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.

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.

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.

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.

Myostatin dysfunction is associated with lower physical activity and reduced improvements in glucose tolerance in response to caloric restriction in Berlin high mice.

Myostatin is an inhibitor of skeletal muscle growth and might be involved in adaptations to caloric restriction (CR). We compared responses to 12-week 30% CR in male mice of Berlin high strain with myostatin dysfunction (BEH) and wild-type myostatin (BEH+/+). BEH mice were heavier than BEH+/+ mice (58.8 ±â€¯2.0 versus 53.1 ±â€¯2.7 g, p < 0.001), had 1.8-fold greater hind limb muscle mass and were less (p < 0.05) physically active when fed ad libitum. After CR, BEH and BEH+/+ strains experienced similar weight loss (24.7 ±â€¯5.7 versus 20.6 ±â€¯6.5%, p > 0.05, respectively) and decreases (p < 0.001) in plasma IGF-1 and total cholesterol, but loss of hind limb muscle mass was greater (p < 0.001) in BEH mice than BEH+/+ mice. BEH mice had better (p < 0.001) glucose tolerance and showed smaller (p < 0.05) improvements of it than BEH+/+ mice after CR (1038.2 ±â€¯174.7 versus 744.4 ±â€¯95.8 glucose mM× 120 min, p < 0.01 for BEH; 1365.8 ±â€¯218.5 versus 831.5 ±â€¯134.4 glucose mM ×120 min, p < 0.001, for BEH+/+, respectively). In summary, myostatin dysfunction is associated with muscle hypertrophy and high glucose tolerance, but greater muscle wasting and smaller improvements in glucose tolerance in response to CR.

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.

Forced Running Endurance Is Influenced by Gene(s) on Mouse Chromosome 10.

Phenotypic diversity between laboratory mouse strains provides a model for studying the underlying genetic mechanisms. The A/J strain performs poorly in various endurance exercise models. The aim of the study was to test if endurance capacity and contractility of the fast- and slow-twitch muscles are affected by the genes on mouse chromosome 10. The C57BL/6J (B6) strain and C57BL/6J-Chr 10A/J/NaJ (B6.A10) consomic strain which carries the A/J chromosome 10 on a B6 strain background were compared. The B6.A10 mice compared to B6 were larger in body weight (p < 0.02): 27.2 ± 1.9 vs. 23.8 ± 2.7 and 23.4 ± 1.9 vs. 22.9 ± 2.3 g, for males and females, respectively, and in male soleus weight (p < 0.02): 9.7 ± 0.4 vs. 8.6 ± 0.9 mg. In the forced running test the B6.A10 mice completed only 64% of the B6 covered distance (p < 0.0001). However, there was no difference in voluntary wheel running (p = 0.6) or in fatigability of isolated soleus (p = 0.24) or extensor digitorum longus (EDL, p = 0.7) muscles. We conclude that chromosome 10 of the A/J strain contributes to reduced endurance performance. We also discuss physiological mechanisms and methodological aspects relevant to interpretation of these findings.