Effect of fasting on body composition and proteolysis gene expression in skeletal muscles and liver of BEH+/+ and BEL mice.

Fasting improves health, but can cause muscle weakness. We assessed body composition in 21-week old males of Berlin high (BEH+/+) and Berlin low (BEL) strains after two bouts of 48-h or 40-h of fasting with 5-day refeeding in between, respectively. BEH+/+ mice tended to loose less weight than BEL in bout 1 and 2 (16.0 ± 2.7 versus 23.5 ± 2.9%, p < 0.001 and 17.1 ± 3.4 versus 20.4 ± 3.4%, p = 0.17, respectively). In spite of greater serum IGF-1 and body fat levels, BEH+/+ mice showed more severe muscle atrophy, but less marked liver wasting and fat depletion than BEL mice. BEH+/+ mice also showed smaller increases in expression of p62, Atrogin-1, and Mstn genes in skeletal muscles. In summary, BEL mice show resistance to fasting-induced muscle wasting in spite of low serum IGF-1 levels and high expression of genes associated with muscle atrophy.

Myostatin dysfunction does not protect from fasting-induced loss of muscle mass in mice.

OBJECTIVES: The aim of the study was to investigate if myostatin dysfunction can ameliorate fasting-induced muscle wasting. METHODS: 18-week old males from Berlin high (BEH) strain with myostatin dysfunction and wild type myostatin (BEH+/+) strain were subjected to 48-h food deprivation (FD). Changes in body composition as well as contractile properties of soleus (SOL) and extensor digitorum longus (EDL) muscles were studied. RESULTS: BEH mice were heavier than BEH+/+ mice (56.0±2.5 vs. 49.9±2.8 g, P<0.001, respectively). FD induced similar loss of body mass in BEH and BEH+/+ mice (16.6±2.4 vs. 17.4±2.2%, P>0.05), but only BEH mice experienced wasting of the gastrocnemius, tibialis anterior and plantaris muscles. FD induced a marked decrease in specific muscle force of SOL. EDL of BEH mice tended to be protected from this decline. CONCLUSION: Myostatin dysfunction does not protect from loss of muscle mass during fasting.

Pre-Exercise Rehydration Attenuates Central Fatigability during 2-Min Maximum Voluntary Contraction in Hyperthermia.

Background and objectives: Hyperthermia with dehydration alters several brain structure volumes, mainly by changing plasma osmolality, thus strongly affecting neural functions (cognitive and motor). Here, we aimed to examine whether the prevention of significant dehydration caused by passively induced whole-body hyperthermia attenuates peripheral and/or central fatigability during a sustained 2-min isometric maximal voluntary contraction (MVC). Materials and Methods: Ten healthy and physically active adult men (21 ± 1 years of age) performed an isometric MVC of the knee extensors for 2 min (2-min MVC) under control (CON) conditions, after passive lower-body heating that induced severe whole-body hyperthermia (HT, Tre > 39 °C) with dehydration (HT-D) and after HT with rehydration (HT-RH). Results: In the HT-D trial, the subjects lost 0.94 ± 0.15 kg (1.33% ± 0.13%) of their body weight; in the HT-RH trial, their body weight increased by 0.1 ± 0.42 kg (0.1% ± 0.58%). After lower-body heating, the HT-RH trial (vs. HT-D trial) was accompanied by a significantly lower physiological stress index (6.77 ± 0.98 vs. 7.40 ± 1.46, respectively), heart rate (47.8 ± 9.8 vs. 60.8 ± 13.2 b min-1, respectively), and systolic blood pressure (-12.52 ± 5.1 vs. +2.3 ± 6.4, respectively). During 2-min MVC, hyperthermia (HT-D; HT-RH) resulted in greater central fatigability compared with the CON trial. The voluntary activation of exercising muscles was less depressed in the HT-RH trial compared with the HT-D trial. Over the exercise period, electrically (involuntary) induced torque decreased less in the HT-D trial than in the CON and HT-RH trials. Conclusions: Our results suggest that pre-exercise rehydration might have the immediate positive effect of reducing physiological thermal strain, thus attenuating central fatigability even when exercise is performed during severe (Tre > 39 °C) HT, induced by passive warming of the lower body.

Efflux of creatine kinase from isolated soleus muscle depends on age, sex and type of exercise in mice.

Elevated plasma creatine kinase (CK) activity is often used as an indicator of exercise-induced muscle damage. Our aim was to study effects of contraction type, sex and age on CK efflux from isolated skeletal muscles of mice. The soleus muscle (SOL) of adult (7.5-month old) female C57BL/6J mice was subjected to either 100 passive stretches, isometric contractions or eccentric contractions, and muscle CK efflux was assessed after two-hour incubation in vitro. SOL of young (3-month old) male and female mice was studied after 100 eccentric contractions. For adult females, muscle CK efflux was larger (p < 0.05) after eccentric contractions than after incubation without exercise (698 ± 344 vs. 268 ± 184 mU·h(-1), respectively), but smaller (p < 0.05) than for young females after the same type of exercise (1069 ± 341 mU·h(-1)). Eccentric exercise-induced CK efflux was larger in muscles of young males compared to young females (2046 ± 317 vs 1069 ± 341 mU · h(-1), respectively, p < 0.001). Our results show that eccentric contractions induce a significant increase in muscle CK efflux immediately after exercise. Isolated muscle resistance to exercise-induced CK efflux depends on age and sex of mice. Key pointsMuscle lengthening contractions induce the highest CK efflux in vitro compared with similar protocol of isometric contractions or passive stretches.Muscle CK efflux in vitro is applicable in studying changes of sarcolemma permeability/integrity, a proxy of muscle damage, in response to muscle contractile activity.Isolated muscle resistance to exercise-induced CK efflux is greater in female compared to male mice of young age and is further increased in adult female mice.

Skeletal muscle hypertrophy rewires glucose metabolism: An experimental investigation and systematic review.

BACKGROUND: Proliferating cancer cells shift their metabolism towards glycolysis, even in the presence of oxygen, to especially generate glycolytic intermediates as substrates for anabolic reactions. We hypothesize that a similar metabolic remodelling occurs during skeletal muscle hypertrophy. METHODS: We used mass spectrometry in hypertrophying C2C12 myotubes in vitro and plantaris mouse muscle in vivo and assessed metabolomic changes and the incorporation of the [U-13C6]glucose tracer. We performed enzyme inhibition of the key serine synthesis pathway enzyme phosphoglycerate dehydrogenase (Phgdh) for further mechanistic analysis and conducted a systematic review to align any changes in metabolomics during muscle growth with published findings. Finally, the UK Biobank was used to link the findings to population level. RESULTS: The metabolomics analysis in myotubes revealed insulin-like growth factor-1 (IGF-1)-induced altered metabolite concentrations in anabolic pathways such as pentose phosphate (ribose-5-phosphate/ribulose-5-phosphate: +40%; P = 0.01) and serine synthesis pathway (serine: -36.8%; P = 0.009). Like the hypertrophy stimulation with IGF-1 in myotubes in vitro, the concentration of the dipeptide l-carnosine was decreased by 26.6% (P = 0.001) during skeletal muscle growth in vivo. However, phosphorylated sugar (glucose-6-phosphate, fructose-6-phosphate or glucose-1-phosphate) decreased by 32.2% (P = 0.004) in the overloaded muscle in vivo while increasing in the IGF-1-stimulated myotubes in vitro. The systematic review revealed that 10 metabolites linked to muscle hypertrophy were directly associated with glycolysis and its interconnected anabolic pathways. We demonstrated that labelled carbon from [U-13C6]glucose is increasingly incorporated by ~13% (P = 0.001) into the non-essential amino acids in hypertrophying myotubes, which is accompanied by an increased depletion of media serine (P = 0.006). The inhibition of Phgdh suppressed muscle protein synthesis in growing myotubes by 58.1% (P < 0.001), highlighting the importance of the serine synthesis pathway for maintaining muscle size. Utilizing data from the UK Biobank (n = 450 243), we then discerned genetic variations linked to the serine synthesis pathway (PHGDH and PSPH) and to its downstream enzyme (SHMT1), revealing their association with appendicular lean mass in humans (P < 5.0e-8). CONCLUSIONS: Understanding the mechanisms that regulate skeletal muscle mass will help in developing effective treatments for muscle weakness. Our results provide evidence for the metabolic rewiring of glycolytic intermediates into anabolic pathways during muscle growth, such as in serine synthesis.

Genetic and genomic analyses of musculoskeletal differences between BEH and BEL strains.

Berlin high (BEH) and Berlin low (BEL) strains selected for divergent growth differ threefold in body weight. We aimed at examining muscle mass, which is a major contributor to body weight, by exploring morphological characteristics of the soleus muscle (fiber number and cross sectional area; CSA), by analyzing the transcriptome of the gastrocnemius and by initiating quantitative trait locus (QTL) mapping. BEH muscles were four to eight times larger than those of BEL. In substrain BEH+/+, mutant myostatin was replaced with a wild-type allele; however, BEH+/+muscles still were two to four times larger compared with BEL. BEH soleus muscle fibers were two times more numerous (P < 0.0001) and CSA was two times larger (P < 0.0001) compared with BEL. In addition, soleus femoral attachment anomaly (SFAA) was observed in all BEL mice. One significant (Chr 1) and four suggestive (Chr 3, 4, 6, and 9) muscle weight QTLs were mapped in a 21-day-old F2 intercross (n = 296) between BEH and BEL strains. The frequency of SFAA incidence in the F2 and in the backcross to BEL strain (BCL) suggested the presence of more than one causative gene. Two suggestive SFAA QTLs were mapped in BCL; however, their peak markers were not associated with the phenotype in F2. RNA-Seq analysis revealed 2,148 differentially expressed (P < 0.1) genes and 45,673 single nucleotide polymorphisms and >2,000 indels between BEH+/+ and BEL males. In conclusion, contrasting muscle traits and genomic and gene expression differences between BEH and BEL strains provide a promising model for the search for genes involved in muscle growth and musculoskeletal morphogenesis.

Effects of Diet Macronutrient Composition on Weight Loss during Caloric Restriction and Subsequent Weight Regain during Refeeding in Aging Mice.

Caloric restriction (CR) induces weight loss, but is associated with rapid weight regain upon return to ad libitum feeding. Our aim was to investigate effects of the macronutrient composition of the diet on weight loss and regain in elderly mice. Males, 18 months old, of the C57BL/6J strain were subjected to 4-week 30% CR followed by 4 weeks of ad libitum refeeding on either high-carb (HC), high-fat (HF) or high-protein (HP) diets (n = 22 each). Mice (n = 11) fed a chow diet ad libitum served as a control group (CON). Body mass and food intake were monitored daily. Twenty-four-hour indirect calorimetry was used to assess energy expenditure and substrate oxidation. Muscle and fat mass were evaluated with dissection of the tissues. Serum leptin and ghrelin levels were also measured. CR-induced weight loss did not differ between the diets. Weight regain was particularly fast for HF as mice overshot their initial weight by 12.8 ± 5.7% after 4-week refeeding when HC and HP mice reached the weight of the CON group. Weight regain strongly correlated with energy intake across the groups. The respiratory exchange ratio was lower in HF mice (0.81 ± 0.03) compared to HC (0.94 ± 0.06, p < 0.001), HP (0.89 ± 0.04, p < 0.001) and CON mice (0.91 ± 0.06, p < 0.01) during the refeeding. Serum leptin levels were higher in HF mice (1.03 ± 0.50 ng/mL) compared to HC (0.46 ± 0.14, p < 0.001), HP (0.63 ± 0.28, p < 0.05) or CON mice (0.41 ± 0.14, p < 0.001). Thus, CR induces similar weight loss in aging mice irrespective of the diet's macronutrient composition. An HF diet leads to excessive energy intake and pronounced gain in body fat in spite of increased fat oxidation and serum leptin during the refeeding after CR.

Stanniocalcin-2 inhibits skeletal muscle growth and is upregulated in functional overload-induced hypertrophy.

AIMS: Stanniocalcin-2 (STC2) has recently been implicated in human muscle mass variability by genetic analysis. Biochemically, STC2 inhibits the proteolytic activity of the metalloproteinase PAPP-A, which promotes muscle growth by upregulating the insulin-like growth factor (IGF) axis. The aim was to examine if STC2 affects skeletal muscle mass and to assess how the IGF axis mediates muscle hypertrophy induced by functional overload. METHODS: We compared muscle mass and muscle fiber morphology between Stc2-/- (n = 21) and wild-type (n = 15) mice. We then quantified IGF1, IGF2, IGF binding proteins -4 and -5 (IGFBP-4, IGFBP-5), PAPP-A and STC2 in plantaris muscles of wild-type mice subjected to 4-week unilateral overload (n = 14). RESULTS: Stc2-/- mice showed up to 10% larger muscle mass compared with wild-type mice. This increase was mediated by greater cross-sectional area of muscle fibers. Overload increased plantaris mass and components of the IGF axis, including quantities of IGF1 (by 2.41-fold, p = 0.0117), IGF2 (1.70-fold, p = 0.0461), IGFBP-4 (1.48-fold, p = 0.0268), PAPP-A (1.30-fold, p = 0.0154) and STC2 (1.28-fold, p = 0.019). CONCLUSION: Here we provide evidence that STC2 is an inhibitor of muscle growth upregulated, along with other components of the IGF axis, during overload-induced muscle hypertrophy.

Resolving candidate genes of mouse skeletal muscle QTL via RNA-Seq and expression network analyses.

BACKGROUND: We have recently identified a number of Quantitative Trait Loci (QTL) contributing to the 2-fold muscle weight difference between the LG/J and SM/J mouse strains and refined their confidence intervals. To facilitate nomination of the candidate genes responsible for these differences we examined the transcriptome of the tibialis anterior (TA) muscle of each strain by RNA-Seq. RESULTS: 13,726 genes were expressed in mouse skeletal muscle. Intersection of a set of 1061 differentially expressed transcripts with a mouse muscle Bayesian Network identified a coherent set of differentially expressed genes that we term the LG/J and SM/J Regulatory Network (LSRN). The integration of the QTL, transcriptome and the network analyses identified eight key drivers of the LSRN (Kdr, Plbd1, Mgp, Fah, Prss23, 2310014F06Rik, Grtp1, Stk10) residing within five QTL regions, which were either polymorphic or differentially expressed between the two strains and are strong candidates for quantitative trait genes (QTGs) underlying muscle mass. The insight gained from network analysis including the ability to make testable predictions is illustrated by annotating the LSRN with knowledge-based signatures and showing that the SM/J state of the network corresponds to a more oxidative state. We validated this prediction by NADH tetrazolium reductase staining in the TA muscle revealing higher oxidative potential of the SM/J compared to the LG/J strain (p<0.03). CONCLUSION: Thus, integration of fine resolution QTL mapping, RNA-Seq transcriptome information and mouse muscle Bayesian Network analysis provides a novel and unbiased strategy for nomination of muscle QTGs.

High-intensity exercise attenuates postprandial lipaemia and markers of oxidative stress.

Regular exercise can reduce the risk of CVD (cardiovascular disease). Although moderate-intensity exercise can attenuate postprandial TAG (triacylglycerol), high-intensity intermittent exercise might be a more effective method to improve health. We compared the effects of high-intensity intermittent exercise and 30 min of brisk walking on postprandial TAG, soluble adhesion molecules and markers of oxidative stress. Nine men each completed three 2-day trials. On day 1, subjects rested (control), walked briskly for 30 min (walking) or performed 5×30 s maximal sprints (high-intensity). On day 2, subjects consumed a high-fat meal for breakfast and 3 h later for lunch. Blood samples were taken at various times and analysed for TAG, glucose, insulin, ICAM-1 (intracellular adhesion molecule-1), VCAM-1 (vascular adhesion molecule-1), TBARS (thiobarbituric acid- reactive substances), protein carbonyls and ß-hydroxybutyrate. On day 2 of the high-intensity trial, there was a lower (P<0.05) incremental TAG AUC (area under the curve; 6.42±2.24 mmol/l per 7 h) compared with the control trial (9.68±4.77 mmol/l per 7 h) with no differences during day 2 of the walking trial (8.98±2.84 mmol/l per 7 h). A trend (P=0.056) for a reduced total TAG AUC was also seen during the high-intensity trial (14.13±2.83 mmol/l per 7 h) compared with control (17.18±3.92 mmol/l per 7 h), walking showed no difference (16.33±3.51 mmol/l per 7 h). On day 2 of the high-intensity trial plasma TBARS and protein carbonyls were also reduced (P<0.05) when compared with the control and walking trials. In conclusion, high-intensity intermittent exercise attenuates postprandial TAG and markers of oxidative stress after the consumption of a high-fat meal.

A longitudinal MRI study of muscle atrophy during lower leg immobilization following ankle fracture.

PURPOSE: To investigate MRI biomarkers of muscle atrophy during cast immobilization of the lower leg. MATERIALS AND METHODS: Eighteen patients (8 male, 10 female), who had one lower leg immobilized in a cast, underwent 3.0 Tesla (T) MR imaging 5, 8, 15, 29, and 43 days after casting. Measurements were made on both lower legs of total muscle volume. Cross-sectional area (CSA), fractional water content, and T(2) were measured in tibialis anterior (TA), gastrocnemius medialis (GM) and lateralis (GL) and soleus (SOL). Fiber pennation angle was measured in GM. RESULTS: Total muscle volume decreased by 17% (P < 0.001) over the 6 weeks of immobilization. The greatest loss in CSA (mean[SD]) was seen in GM (-23.3(8.7)%), followed by SOL (-19.0(9.8)%), GL (-17.1(6.5)%), and TA (-10.7(5.9)%). Significant reductions of CSA were also detectable in the contra-lateral leg. T(2) increased in all muscles: TA 27.0(2.5) ms to 29.6(2.8) ms (P < 0.001), GM 34.6(2.9) ms to 39.8(5.4) ms (P < 0.001) and SOL 34.4 (2.9) ms to 44.9(5.9) ms (P < 0.001). Small reductions were found in fractional water content. Pennation angle decreased in the cast leg (P < 0.001). CONCLUSION: Quantitative MR imaging can detect and monitor progressive biochemical and biophysical changes in muscle during immobilization.

Caloric Restriction per se Rather Than Dietary Macronutrient Distribution Plays a Primary Role in Metabolic Health and Body Composition Improvements in Obese Mice.

Caloric restriction (CR) is of key importance in combating obesity and its associated diseases. We aimed to examine effects of dietary macronutrient distribution on weight loss and metabolic health in obese mice exposed to CR. Male C57BL/6J mice underwent diet-induced obesity for 18 weeks. Thereafter mice were exposed to a 6-week CR for up to 40% on either low-fat diet (LFD; 20, 60, 20% kcal from protein, carbohydrate, fat), low-carb diet (LCD; 20, 20, 60% kcal, respectively) or high-pro diet (HPD; 35, 35, 30% kcal, respectively) (n = 16 each). Ten mice on the obesogenic diet served as age-matched controls. Body composition was evaluated by tissue dissections. Glucose tolerance, bloods lipids and energy metabolism were measured. CR-induced weight loss was similar for LFD and LCD while HPD was associated with a greater weight loss than LCD. The diet groups did not differ from obese controls in hindlimb muscle mass, but showed a substantial decrease in body fat without differences between them. Glucose tolerance and blood total cholesterol were weight-loss dependent and mostly improved in LFD and HPD groups during CR. Blood triacylglycerol was lowered only in LCD group compared to obese controls. Thus, CR rather than macronutrient distribution in the diet plays the major role for improvements in body composition and glucose control in obese mice. Low-carbohydrate-high-fat diet more successfully reduces triacylglycerol but not cholesterol levels compared to isocaloric high-carbohydrate-low-fat weight loss diets.

Effects of fasting on skeletal muscles and body fat of adult and old C57BL/6J mice.

Fasting improves metabolic health, but is also associated with loss of lean body mass. We investigated if old mice are less resistant to fasting-induce muscle wasting than adult mice. We compared changes in skeletal muscles and fat distribution in C57BL/6J mice subjected to 48-hour fasting at adult (6-month old) or old (24-month old) age. Old mice lost less weight (11.9 ± 1.5 vs 16.9 ± 2.8%, p < 0.001) and showed less (p < 0.01) pronounced muscle wasting than adult mice. Extensor digitorum longus (EDL) muscle force decreased only in adult mice after fasting. Serum IGF-1 levels were higher (p < 0.01) and showed greater (p < 0.01) decline in adult mice compared to old mice. Phosphorylation of 4EBP1 was reduced in the gastrocnemius muscles of adult mice only. Energy expenditure was slower in old mice and showed smaller fasting-induced decline than in adult mice when adjusted for variations in physical activity. There was a loss of fat mass in both age groups, but it was more pronounced in adult mice than old mice. Our results suggest that ageing-related decrease in metabolic rate protects old mice from skeletal muscle wasting during fasting.

H55N polymorphism as a likely cause of variation in citrate synthase activity of mouse skeletal muscle.

Citrate synthase (CS) is an enzyme of the Krebs cycle that plays a key role in mitochondrial metabolism. The aim of this study was to investigate the mechanisms underlying low activity of citrate synthase (CS) in A/J mice compared with other inbred strains of mice. Enzyme activity, protein content, and mRNA levels of CS were studied in the quadriceps muscles of A/J, BALB/cByJ, C57BL/6J, C3H/HeJ, DBA/2J, and PWD/PhJ strains of mice. Cytochrome c protein content was also measured. The results of the study indicate that A/J mice have a 50-65% reduction in CS activity compared with other strains despite similar levels of Cs mRNA and lack of differences in CS and cytochrome c protein content. CS from A/J mice also showed lower Michaelis constant (K(m)) for both acetyl CoA and oxaloacetate compared with the other strains of mice. In silico analysis of the genomic sequence identified a nonsynonymous single nucleotide polymorphism (SNP) (rs29358506, H55N) in Cs gene occurring near the site of the protein interacting with acetyl CoA. Allelic variants of the polymorphism segregated with the catalytic properties of CS enzyme among the strains. In summary, H55N polymorphism in Cs could be the underlying cause of low CS activity and its high affinity for substrates in A/J mice compared with other strains. This SNP might also play a role in resistance to obesity of A/J mice.

Yap is a novel regulator of C2C12 myogenesis.

The expression, regulation and function of mammalian Hippo pathway members in skeletal muscle is largely unknown. The aim of this study was thus to test the hypothesis that core members of the mammalian Hippo pathway are expressed in skeletal muscle and that the transcriptional co-factor Yap, a core member of the Hippo pathway, regulates C2C12 myogenesis. We found that the major components of the mammalian Hippo pathway including Yap are all expressed in skeletal muscles, C2C12 myoblasts and myotubes. In C2C12 myoblasts, Yap Ser127 phosphorylation is low and Yap localises to nuclei. Upon differentiation, Yap Ser127 phosphorylation increases approximately 20-fold and Yap translocates from the nucleus to the cytosol. To test whether the observed increase of Yap Ser127 phosphorylation is required for differentiation we overexpressed hYAP1 S127A, a mutant that can not be phosphorylated at Ser127, in C2C12 myoblasts. We found that overexpression of hYAP S127A prevented myotube formation, whereas the overexpression of wildtype hYAP1 or empty vector had no effect. In addition, more hYAP1 S127A overexpressing cells progressed through the S phase of the cell cycle and the expression of MRFs (myogenin, Myf5), Mef2c and cell cycle regulators (p21, cyclin D1) was significantly changed when compared to wildtype hYAP1 and empty vector overexpressing cells. This data suggests that the phosphorylation of Yap at Ser127 leads to a changed expression of MRFs and cell cycle regulators and is required for C2C12 myoblasts to differentiate into myotubes.

SB431542 treatment promotes the hypertrophy of skeletal muscle fibers but decreases specific force.

The small molecule inhibitor SB431542 inhibits activin type I receptors. The muscle growth-inhibitor myostatin binds to and signals via these receptors. The aim of this study was to test the hypothesis that SB431542 can inhibit myostatin-related Smad signaling and induce muscle growth in cultured C2C12 myotubes and increase growth and specific force in cultured Xenopus muscle fibers. The effect of SB431542 was assessed in vitro on C2C12 myotubes and ex vivo using mature Xenopus muscle fibers. SB431542 treatment reduced myostatin-induced C-terminal Smad2 phosphorylation and resulted in the formation of enlarged myotubes. However myogenin expression was unchanged, while p70 S6k phosphorylation at Thr389, total myosin heavy chain, and the rate of protein synthesis were all reduced. Mature Xenopus muscle fibers that were treated with SB431542 had a higher fiber cross-sectional area but decreased specific force production than control. SB431542 can initially antagonize myostatin signaling, but long-term unexpected signaling effects occur. Muscle fibers hypertrophy, but their specific force decreases compared to control.

Effects of ten-week 30% caloric restriction on metabolic health and skeletal muscles of adult and old C57BL/6J mice.

Caloric restriction (CR) can improve health, but the benefits are age-dependant. We studied effects of ten-week 30 % CR on skeletal muscles of adult (7-month old) and old (24-month old) C57BL/6 J mice. Old mice were heavier than adult mice (36.1 ± 4.0 g versus 32.9 ± 2.3 g, p < 0.05, respectively), but lost more weight (34.7 ± 6.0 % versus 23.9 ± 3.3 %, p < 0.001, respectively) during CR. Old mice did not differ from adult mice in extent of hind-limb muscle wasting or improvement in glucose tolerance after CR. Ageing and CR had an additive effect on increase in percentage of type 1 fibres in the soleus (SOL) muscle. CR was associated with greater atrophy of fast-twitch extensor digitorum longus (EDL) compared to slow-twitch SOL muscle. Old mice showed reduced gene expression of lysosomal markers, p62 and LC3B, while CR tended to upregulate the proteolysis genes. CR was also associated with increase in specific force of EDL muscle, but did not affect it in SOL muscle. In summary, ten-week CR induces only limited improvements in skeletal muscle function, but leads to significant muscle wasting and weakness in both adult and old mice.

Hypocaloric Low-Carbohydrate and Low-Fat Diets with Fixed Protein Lead to Similar Health Outcomes in Obese Mice.

OBJECTIVE: It is controversial whether low-carbohydrate diets are better suited for weight control and metabolic health than high-carbohydrate diets. This study examined whether these diets induce different improvements in body composition and glucose tolerance in obese mice during caloric restriction (CR). METHODS: Male C57BL/6J mice were fed an obesogenic diet ad libitum for 18 weeks and then subjected to 6-week progressive CR of up to 40%, using either a low-fat or low-carbohydrate diet with equal protein content. Mice fed a regular chow diet ad libitum served as controls. Body mass, hindlimb muscle mass, fat mass, energy expenditure, and glucose tolerance were compared between the groups. RESULTS: Initially low-fat and low-carbohydrate groups had similar body mass, which was 30% greater compared with controls. CR induced similar weight loss in low-fat and low-carbohydrate groups. This weight loss was mainly due to fat loss in both groups. Energy expenditure of freely moving mice did not differ between the groups. Glucose tolerance improved compared with the values before CR and in controls but did not differ between the diets. CONCLUSIONS: Dietary carbohydrate or fat content does not affect improvements in body composition and metabolic health in obese mice exposed to CR with fixed energy and protein intake.

The effect of interleukin-6 and the interleukin-6 receptor on glucose transport in mouse skeletal muscle.

Exercise results in an increase in interleukin-6 (IL-6), its receptor (IL-6R) and skeletal muscle glucose transport. Interleukin-6 has been found to have equivocal effects on glucose transport, with no studies, to our knowledge, investigating any potential role of IL-6R. In the present study, we hypothesized that a combined preparation of IL-6 and soluble IL-6R (sIL-6R) would stimulate glucose transport. Mouse soleus muscles were incubated with physiological and supraphysiological concentrations of IL-6 and a combination of IL-6 and sIL-6R. Total and phosphorylated AMP-activated protein kinase (AMPK) and Protein Kinase B (PKB/Akt) were also measured by Western blotting. Exposure to both physiological (80 pg ml(-1)) and supraphysiological IL-6 (120 ng ml(-1)) had no effect on glucose transport. At physiological levels, exposure to a combination of IL-6 and sIL-6R (32 ng ml(-1)) resulted in a 1.4-fold increase (P < 0.05) in basal glucose transport with no change to the phosphorylation of AMPK. Exposure to supraphysiological levels of IL-6 and sIL-6R (120 ng ml(-1)) resulted in an approximately twofold increase (P < 0.05) in basal glucose transport and an increase (P < 0.05) in AMPK phosphorylation. No effect of IL-6 or sIL-6R was observed on insulin-stimulated glucose transport. These findings demonstrate that, while IL-6 alone does not stimulate glucose transport in mouse soleus muscle, when sIL-6R is introduced glucose transport is directly stimulated, partly through AMPK-dependent signalling.

Signal transduction pathways that regulate muscle growth.

Progressive high-resistance exercise with 8-12 repetitions per set to near failure for beginners and 1-12 repetitions for athletes will increase muscle protein synthesis for up to 72 h; approx. 20 g of protein, especially when ingested directly after exercise, will promote high growth by elevating protein synthesis above breakdown. Muscle growth is regulated by signal transduction pathways that sense and compute local and systemic signals and regulate various cellular functions. The main signalling mechanisms are the phosphorylation of serine, threonine and tyrosine residues by kinases and their dephosphorylation by phosphatases. Muscle growth is stimulated by the mTOR (mammalian target of rapamycin) system, which senses (i) IGF-1 (insulin-like growth factor 1)/MGF (mechano-growth factor)/insulin and/or (ii) mechanical signals, (iii) amino acids and (iv) the energetic state of the muscle, and regulates protein synthesis accordingly. The action of the mTOR system is opposed by myostatin-Smad signalling which inhibits muscle growth via gene transcription.