Dose-Response of Myofibrillar Protein Synthesis To Ingested Whey Protein During Energy Restriction in Overweight Postmenopausal Women: A Randomized, Controlled Trial.

BACKGROUND: Diet-induced weight loss is associated with a decline in lean body mass, as mediated by an impaired response of muscle protein synthesis (MPS). The dose-response of MPS to ingested protein, with or without resistance exercise, is well characterized during energy balance but limited data exist under conditions of energy restriction in clinical populations. OBJECTIVE: To determine the dose-response of MPS to ingested whey protein following short-term diet-induced energy restriction in overweight, postmenopausal, women at rest and postexercise. DESIGN: Forty middle-aged (58.6±0.4 y), overweight (BMI: 28.6±0.4), postmenopausal women were randomly assigned to 1 of 4 groups: Three groups underwent 5 d of energy restriction (∼800 kcal/d). On day 6, participants performed a unilateral leg resistance exercise bout before ingesting either a bolus of 15g (ERW15, n = 10), 35g (ERW35, n = 10) or 60g (ERW60, n = 10) of whey protein. The fourth group (n = 10) ingested a 35g whey protein bolus after 5 d of an energy balanced diet (EBW35, n = 10). Myofibrillar fractional synthetic rate (FSR) was calculated under basal, fed (FED) and postexercise (FED-EX) conditions by combining an L-[ring-13C6] phenylalanine tracer infusion with the collection of bilateral muscle biopsies. RESULTS: Myofibrillar FSR was greater in ERW35 (0.043±0.003%/h, P = 0.013) and ERW60 (0.042±0.003%/h, P = 0.026) than ERW15 (0.032 ± 0.003%/h), with no differences between ERW35 and ERW60 (P = 1.000). Myofibrillar FSR was greater in FED (0.044 ± 0.003%/h, P < 0.001) and FED-EX (0.048 ± 0.003%/h, P < 0.001) than BASAL (0.027 ± 0.003%/h), but no differences were detected between FED and FED-EX (P = 0.732) conditions. No differences in myofibrillar FSR were observed between EBW35 (0.042 ± 0.003%/h) and ERW35 (0.043 ± 0.003%/h, P = 0.744). CONCLUSION: A 35 g dose of whey protein, ingested with or without resistance exercise, is sufficient to stimulate a maximal acute response of MPS following short-term energy restriction in overweight, postmenopausal women, and thus may provide a per serving protein recommendation to mitigate muscle loss during a weight loss program. TRIAL REGISTRY: clinicaltrials.gov (ID: NCT03326284).

Effect of 1-year daily protein supplementation and physical exercise on muscle protein synthesis rate and muscle metabolome in healthy older Danes: a randomized controlled trial.

BACKGROUND: The skeletal muscle mass decreases with age and the responsiveness of aging muscles' protein synthesis rate (MPS) to protein intake seems to deteriorate. OBJECTIVE: This study investigated the impact of 12 months of protein supplementation with or without physical exercise training on the basal and postprandial MPS and the skeletal muscle metabolome of healthy older Danes (> 65 years, 29 females/37 males). METHODS: Subjects were randomized to follow one of five intervention groups: (1) carbohydrate, (2) collagen protein, (3) whey protein, (4) home-based light resistance training with whey protein, and (5) center-based heavy-load resistance training with whey protein. Before and after the intervention, a tracer infusion trial was conducted to measure basal and postprandial MPS in response to intake of a cocktail consisting of 20 g whey hydrolysate + 10 g glucose. In addition, the skeletal muscle metabolome was measured using gas chromatography-mass spectrometry (GC-MS) at basal state and 4 h after the intake of the cocktail. RESULTS: One year of daily protein or carbohydrate supplementation did not alter the basal and protein-stimulated postprandial muscle protein synthesis rate or the muscle metabolome of healthy older Danes. Basal MPS (%/h) at baseline for all subjects were 0.0034 ± 0,011 (mean ± SD). In contrast to previous studies, no difference was observed in basal MPS between males and females (p = 0.75). With the developed untargeted GC-MS methodology, it was possible to detect and tentatively annotate > 70 metabolites from the human skeletal muscle samples. CONCLUSION: One year of protein supplementation in comparison to an isocaloric-control supplement seems to affect neither the MPS at basal or postprandial state nor the skeletal muscle metabolome. CLINICAL TRIAL REGISTRY: Number: NCT02115698, clinicaltrials.gov/ct2/show/NCT02115698.

Dietary protein requirements and recommendations for healthy older adults: a critical narrative review of the scientific evidence.

Adequate protein intake is essential for the maintenance of whole-body protein mass. Different methodological approaches are used to substantiate the evidence for the current protein recommendations, and it is continuously debated whether older adults require more protein to counteract the age-dependent loss of muscle mass, sarcopenia. Thus, the purpose of this critical narrative review is to outline and discuss differences in the approaches and methodologies assessing the protein requirements and, hence, resulting in controversies in current protein recommendations for healthy older adults. Through a literature search, this narrative review first summarises the historical development of the Food and Agriculture Organization/World Health Organization/United Nations University setting of protein requirements and recommendations for healthy older adults. Hereafter, we describe the various types of studies (epidemiological studies and protein turnover kinetic measurements) and applied methodological approaches founding the basis and the different recommendations with focus on healthy older adults. Finally, we discuss important factors to be considered in future studies to obtain evidence for international agreement on protein requirements and recommendations for healthy older adults. We conclude by proposing future directions to determine 'true' protein requirements and recommendations for healthy older adults.

Changes in Muscle Mass and Strength During Follow-Up After One-Year Resistance Training Interventions in Older Adults.

ABSTRACT: Mertz, KH, Reitelseder, S, Rasmussen, MA, Bülow, J, Højfeldt, G, Jensen, M, Hjulmand, M, Lindberg, J, Kramer, MU, Bechshøft, R, and Holm, L. Changes in muscle mass and strength during follow-up after one-year resistance training interventions in older adults. J Strength Cond Res 37(10): 2064-2070, 2023-The aim of this study was to investigate if home-based resistance training compared with center-based resistance training was associated with better preservation of muscle mass and strength in older individuals, 6 months after the interventions ended. One hundred four healthy older individuals (>65 years) who had completed 1 year of either home-based light-intensity training with daily whey protein supplementation (LITW), center-based heavy resistance training with whey protein supplementation (HRTW), or daily whey protein supplementation alone (WHEY) returned for follow-up measurement 6 months after the interventions. Measures of muscle mass, strength, and power were assessed at the end of intervention as well as at follow-up. Furthermore, we compared changes in these parameters between subjects who continued resistance training (≥1 weekly training session) during follow-up (CONT) with those who stopped (STOP). Resistance training continuation during follow-up did not differ between HRTW and LITW (41 vs. 41%, P = 1.0) but was higher for both groups compared with WHEY (18%, P = 0.04-0.05). However, no between-group differences were observed between LITW/HRTW/WHEY in changes in muscle mass, strength, or power during follow-up. STOP was associated with a poorer preservation of quadriceps cross-sectional area compared with CONT (-1.7 cm 2 [-0.4 to -3.0], P = 0.01, effect size: 0.79). No effect of training continuation was observed on changes in muscle strength and power. In conclusion, maintenance of muscle mass and strength is not superior after home-based resistance training compared with center-based training. However, training continuation seems crucial for the maintenance of muscle mass, irrespective of the training intervention.

Ubiquitin E3 ligase Atrogin-1 protein is regulated via the rapamycin-sensitive mTOR-S6K1 signaling pathway in C2C12 muscle cells.

Atrogin-1 and Muscle-specific RING finger protein 1 (MuRF1) are highly expressed in multiple conditions of skeletal muscle atrophy. The phosphoinositide 3-kinase (PI3K)/Akt/forkhead box (FoxO) signaling pathway is well known to regulate Atrogin-1 and MuRF1 gene expressions. However, Akt activation also activates the mechanistic target of rapamycin complex 1 (mTORC1), which induces skeletal muscle hypertrophy. Whether mTORC1-dependent signaling has a role in regulating Atrogin-1 and/or MuRF1 gene and protein expression is currently unclear. In this study, we showed that activation of insulin-mediated Akt signaling suppresses both Atrogin-1 and MuRF1 protein contents and that inhibition of Akt increases both Atrogin-1 and MuRF1 protein contents in C2C12 myotubes. Interestingly, inhibition of mTORC1 with a specific mTORC1 inhibitor, rapamycin, increased Atrogin-1, but not MuRF1, protein content. Furthermore, activation of AMP-activated protein kinase (AMPK), a negative regulator of the mTORC1 signaling pathway, also showed distinct time-dependent changes between Atrogin-1 and MuRF1 protein contents, suggesting differential regulatory mechanisms between Atrogin-1 and MuRF1 protein content. To further explore the downstream of mTORC1 signaling, we employed a specific S6K1 inhibitor, PF-4708671. We found that Atrogin-1 protein content was dose-dependently increased with PF-4708671 treatment, whereas MuRF1 protein content was decreased at 50 µM of PF-4708671 treatment. However, MuRF1 protein content was unexpectedly increased by PF-4708671 treatment for a longer period. Overall, our results indicate that Atrogin-1 and MuRF1 protein contents are regulated by different mechanisms, the downstream of Akt, and that Atrogin-1 protein content can be regulated by the rapamycin-sensitive mTOR-S6K1-dependent signaling pathway.

Human Blood Lipoprotein Predictions from 1H NMR Spectra: Protocol, Model Performances, and Cage of Covariance.

Lipoprotein subfractions are biomarkers for the early diagnosis of cardiovascular diseases. The reference method, ultracentrifugation, for measuring lipoproteins is time-consuming, and there is a need to develop a rapid method for cohort screenings. This study presents partial least-squares regression models developed using 1H nuclear magnetic resonance (NMR) spectra and concentrations of lipoproteins as measured by ultracentrifugation on 316 healthy Danes. This study explores, for the first time, different regions of the 1H NMR spectrum representing signals of molecules in lipoprotein particles and different lipid species to develop parsimonious, reliable, and optimal prediction models. A total of 65 lipoprotein main and subfractions were predictable with high accuracy, Q2 of >0.6, using an optimal spectral region (1.4-0.6 ppm) containing methylene and methyl signals from lipids. The models were subsequently tested on an independent cohort of 290 healthy Swedes with predicted and reference values matching by up to 85-95%. In addition, an open software tool was developed to predict lipoproteins concentrations in human blood from standardized 1H NMR spectral recordings.

Co-ingestion of cluster dextrin carbohydrate does not increase exogenous protein-derived amino acid release or myofibrillar protein synthesis following a whole-body resistance exercise in moderately trained younger males: a double-blinded randomized controlled crossover trial.

PURPOSE: This study investigates if co-ingestion of cluster dextrin (CDX) augments the appearance of intrinsically labeled meat protein hydrolysate-derived amino acid (D5-phenylalanine), Akt/mTORC1 signaling, and myofibrillar protein fractional synthetic rate (FSR). METHODS: Ten moderately trained healthy males (age: 21.5 ± 2.1 years, body mass: 75.7 ± 7.6 kg, body mass index (BMI): 22.9 ± 2.1 kg/m2) were included for a double-blinded randomized controlled crossover trial. Either 75 g of CDX or glucose (GLC) was given in conjunction with meat protein hydrolysate (0.6 g protein * FFM-1) following a whole-body resistance exercise. A primed-continuous intravenous infusion of L-[15N]-phenylalanine with serial muscle biopsies and venous blood sampling was performed. RESULTS: A time × group interaction effect was found for serum D5-phenylalanine enrichment (P < 0.01). Serum EAA and BCAA concentrations showed a main effect for group (P < 0.05). Tmax serum BCAA was greater in CDX as compared to GLC (P < 0.05). However, iAUC of all serum parameters did not differ between CDX and GLC (P > 0.05). Tmax serum EAA showed a trend towards a statistical significance favoring CDX over GLC. The phosphorylation of p70S6KThr389, rpS6Ser240/244, ERK1/2Thr202/Tyr204 was greater in CDX compared to GLC (P < 0.05). However, postprandial myofibrillar FSR did not differ between CDX and GLC (P = 0.17). CONCLUSION: In moderately trained younger males, co-ingestion of CDX with meat protein hydrolysate does not augment the postprandial amino acid availability or myofibrillar FSR as compared to co-ingestion of GLC during the recovery from a whole-body resistance exercise despite an increased intramuscular signaling. TRIAL REGISTRATION: ClinicalTrials.gov ID: NCT03303729 (registered on October 3, 2017).

A frameshift deletion in the GDF9 gene in Icelandic Loa prolific sheep.

Several genetic variants have been shown to affect the mean number of offspring in different sheep breeds. Here, we analyzed samples from Icelandic sheep with the aim of identifying the genetic cause of the Icelandic Loa phenotype using three previously identified prolificacy genes as candidates. We demonstrate that a 4-bp frameshift deletion positioned in the mature region of the GDF9 protein in the Loa animals is a likely causal mutation for the observed increase in prolificacy; however, sequencing showed that not all ewes with a high number of offspring carried the deletion, suggesting the presence of a second mutation segregating within this group of animals.

Recent advances in measuring and understanding the regulation of exercise-mediated protein degradation in skeletal muscle.

Skeletal muscle protein turnover plays a crucial role in controlling muscle mass and protein quality control, including sarcomeric (structural and contractile) proteins. Protein turnover is a dynamic and continual process of protein synthesis and degradation. The ubiquitin proteasome system (UPS) is a key degradative system for protein degradation and protein quality control in skeletal muscle. UPS-mediated protein quality control is known to be impaired in aging and diseases. Exercise is a well-recognized, nonpharmacological approach to promote muscle protein turnover rates. Over the past decades, we have acquired substantial knowledge of molecular mechanisms of muscle protein synthesis after exercise. However, there have been considerable gaps in the mechanisms of how muscle protein degradation is regulated at the molecular level. The main challenge to understand muscle protein degradation is due in part to the lack of solid stable isotope tracer methodology to measure muscle protein degradation rate. Understanding the mechanisms of UPS with the concomitant measurement of protein degradation rate in skeletal muscle will help identify novel therapeutic strategies to ameliorate impaired protein turnover and protein quality control in aging and diseases. Thus, the goal of this present review was to highlight how recent advances in the field may help improve our understanding of exercise-mediated protein degradation. We discuss 1) the emerging roles of protein phosphorylation and ubiquitylation modifications in regulating proteasome-mediated protein degradation after exercise and 2) methodological advances to measure in vivo myofibrillar protein degradation rate using stable isotope tracer methods.

Postprandial amino acid availability after intake of intact or hydrolyzed meat protein in a mixed meal in healthy elderly subjects: a randomized, single blind crossover trial.

The absorption of dietary proteins affects the anabolic response, among others protein synthesis. For elderly, optimal amino acid absorption is warranted to preserve the amino acid pool of the body, especially skeletal muscle proteins. Therefore, the aim of this study was to characterize if hydrolyzing meat protein (HMP) would improve the amino acid absorption after ingestion of meat compared to equal amounts of the same meat proteins but present in a different structure; steak or minced meat. With a crossover study design on 12 healthy older adults (> 65 years of age, BMI 18.5-30), the amino acid absorption kinetics were explored by ingesting 0.55 g protein/kg LBM as a mixed meal together with intrinsically [2H5]phenylalanine labeled meat proteins prepared as a STEAK, MINCED meat, or HMP. Plasma [2H5]phenylalanine enrichment as well as AA concentrations were measured by mass spectrometry from blood samples drawn during a 5-h postprandial period. After HMP ingestion, [2H5]phenylalanine was faster absorbed in the initial 2 h compared to STEAK and MINCED. The peak time in AA concentrations was faster in HMP compared to STEAK and MINCED. However, the peak AA concentrations were not different between STEAK, MINCED, and HMP. Although HMP showed to have the fastest initial amino acid appearance in older adults, the peak EAA concentrations were similar after ingesting meal with either STEAK, MINCED, or HMP in the 5-h postprandial period.

Postprandial muscle protein synthesis rate is unaffected by 20-day habituation to a high protein intake: a randomized controlled, crossover trial.

PURPOSE: During the last decade more researchers have argued in favor of an increased protein intake for older adults. However, there is a lack of knowledge on the long-term effects of conforming to such a high protein intake with regards to the basal and postprandial muscle protein turnover. The purpose of this study was to compare the postprandial synthesis response in muscle proteins, and the abundance of directly incorporated food-derived amino acids following habituation to high vs. recommended level of protein intake. METHODS: In a double blinded crossover intervention 11 older male participants (66.6 ± 1.7 years of age) were habituated for 20 days to a recommended protein (RP) intake (1.1 g protein/kg lean body mass (LBM)/day) and a high protein (HP) intake (> 2.1 g protein/kg LBM/day). Following each habituation period, intrinsically labelled proteins were ingested as part of a mixed meal to determine the incorporation of meal protein-derived amino acids into myofibrillar proteins. Furthermore, the myofibrillar fractional synthesis rate (FSR) and amino acid kinetics across the leg were determined using gold standard stable isotope tracer methodologies. RT qPCR was used to assess the expression of markers related to muscle proteinsynthesis and breakdown. RESULTS: No impact of habituation was observed on skeletal muscle amino acid or protein kinetics. However, the shunting of amino acids directly from artery to vein was on average 2.9 [Formula: see text]mol/min higher following habituation to HP compared to RP. CONCLUSIONS: In older males, habituation to a higher than the currently recommended protein intake did not demonstrate any adaptions in the muscle protein turnover or markers hereof when subjected to an intake of an identical mixed meal. CLINICAL TRIAL REGISTRY: Journal number NCT02587156, Clinicaltrials.org. Date of registration: October 27th, 2015.

Muscle protein breakdown is impaired during immobilization compared to during a subsequent retraining period in older men: no effect of anti-inflammatory medication.

Muscle inactivity reduces muscle protein synthesis (MPS), whereas a subsequent period of rehabilitation resistance training (retraining) increases MPS. However, less is known regarding muscle protein breakdown (MPB) during such conditions. Furthermore, nonsteroidal anti-inflammatory drugs (NSAIDs) may have a dampening effect on MPB during periods of inactivity in older individuals. Thus, we measured the average MPB, by use of the deuterated water methodology, during an immobilization period and a subsequent retraining period in older individuals with and without NSAID treatment. Eighteen men (60-80 years: range) were randomly assigned to ibuprofen (1200 mg/d, Ibu) or placebo (Plc). One lower limb was immobilized in a cast for 2 weeks and retrained for 2 weeks, and 2 × 20 g of whey protein was ingested daily during both periods. Besides MPB, the protein expression of different muscle degradation signaling molecules was investigated. MPB was lower during immobilization compared to retraining (p < 0.01). NSAID treatment did not affect the MPB rate during immobilization or retraining (p > 0.05). The protein expression of muscle degradation signaling molecules changed during the study intervention but were unaffected by NSAID treatment. The finding that MPB was lower during immobilization than during retraining indicates that an increased MPB may play an important role in the muscle protein remodeling processes taking place within the initial retraining period. Moreover, NSAID treatment did not significantly influence the MPB rate during 2 weeks of lower limb immobilization or during 2 weeks of subsequent retraining in older individuals.

Postprandial Nutrient Handling and Gastrointestinal Hormone Secretion After Roux-en-Y Gastric Bypass vs Sleeve Gastrectomy.

BACKGROUND & AIMS: Sleeve gastrectomy (SG) and Roux-en-Y gastric bypass (RYGB) induce substantial weight loss and improve glycemic control in patients with type 2 diabetes, but it is not clear whether these occur via the same mechanisms. We compared absorption rates of glucose and protein, as well as profiles of gastro-entero-pancreatic hormones, in patients who had undergone SG or RYGB vs controls. METHODS: We performed a cross-sectional study of 12 patients who had undergone sleeve gastrectomy, 12 patients who had undergone RYGB, and 12 individuals who had undergone neither surgery (controls), all in Denmark. Study participants were matched for body mass index, age, sex, and postoperative weight loss, and all had stable weights. They received continuous infusions of stable isotopes of glucose, glycerol, phenylalanine, tyrosine, and urea before and during a mixed meal containing labeled glucose and intrinsically phenylalanine-labeled caseinate. Blood samples were collected for 6 hours, at 10- to 60-minute intervals, and analyzed. RESULTS: The systemic appearance of ingested glucose was faster after RYGB and SG vs controls; the peak glucose appearance rate was 64% higher after RYGB, and 23% higher after SG (both P < .05); the peak phenylalanine appearance rate from ingested casein was 118% higher after RYGB (P < .01), but similar between patients who had undergone SG and controls. Larger, but more transient increases in levels of plasma glucose and amino acids were accompanied by higher secretion of insulin, glucagon-like peptide 1, peptide YY, and cholecystokinin after RYGB, whereas levels of ghrelin were lower after SG, compared with RYGB and controls. Total 6-hour oral recovery of ingested glucose and protein was comparable among groups. CONCLUSIONS: Postprandial glucose and protein absorption and gastro-entero-pancreatic hormone secretions differ after SG and RYGB. RYGB was characterized by accelerated absorption of glucose and amino acids, whereas protein metabolism after SG did not differ significantly from controls, suggesting that different mechanisms explain improved glycemic control and weight loss after these surgical procedures. ClinicalTrials.gov ID NCT03046186.

Even effect of milk protein and carbohydrate intake but no further effect of heavy resistance exercise on myofibrillar protein synthesis in older men.

PURPOSE: The responsiveness of older individuals' skeletal muscle to anabolic strategies may be impaired. However, direct comparisons within the same experimental setting are sparse. The aim of this study was to assess the resting and post-resistance exercise muscle protein synthesis rates in response to two types of milk protein and carbohydrate using a unilateral exercise leg model. METHODS: Twenty-seven older (69 ± 1 year, mean ± SE) men were randomly assigned one of three groups: Whey hydrolysate (WH), caseinate (CAS), or carbohydrate (CHO). By applying stable isotope tracer techniques (L-[15N]phenylalanine), the fasted-rested (basal) myofibrillar fractional synthesis rate (FSR) was measured. Hereafter, FSR was measured in the postprandial phase (0.45 g nutrient/kg LBM) in both legs, one rested (fed-rest) and one exercised (10 × 8 reps at 70% 1RM; fed-exercise). In addition, the activity of p70S6K and venous plasma insulin, phenylalanine, and leucine concentrations were measured. RESULTS: Insulin, phenylalanine, and leucine concentrations differed markedly after intake of the different study drinks. The basal FSR in WH, CAS, and CHO were 0.027 ± 0.003, 0.030 ± 0.003, and 0.030 ± 0.004%/h, the fed-rested FSR were 0.043 ± 0.004, 0.045 ± 0.003, and 0.035 ± 0.004%/h, and the fed-exercised FSR were 0.041 ± 0.004, 0.043 ± 0.004, and 0.034 ± 0.004%/h, respectively. No significant differences were observed at any state between the groups. Fed-rested- and fed-exercised FSR were higher than basal (P < 0.001). 3 h after exercise and feeding, no significant group differences were detected in the activity of p70S6K. CONCLUSIONS: Milk protein and carbohydrate supplementation stimulate myofibrillar protein synthesis in older men, with no further effect of heavy resistance exercise within 0-3 h post exercise.

Influence of between-limb asymmetry in muscle mass, strength, and power on functional capacity in healthy older adults.

PURPOSE: Numerous daily tasks such as walking and rising from a chair involve bilateral lower limb movements. During such tasks, lower extremity function (LEF) may be compromised among older adults. LEF may be further impaired due to high degrees of between-limb asymmetry. The present study investigated the prevalence of between-limb asymmetry in muscle mass, strength, and power in a cohort of healthy older adults and examined the influence of between-limb asymmetry on LEF. METHODS: Two hundred and eight healthy older adults (mean age 70.2 ± 3.9 years) were tested for LEF (400 m walking and 30-seconds chair stand). Furthermore, maximal isometric and dynamic knee extensor strength, leg extensor power, and lower limb lean tissue mass (LTM) were obtained unilaterally. RESULTS: Mean between-limb asymmetry in maximal muscle strength and power ranged between 10% and 13%, whereas LTM asymmetry was 3 ± 2.3%. Asymmetry in dynamic knee extensor strength was larger for women compared with men (15.0 ± 11.8% vs 11.1 ± 9.5%; P = .005) Leg strength and power were positively correlated with LEF (r2  = .43-.46, P < .001). The weakest leg was not a stronger predictor of LEF than the strongest leg. Between-limb asymmetry in LTM and isometric strength was negatively associated with LEF (LTM; r2  = .12, P = .005, isometric peak torque; r2  = 0.40, P = .03.) but dynamic strength and power were not. CONCLUSION: The present study supports the notion that in order to improve or maintain LEF, healthy older adults should participate in training interventions that increase muscle strength and power, whereas the effects of reducing between-limb asymmetry in these parameters might be of less importance.

Beta2 -adrenoceptor agonist salbutamol increases protein turnover rates and alters signalling in skeletal muscle after resistance exercise in young men.

KEY POINTS: Animal models have shown that beta2 -adrenoceptor stimulation increases protein synthesis and attenuates breakdown processes in skeletal muscle. Thus, the beta2 -adrenoceptor is a potential target in the treatment of disuse-, disease- and age-related muscle atrophy. In the present study, we show that a few days of oral treatment with the commonly prescribed beta2 -adrenoceptor agonist, salbutamol, increased skeletal muscle protein synthesis and breakdown during the first 5 h after resistance exercise in young men. Salbutamol also counteracted a negative net protein balance in skeletal muscle after resistance exercise. Changes in protein turnover rates induced by salbutamol were associated with protein kinase A-signalling, activation of Akt2 and modulation of mRNA levels of growth-regulating proteins in skeletal muscle. These findings indicate that protein turnover rates can be augmented by beta2 -adrenoceptor agonist treatment during recovery from resistance exercise in humans. ABSTRACT: The effect of beta2 -adrenoceptor stimulation on skeletal muscle protein turnover and intracellular signalling is insufficiently explored in humans, particularly in association with exercise. In a randomized, placebo-controlled, cross-over study investigating 12 trained men, the effects of beta2 -agonist (6 × 4 mg oral salbutamol) on protein turnover rates, intracellular signalling and mRNA response in skeletal muscle were investigated 0.5-5 h after quadriceps resistance exercise. Each trial was preceded by a 4-day lead-in treatment period. Leg protein turnover rates were assessed by infusion of [13 C6 ]-phenylalanine and sampling of arterial and venous blood, as well as vastus lateralis muscle biopsies 0.5 and 5 h after exercise. Furthermore, myofibrillar fractional synthesis rate, intracellular signalling and mRNA response were measured in muscle biopsies. The mean (95% confidence interval) myofibrillar fractional synthesis rate was higher for salbutamol than placebo [0.079 (95% CI, 0.064 to 0.093) vs. 0.066 (95% CI, 0.056 to 0.075%) × h-1 ] (P < 0.05). Mean net leg phenylalanine balance 0.5-5 h after exercise was higher for salbutamol than placebo [3.6 (95% CI, 1.0 to 6.2 nmol) × min-1  × 100 gLeg Lean Mass-1 ] (P < 0.01). Phosphorylation of Akt2, cAMP response element binding protein and PKA substrate 0.5 and 5 h after exercise, as well as phosphorylation of eEF2 5 h after exercise, was higher (P < 0.05) for salbutamol than placebo. Calpain-1, Forkhead box protein O1, myostatin and Smad3 mRNA content was higher (P < 0.01) for salbutamol than placebo 0.5 h after exercise, as well as Forkhead box protein O1 and myostatin mRNA content 5 h after exercise, whereas ActivinRIIB mRNA content was lower (P < 0.01) for salbutamol 5 h after exercise. These observations suggest that beta2 -agonist increases protein turnover rates in skeletal muscle after resistance exercise in humans, with concomitant cAMP/PKA and Akt2 signalling, as well as modulation of mRNA response of growth-regulating proteins.

Light-load resistance exercise increases muscle protein synthesis and hypertrophy signaling in elderly men.

The present study investigated whether well-tolerated light-load resistance exercise (LL-RE) affects skeletal muscle fractional synthetic rate (FSR) and anabolic intracellular signaling as a way to counteract age-related loss of muscle mass. Untrained healthy elderly (>65-yr-old) men were subjected to 13 h of supine rest. After 2.5 h of rest, unilateral LL-RE, consisting of leg extensions (10 sets, 36 repetitions) at 16% of 1 repetition maximum (RM), was conducted. Subsequently, the subjects were randomized to oral intake of 4 g of whey protein per hour (PULSE, n = 10), 28 g of whey protein at 0 h and 12 g of whey protein at 7 h postexercise (BOLUS, n = 10), or 4 g of maltodextrin per hour (placebo, n = 10). Quadriceps muscle biopsies were taken at 0, 3, 7, and 10 h postexercise from the resting and the exercised leg of each subject. Myofibrillar FSR and activity of select targets from the mechanistic target of rapamycin complex 1-signaling cascade were analyzed from the biopsies. LL-RE increased myofibrillar FSR compared with the resting leg throughout the 10-h postexercise period. Phosphorylated (T308) AKT expression increased in the exercised leg immediately after exercise. This increase persisted in the placebo group only. Levels of phosphorylated (T37/46) eukaryotic translation initiation factor 4E-binding protein 1 increased throughout the postexercise period in the exercised leg in the placebo and BOLUS groups and peaked at 7 h. In all three groups, phosphorylated (T56) eukaryotic elongation factor 2 decreased in response to LL-RE. We conclude that resistance exercise at only 16% of 1 RM increased myofibrillar FSR, irrespective of nutrient type and feeding pattern, which indicates an anabolic effect of LL-RE in elderly individuals. This finding was supported by increased signaling for translation initiation and translation elongation in response to LL-RE.

Contraction mode and whey protein intake affect the synthesis rate of intramuscular connective tissue.

INTRODUCTION: In this study we investigated the impact of whey protein hydrolysate and maltodextrin (WPH) intake on intramuscular connective tissue (IMCT) protein fractional synthesis rate (FSR) after maximal shortening and lengthening contractions. METHODS: Twenty young men were randomized to receive either WPH or maltodextrin [carbohydrate (CHO)] immediately after completion of unilateral shortening and lengthening knee extensions. Ring-13 C6 -phenylalanine was infused, and muscle biopsies were obtained. IMCT protein FSR was measured at 1-5, as well as 1-3 and 3-5 hours after contractions and nutrient intake. RESULTS: During the 1-3-hour recovery, lengthening contractions resulted in a higher FSR than shortening contractions (P < 0.01), independent of supplementation type and, during the 3-5-hour recovery, WPH had a higher FSR than CHO (P < 0.05), independent of prior contraction mode. CONCLUSIONS: The later appearance of a stimulating effect of WPH on the IMCT FSR after strenuous muscle contractions lends support to its ability to promote recovery of the muscle connective tissue matrix after exercise. Muscle Nerve 55: 128-130, 2017.

Deep sequencing of Danish Holstein dairy cattle for variant detection and insight into potential loss-of-function variants in protein coding genes.

BACKGROUND: Over the last few years, continuous development of high-throughput sequencing platforms and sequence analysis tools has facilitated reliable identification and characterization of genetic variants in many cattle breeds. Deep sequencing of entire genomes within a cattle breed that has not been thoroughly investigated would be imagined to discover functional variants that are underlying phenotypic differences. Here, we sequenced to a high coverage the Danish Holstein cattle breed to detect and characterize single nucleotide polymorphisms (SNPs), insertion/deletions (Indels), and loss-of-function (LoF) variants in protein-coding genes in order to provide a comprehensive resource for subsequent detection of causal variants for recessive traits. RESULTS: We sequenced four genetically unrelated Danish Holstein cows with a mean coverage of 27X using an Illumina Hiseq 2000. Multi-sample SNP calling identified 10,796,794 SNPs and 1,295,036 indels whereof 482,835 (4.5 %) SNPs and 231,359 (17.9 %) indels were novel. A comparison between sequencing-derived SNPs and genotyping from the BovineHD BeadChip revealed a concordance rate of 99.6-99.8 % for homozygous SNPs and 93.3-96.5 % for heterozygous SNPs. Annotation of the SNPs discovered 74,886 SNPs and 1937 indels affecting coding sequences with 2145 being LoF mutations. The frequency of LoF variants differed greatly across the genome, a hot spot with a strikingly high density was observed in a 6 Mb region on BTA18. LoF affected genes were enriched for functional categories related to olfactory reception and underrepresented for genes related to key cellular constituents and cellular and biological process regulation. Filtering using sequence derived genotype data for 288 Holstein animals from the 1000 bull genomes project removing variants containing homozygous individuals retained 345 of the LoF variants as putatively deleterious. A substantial number of the putative deleterious LoF variants had a minor allele frequency >0.05 in the 1000 bull genomes data set. CONCLUSIONS: Deep sequencing of Danish Holstein genomes enabled us to identify 12.1 million variants. An investigation into LoF variants discovered a set of variants predicted to disrupt protein-coding genes. This catalog of variants will be a resource for future studies to understand variation underlying important phenotypes, particularly recessively inherited lethal phenotypes.