The myofibrillar protein synthetic response to the ingestion of 35 g casein is greater than after an equal amount of wheat protein. Ingesting a larger amount of wheat protein (i.e., 60 g) substantially increases myofibrillar protein synthesis rates in healthy older men. This trial was registered at clinicaltrials.gov as NCT01952639.

Background: Anemia is a major complication of end stage renal disease. The anemia is mainly the result of impaired formation of erythrocytes due to lack of erythropoietin and iron deficiency. Compelling evidence, however, points to the contribution of accelerated erythrocyte death, which decreases the life span of circulating erythrocytes. Erythrocytes may enter suicidal death or eryptosis, which is characterized by cell shrinkage and by cell membrane scrambling with phosphatidylserine-exposure at the erythrocyte surface. Triggers of eryptosis include increase of cytosolic Ca²⁺-activity ([Ca²⁺]_i). Erythrocytes could be sensitized to cytosolic Ca²⁺ by ceramide. In end stage renal disease, eryptosis may possibly be stimulated by uremic toxins. The present study explored, whether the uremic toxin acrolein could trigger eryptosis. Methods: Cell volume was estimated from forward scatter, phosphatidylserine-exposure from annexin-V-binding, hemolysis from hemoglobin release, [Ca²⁺]_i from Fluo3-fluorescence, and ceramide from fluorescent antibodies. Results: A 48 h exposure to acrolein (30 － 50 µM) did not significantly modify [Ca²⁺]_i but significantly decreased forward scatter and increased annexin-V-binding. Acrolein further triggered slight, but significant hemolysis and increased ceramide formation in erythrocytes. Acrolein (50 µM) induced annexin-V-binding was significantly blunted in the nominal absence of extracellular Ca²⁺. Acrolein augmented the annexin-V-binding following treatment with Ca²⁺ ionophore ionomycin (1 µM). Conclusion: Acrolein stimulates suicidal erythrocyte death or eryptosis, an effect at least in part due to stimulation of ceramide formation with subsequent sensitisation of the erythrocytes to cytosolic Ca²⁺.

Resistance-type exercise performed in the evening augments the overnight muscle protein synthetic response to presleep protein ingestion and allows more of the ingested protein-derived amino acids to be used for de novo myofibrillar protein synthesis during overnight sleep.

Muscle loss is a severe complication of many medical conditions such as cancer, cardiac failure, muscular dystrophies, and nerve damage. The contribution of myofibrillar protein synthesis (MPS) to the loss of muscle mass after nerve damage is not clear. Using deuterium oxide (D2O) labeling, we demonstrate that MPS is significantly increased in rat m. tibialis anterior (TA) compared to control (3.23 ± 0.72 [damaged] to 2.09 ± 0.26%∗day−1 [control]) after 4 weeks of nerve constriction injury. This is the case despite substantial loss of mass of the TA (350 ± 96 mg [damaged] to 946 ± 361 mg [control]). We also show that expression of regulatory proteins involved with MPS (p70s6k1: 2.4 ± 0.3 AU [damaged] to 1.8 ± 0.2 AU [control]) and muscle protein breakdown (MPB) (MAFbx: 5.3 ± 1.2 AU [damaged] to 1.4 ± 0.4 AU [control]) are increased in nerve damaged muscle. Furthermore, the expression of p70s6k1 correlates with MPS rates (r2 = 0.57). In conclusion, this study shows that severe muscle wasting following nerve damage is accompanied by increased as opposed to decreased MPS.

Background Desminopathy is a clinically heterogeneous muscle disease caused by over 60 different mutations in desmin. The most common mutation with a clinical phenotype in humans is an exchange of arginine to proline at position 350 of desmin leading to p.R350P. We created the first CRISPR-Cas9 engineered rat model for a muscle disease by mirroring the R350P mutation in humans. Methods Using CRISPR-Cas9 technology, Des c.1045-1046 (AGG > CCG) was introduced into exon 6 of the rat genome causing p.R349P. The genotype of each animal was confirmed via quantitative PCR. Six male rats with a mutation in desmin (n = 6) between the age of 120-150 days and an equal number of wild type littermates (n = 6) were used for experiments. Maximal plantar flexion force was measured in vivo and combined with the collection of muscle weights, immunoblotting, and histological analysis. In addition to the baseline phenotyping, we performed a synergist ablation study in the same animals. Results We found a difference in the number of central nuclei between desmin mutants (1 ± 0.4%) and wild type littermates (0.2 ± 0.1%; P < 0.05). While muscle weights did not differ, we found the levels of many structural proteins to be altered in mutant animals. Dystrophin and syntrophin were increased 54% and 45% in desmin mutants, respectively (P < 0.05). Dysferlin and Annexin A2, proteins associated with membrane repair, were increased twofold and 32%, respectively, in mutants (P < 0.05). Synergist ablation caused similar increases in muscle weight between mutant and wild type animals, but changes in fibre diameter revealed that fibre hypertrophy in desmin mutants was hampered compared with wild type animals (P < 0.05). Conclusions We created a novel animal model for desminopathy that will be a useful tool in furthering our understanding of the disease. While mutant animals at an age corresponding to a preclinical age in humans show no macroscopic differences, microscopic and molecular changes are already present. Future studies should aim to further decipher those biological changes that precede the clinical progression of disease and test therapeutic approaches to delay disease progression.

Perturbations in skeletal muscle metabolism have been reported for a variety of neuromuscular diseases. However, the role of metabolism after constriction injury to a nerve and the associated muscle atrophy is unclear. We have analyzed rat tibialis anterior (TA) four weeks after unilateral constriction injury to the sciatic nerve (DMG) and in the contralateral control leg (CTRL) (n = 7) to investigate changes of the metabolome, immunohistochemistry and protein levels. Untargeted metabolomics identified 79 polar metabolites, 27 of which were significantly altered in DMG compared to CTRL. Glucose concentrations were increased 2.6-fold in DMG, while glucose 6-phosphate (G6-P) was unchanged. Intermediates of the polyol pathway were increased in DMG, particularly fructose (1.7fold). GLUT4 localization was scattered as opposed to clearly at the sarcolemma. Despite the altered localization, we found GLUT4 protein levels to be increased 7.8-fold while GLUT1 was decreased 1.7fold in nerve damaged TA. PFK1 and GS levels were both decreased 2.1-fold, indicating an inability of glycolysis and glycogen synthesis to process glucose at sufficient rates. In conclusion, chronic nerve constriction causes increased GLUT4 levels in conjunction with decreased glycolytic activity and glycogen storage in skeletal muscle, resulting in accumulation of intramuscular glucose and polyol pathway intermediates. Skeletal muscle atrophy is a pathological condition associated with many diseases. While protein metabolism is thought to be the main regulator of muscle size, many situations of atrophy and neuromuscular diseases are accompanied by changes to substrate metabolism as well. Critical illness myopathy (CIM) is a condition for which disturbances of glucose metabolism have been reported by our laboratory and others 1. Specifically, we have found the primary glucose transporter GLUT4 to be insufficiently translocated in CIM patients, resulting in decreased glucose supply and reduced AMPK activity 2. Besides CIM, a number of other neuromuscular disorders have been found to be show signs of changes to glucose metabolism in skeletal muscle such as amyotrophic lateral sclerosis (ALS), Charcot-Marie-Tooth neuropathy (CMT) or spinal muscular atrophy (SMA) 3-5. For example, it has been found that concentrations of glucose as well as fructose are increased in skeletal muscle samples of ALS patients, often accompanied by an early onset of insulin resistance 6. Early research in respect to nerve damage and glucose metabolism has reported that denervation is followed by insulin resistance, reduced glucose transport into the muscle, less glucose abundance and transiently decreased GLUT4 levels 7-9. A more recent study in mice found that despite decreased GLUT4 mRNA abundance, long term denervation was associated with increased GLUT4 protein levels potentially regulated by increased Akt activity 10. The same study found that glucose uptake in

Cachexia is a debilitating comorbidity affecting many lung cancer patients. We have previously found that cachectic mice with lung cancer have reduced serum ketone body levels due to low PPARα activity in the liver. Restoring hepatic PPARα activity with fenofibrate increased circulating ketones and delayed muscle and white adipose tissue wasting. We hypothesized that the loss of circulating ketones plays a pathophysiologic role in cachexia and performed two dietary intervention studies to test this hypothesis. In the first study, male and female mice were randomized to consume either a very low carbohydrate, ketogenic diet (KD) or normal chow (NC) after undergoing tumor induction. The KD successfully restored serum ketone levels and decreased blood glucose in cachectic mice but did not improve body weight maintenance or survival. In fact, there was a trend for the KD to worsen survival in male but not in female mice. In the second study, we compounded a ketone ester supplement into the NC diet (KE) and randomized tumor-bearing mice to KE or NC after tumor induction. We confirmed that KE was able to acutely and chronically increase ketone body abundance in the serum compared to NC. However, the restoration of ketones in the circulation was not able to improve body weight maintenance or survival in male or female mice with lung cancer. Finally, we investigated PPARα activity in the liver of mice fed KE and NC and found that animals fed a ketone ester supplement showed a significant increase in mRNA expression of several PPARα targets. These data negate our initial hypothesis and suggest that restoring ketone body availability in the circulation of mice with lung cancer does not alter cachexia development or improve survival, despite increasing hepatic PPARα activity.

Cannabidiol (CBD) has proven clinical benefits in the treatment of seizures, inflammation, and pain. The recent legalization of CBD in many countries has caused increased interest in the drug as an over-the-counter treatment for athletes looking to improve recovery. However, no data on the effects of CBD on the adaptive response to exercise in muscle are available. To address this gap, we eccentrically loaded the tibialis anterior muscle of 14 rats, injected them with a vehicle (n = 7) or 100 mg/kg CBD (n = 7), and measured markers of injury, inflammation, anabolic signaling, and autophagy 18 hr later. Pro-inflammatory signaling through nuclear factor kappa B (NF-kB) (Ser536) increased with loading in both groups; however, the effect was significantly greater (36%) in the vehicle group (p < .05). Simultaneously, anabolic signaling through ribosomal protein S6 kinase beta-1 (S6K1) (Thr389) increased after eccentric contractions in both groups with no difference between vehicle and CBD (p = .66). The ribosomal protein S6 phosphorylation (240/244) increased with stimulation (p < .001) and tended to be higher in the CBD group (p = .09). The ubiquitin-binding protein p62 levels were not modulated by stimulation (p = .6), but they were 46% greater in the CBD compared with the vehicle group (p = .01). Although liver weight did not differ between the groups (p = .99) and levels of proteins associated with stress were similar, we did observe serious side effects in one animal. In conclusion, an acute dose of CBD decreased pro-inflammatory signaling in the tibialis anterior without blunting the anabolic response to exercise in rats. Future research should determine whether these effects translate to improved recovery without altering adaptation in humans.