T2DM.

Long-term therapy with Bendavia (MTP-131), a novel mitochondria-targeting peptide, normalizes skeletal muscle fiber type composition in dogs with chronic heart failure

Background: Chronic heart failure (HF) is associated with a shift in skeletal muscle (SM) fiber type composition manifested by a decline in the number of aerobic, slow-twitch, fatigue-resistant type-1 fibers and an increase in the number of glycolytic, fast-twitch type-2 fibers. This fiber type shift is responsible, in part, for exercise intolerance in HF. Bendavia (MTP-131), a novel, first in class, MITO-targeting peptide, improves LV systolic function in dogs with chronic HF and restores ATP synthesis in multiple organs including heart, kidney and SM in other animal models of disease. We tested the hypothesis that long-term therapy with Bendavia can reverse the maladaptive SM fiber type shift in dogs with chronic HF possibly through restored MITO oxidative capacity. Methods: Studies were performed in triceps SM samples of 14 HF dogs produced by intracoronary microembolizations (LV ejection fraction ∼30%) and from 9 normal dogs. HF dogs were randomized to 3 months therapy with subcutaneous injections of Bendavia (0.5 mg/kg once daily, n=7) or saline (Control, n=7). SM type-1 and -2 fibers were differentiated histologically by myofibrillar adenosine triphosphatase staining. The proportion of Type-1 and -2 fibers and the average cross-sectional area (CSA) of each fiber type was assessed in 5 randomly selected SM fields/dog each containing ∼100 fibers. Results: The proportion of SM type-1 fibers was lower and type-2 fibers higher in HF-Control compared to normal (table) leading to a significantly lower fiber type ratio. Treatment with Bendavia restored a near normal fiber type composition. There were no differences in SM CSA among study groups (table). Skeletal Muscle Fiber Type Composition *p<0.05 vs. Normal; †p<0.05 vs. HF-Control. Conclusions: Therapy with Bendavia reverses abnormalities of SM fiber type without influencing SM CSA. Reversal of this SM maladaptation following therapy with Bendavia can lead to improved exercise tolerance in HF.

Reduced Satellite Cell Numbers with Spinal Cord Injury and Aging in Humans

Both sarcopenia and spinal cord injury (SCI) are characterized by the loss of skeletal muscle mass and function. Despite obvious similarities in atrophy between both models, differences in muscle fiber size and satellite cell content may exist on a muscle fiber type-specific level. In the present study, we compared skeletal muscle fiber characteristics between wheelchair-dependent young males with SCI (n = 8, 32 ± 4 yr), healthy elderly males (n = 8, 75 ± 2 yr), and young controls (n = 8, 31 ± 3 yr). Muscle biopsies were collected to determine skeletal muscle fiber type composition, fiber size, and satellite cell content. Severe atrophy and a shift toward approximately 90% Type II muscle fibers were observed in muscle obtained from males with SCI. Muscle fiber size was substantially smaller in both the SCI (Types I and II fibers) and elderly subjects (Type II fibers) when compared with the controls. Satellite cell content was substantially lower in the wheelchair-dependent SCI subjects in both the Types I and II muscle fibers (0.049 ± 0.019 and 0.050 ± 0.005 satellite cells per fiber, respectively) when compared with the young controls (0.104 ± 0.011 and 0.117 ± 0.009 satellite cells per fiber, respectively). In the elderly, the number of satellite cells was lower in the Type II muscle fibers only (0.042 ± 0.005 vs 0.117 ± 0.009 satellite cells per fiber in the elderly vs young controls, respectively). This is the first study to show that muscle fiber atrophy as observed with SCI (Types I and II fibers) and aging (Type II fibers) is accompanied by a muscle fiber type-specific reduction in satellite cell content in humans.

Between-day reliability of a method for non-invasive estimation of muscle composition

Tensiomyography is a method for valid and non-invasive estimation of skeletal muscle fibre type composition. The validity of selected temporal tensiomyographic measures has been well established recently; there is, however, no evidence regarding the method’s between-day reliability. Therefore it is the aim of this paper to establish the between-day repeatability of tensiomyographic measures in three skeletal muscles.For three consecutive days, 10 healthy male volunteers (mean±SD: age 24.6±3.0years; height 177.9±3.9cm; weight 72.4±5.2kg) were examined in a supine position. Four temporal measures (delay, contraction, sustain, and half-relaxation time) and maximal amplitude were extracted from the displacement–time tensiomyogram. A reliability analysis was performed with calculations of bias, random error, coefficient of variation (CV), standard error of measurement, and intra-class correlation coefficient (ICC) with a 95% confidence interval.An analysis of ICC demonstrated excellent agreement (ICC were over 0.94 in 14 out of 15 tested parameters). However, lower CV was observed in half-relaxation time, presumably because of the specifics of the parameter definition itself. These data indicate that for the three muscles tested, tensiomyographic measurements were reproducible across consecutive test days. Furthermore, we indicated the most possible origin of the lowest reliability detected in half-relaxation time.

When less is more: a simple Western blotting amendment allowing data acquisition on human single fibers

the subdivision of skeletal muscle into distinct fiber types based on myosin heavy chain (MHC) isoform expression [I, IIa, and IIx in humans; I, IIa, IIx, and IIb in rodents ([9][1])], has proven to be a practical and lasting working concept not only in studies of contractile properties but in the

Myosin heavy chain expression pattern as a marker for anabolic potency: desoxymethyltestosterone (madol), norandrostendione and testosterone repress MHC-IIb expression and stimulate MHC-IId/x expression in orchiectomized rat gastrocnemius muscle

Both 19-norandrostenedione (estr-4-ene-3,17-dione, NOR) and desoxymethyltestosterone (17alpha-methyl-5alpha-androst-2-en-17beta-ol, DMT or "madol") are 'designer steroids' misused for doping purposes in the bodybuilding scene. We have previously characterized the pharmacological profile of madol and identified potential adverse side effects. The aim of this study was to investigate the anabolic potency of NOR, madol and the reference substance testosterone propionate (TP). Besides wet weight of the M.levator ani (LA), we examined the effects on muscle fiber type composition and myosin heavy chain (MHC) expression in the M.gastrocnemius (Gas) muscle as additional markers for anabolic potency. A Hershberger assay was performed, where orchiectomized (orchi) male Wistar rats were treated subcutaneously with NOR, madol, TP or vehicle control (all 1 mg/kg BW/day) for 12 days. Wet weights of the Gas, LA, prostate and seminal vesicle were examined to determine anabolic and androgenic effects. Fiber type composition of the Gas muscle was analyzed using ATPase staining, and MHC protein profiles were determined by silver stain and Western blot analysis. NOR and madol exhibited strong anabolic and weak androgenic potency by stimulating growth of the LA but not the prostate and seminal vesicle. Skeletal muscle fiber type composition characterized by ATPase staining was not significantly altered between the treatment groups, although there was a tendency toward lower levels of type IIB and increased type IIA fibers in all treatment groups relative to orchi. MHC protein expression determined by Western blot and silver stain analysis revealed that MHC IId/x was significantly up-regulated, while MHC IIb was significantly down-regulated in NOR, madol and TP groups relative to orchi. There were no significant differences for MHC IIa and MHC I expression between groups. Results suggest that the observed MHC expression shift could serve as a molecular marker to determine anabolic activity of anabolic steroids at least in skeletal muscle of orchi rats. The molecular mechanisms as well as the androgen-dependent regulation of MHC expression in intact skeletal muscle remain to be further investigated.

GLUT4, TBC1D1, TBC1D4, TUG and RUVBL2: Relationships with Each Other and Rat Muscle Fiber Type

Skeletal muscle is composed of multiple fiber types that vary greatly in capacity for both exercise- and insulin-stimulated glucose transport (GT). Relative expression of GLUT4 and key regulatory proteins is crucial for the range in GT capacity. PURPOSE: To assess the relationships between fiber type (myosin heavy chain, MHC, isoform) and abundance of GLUT4, 2 Rab GTPase activating proteins (TBC1D1 and TBC1D4) which regulate subcellular GLUT4 localization, and 2 other proteins proposed to regulate GT [TUG (tethering protein containing a UBX domain for GLUT4) and RuvB-like protein 2 (RUVBL2)]. METHODS: 12 rat skeletal muscle or muscle regions: adductor longus (AL); extensor digitorum longus (EDL); epitrochlearis (EPI); mixed, red and white gastrocnemius (GASM, GASR and GASW); plantaris (PLAN); Soleus (SOL); red and white tibialis anterior (TAR and TAW); tensor fasciae latae (TFL); and white vastus lateralis (VLW) were dissected out and immediately freeze clamped. Muscles were homogenized and muscle lysates were used for SDS-PAGE to determine MHC isoforms expression using coomassie staining, and GLUT4, TUG, RUVBL2, TBC1D1, and TBC1D4 total abundance were determined via western blotting. RESULTS: The major findings are summarized below.TABLE CONCLUSION: The high correlation between TUG and GLUT4 is consistent with published evidence indicating that TUG binds GLUT4 and modulates its subcellular distribution. The lack of a relationship between RUVBL2 and TBC1D4 (which RUVBL2 reportedly binds and regulates in 3T3-L1 cells) may reflect the multiple functions of both proteins. The biological significance of RUVBL2's correlation with GLUT4 and TUG remains to be clarified. Recently published data suggested there might be reciprocal expression of TBC1D1 and TBC1D4 in mouse muscles, but such a relationship was not evident in rat muscles. The expression levels for 3 of 5 proteins studied which have established or potential GT regulatory roles corresponded to differences in skeletal muscle fiber type composition. Supported by the NIH (AG-010026 and NIDDK-071771)