Optimizing protein intake in aging

PURPOSE OF REVIEW: Our current knowledge on the causes of sarcopenia is still fragmentary. The most evident factors of age-related muscle alterations comprise impairment in protein turnover rate, neurodegenerative processes, reduction in anabolic hormone production and action, dysregulation of cytokine secretions, changes in the response to inflammatory events, sedentary lifestyle and inadequate nutritional intakes. Indeed, nutrition, in particular low protein intakes, may participate in the loss of protein mass during aging. Consequently, numerous studies have focused on the possibility to counteract the age effect on muscle loss by changing the quantity but also the quality of ingested proteins, aiming to optimize protein intake and retention through an improvement in amino acid bioavailability for the muscle tissues. RECENT FINDINGS: It has long been recognized that numerous dietary parameters, such as the amount of dietary proteins, affect protein metabolism. Recently, new concepts have been developed by testing either different types of protein sources, proteins with various digestion rates, meals with pulse or spread protein-feeding patterns or amino acids with specific anabolic function. This is therefore a new area in which not only quantitative but also qualitative properties of dietary protein are considered as a strategy to limit body protein loss during aging. SUMMARY: Recent studies have reported sarcopenia to be preventable by using nutritional intervention, but more experimental and clinical evidence is needed to modify the current recommendation in daily protein intake. The new concepts currently developed must be validated using large epidemiological studies in the long term before being applied to the elderly population.     

老年性肌少症（Sarcopenia）与营养干预

肌少症（Sarcopenia）是老龄化进程中以骨骼肌质量及其力量下降为特征的一类临床综合征,可降低老年人生活质量,增加老年人残疾发生率和疾病死亡率。营养素缺乏及其导致的肌蛋白合成减少、肌肉组织的特殊变化等都是肌少症发生和进展的重要原因。因此,深入探讨营养干预对老年性肌少症的防治具有重要的理论和临床意义。     

Effects of protein deficiency on the rate of radioactivity loss from body constituents in adult rats given 14C-amino acids.

The effect of protein deficiency on the rate of loss of radioactivity from body constituents was studied in adult rats administered 14C-Chlorella protein hydrolysate or 14C-lysine. Rats were kept on a protein-free diet for 3 weeks and then injected with labelled amino acids and fed on a protein-free diet for 3 more days to allow 14C deposition in tissues. Then they were given experimental diets (protein-free diet, 1% and 10% wheat gluten diets pair-fed with the protein-free diet, and 10% wheat gluten diet ad libitum) for 7 days and sacrificed. The rates of loss of radioactivity from tissue proteins became low in general with the extent of protein deficiency. This increased capacity of tissues to retain 14C-amino acids may result from higher efficiency of protein utilization in protein deficiency. The reutilization of free amino acids and the rate of catabolism of tissue proteins are discussed on the basis of the results. The half-life of muscle protein was too long to observe the effects of experimental diets given for 7 days on the rate of loss of radioactivity.     

Catabolic response to stress and potential benefits of nutrition support

The catabolic response to sepsis, severe injury, and burn is characterized by whole-body protein loss, mainly reflecting increased breakdown of muscle proteins, in particular myofibrillar proteins. Glucocorticoids and various proinflammatory cytokines are important regulators of muscle proteolysis in stressed patients. There is evidence that breakdown of proteins by the ubiquitin-proteasome pathway plays an important role in muscle cachexia, although other mechanisms may participate, such as calcium- and calpain-dependent release of myofilaments from the sarcomere. Three types of treatments have been used to reduce or prevent the catabolic response to injury and sepsis: 1). nutritional, 2). hormonal, and 3). pharmacologic. With regard to nutrition support, it is generally believed that enteral feeding is superior to parenteral feeding and that early feeding is better than late feeding. Although "immune-enhancing" enteral nutrition has been shown in several recent studies to improve outcome in critically ill patients, the specific effects of these treatments on the catabolic response in muscle are not known. In addition to nutrition support, various hormones, including insulin, growth hormone, and insulin-like growth factor-1, may blunt the catabolic response in patients with stress. Experimental studies have indicated that other treatments may become available in the future, including cytokine antibodies, calcium antagonists, and induction of heat shock response. Methods to prevent or reduce the catabolic response to stress are important considering the significant clinical consequences of muscle cachexia.     

An integrative approach to in-vivo protein synthesis measurement: from whole tissue to specific proteins

PURPOSE OF REVIEW: In-vivo estimation of protein turnover by stable isotopes in animals and humans has provided much relevant information on metabolic regulation and alterations for decades. While it was first appreciated at the whole body level in the 1970s and 1980s, new approaches have allowed inter-organ or tissue protein turnover rates to be measured, notably the incorporation rate of a labelled amino acid in muscle. These technical improvements have recently been completed by new isolation methods for the study of protein synthesis rates in various muscle and hepatic protein fractions in different blood cells or tissues such as bone and skin. RECENT FINDINGS: This new insight into tissue protein synthesis opens the door for exploration of single proteins, which may be fully achievable in the near future through the combination of proteomics analysis and technical progress in mass spectrometry. This is, therefore, a new area in which not only quantitative but also qualitative changes in specific proteins will be considered for a fully integrative approach to assessing protein metabolism in physiology and disease. SUMMARY: To understand the mechanisms by which protein metabolism is altered during physiopathological situations, it is of importance to measure the effect on specific proteins rather than on the body as a whole. Procedures are currently under development with the aim of isolating individuals proteins and to measure their synthesis rates by isotopic methods. Such technical progress is needed to gain a better understanding of the regulation of protein metabolism in situations in which loss of body protein mass occurs.     

Pulse protein feeding pattern restores stimulation of muscle protein synthesis during the feeding period in old rats

Muscle loss during aging could be related to a lower sensitivity of muscle protein synthesis to feeding. To overcome this decrease without increasing protein intake, we proposed to modulate the daily protein feeding pattern. We showed that consuming 80% of dietary proteins at noon (pulse pattern) improved nitrogen balance in elderly women. The present study was undertaken in rats to determine which tissues are the targets of the pulse pattern and what mechanisms are involved. Male Sprague-Dawley 11- and 23-mo-old rats (n = 32 per age) were fed 4 isoproteic (18% protein) meals/d for 10 d. Then half of the rats at each age were switched to a 11/66/11/11% repartition of daily proteins (pulse pattern) for 21 d. On d 21, rats were injected with a flooding dose of L-(13)C-valine (50 atom% excess, 150 micromol/100 g body) and protein synthesis rates were measured in liver, small intestine and gastrocnemius muscle in either the postabsorptive or the fed state. Epitrochlearis muscle degradation rates and plasma amino acid concentrations were measured at the same times. The pulse pattern had the following effects: 1) it significantly increased liver protein synthesis response to feeding and postprandial plasma amino acid concentrations at both ages; 2) it restored a significant response to feeding of gastrocnemius muscle protein synthesis in old rats; and 3) it had no effect in small intestine or on muscle breakdown. Thus, using a pulse pattern could be useful in preventing the age-related loss of muscle by increasing feeding-induced stimulation of muscle protein synthesis.     

Methods for measuring tissue protein breakdown rate in vivo

PURPOSE OF REVIEW: To describe the latest innovations in measuring protein breakdown in vivo, particularly in muscle. RECENT FINDINGS: The traditional method of using 3-methylhistidine excretion to measure muscle protein breakdown has been updated to include arteriovenous or microdialysis measurements, which address the concern that there are alternative sources of 3-methylhistidine in the body other than muscle. Several variations of a precursor-product method to measure fractional breakdown rate of tissues have been developed that are analogous to fractional synthesis rate of tissues. These methods are more generally applicable than the 3-methylhistidine methods and are less invasive than arteriovenous methods. The various precursor-product methods are distinguished by whether they require an isotopic steady state or multiple tracers and by how many biopsies are required. SUMMARY: The new precursor-product methods have enabled assessment in clinical trials of protein breakdown for proteins other than myofibrillar proteins and in circumstances in which arteriovenous sampling is not feasible.     

Perilipin family (PLIN) proteins in human skeletal muscle: the effect of sex, obesity, and endurance training

Proteins that coat the lipid droplets (also known as PAT proteins or perilipin (PLIN) family proteins) have diverse functions that are not well elucidated in many tissues. In skeletal muscle, there is even less known about the functions or characteristics of these proteins or how they might change in response to perturbations that alter both intramyocellular lipid (IMCL) content and fat utilization and oxidation. Therefore, the purpose of this study was to examine the human muscle content and gene expression of the four skeletal muscle PLIN proteins in both lean and obese men and women and how this was changed following a 12-week endurance training protocol. PLIN2-PLIN5 proteins were all more abundant in women than in men (p?= 0.037 and?p?< 0.0001, respectively), consistent with higher IMCL content observed in female skeletal muscle. PLIN5 (previously known as OXPAT) is of particular interest because it has previously been associated primarily with oxidative tissues that rely heavily on fat oxidation for energy production. Although PLIN5 was not different between lean and obese subjects, it was the only PLIN protein to increase in response to endurance training in both sexes. PLIN5 correlated with IMCL volume (p?< 0.0001), but in general, the other PLIN proteins did not correlate well with IMCL volume, suggesting that the relationship between lipid accumulation and PLIN family protein content is not a simple one. Although more work is necessary, it is clear that PLIN5 likely plays an important role in IMCL accumulation and oxidation, both of which increase with endurance training in human skeletal muscle.     

Vitamin E levels in sheep tissues at various times after a single oral administration of d-alpha-tocopherol acetate

Five groups, each of 5 sheep of 40-45 kg body weight were used. One group acted as control (killed at 0 time, no vitamin E dose) while the other four groups were given a single oral dose of d-alpha-tocopherol acetate (100 mg/kg body weight) and killed 24, 48, 72 and 240 h after dosing. Samples of adrenal, adipose, heart, liver, kidney, lung, muscle, pancreas and spleen were taken from all 25 sheep and were analyzed for their vitamin E content. In the control sheep the tocopherol concentration in some tissues such as pancreas and adrenal were substantially higher than in the other tissues. Muscle and adipose tissue contained the lowest tocopherol concentrations among the various tissues. Tissues responded differently to the vitamin E dosing. Liver was characterized by a rapid accumulation of alpha-tocopherol at 24 h followed by a progressive loss. In the adrenal and lung the peak occurred 48 h after vitamin E loading. The variable pattern in tocopherol concentrations in the various sheep tissues following vitamin E dosing was considered a reflection of their different metabolic activities.     

Mechanisms activating proteolysis to cause muscle atrophy in catabolic conditions.

The daily turnover of cellular proteins is the same as the amount of protein contained in 1 to 1.5 kg of muscle. Consequently, even a small but persistent increase in protein degradation or decrease in protein synthesis results in substantial loss of muscle mass, as shown in patients with trauma, sepsis, or kidney failure. Activation of the ubiquitin-proteasome proteolytic system in muscle is the major pathway contributing to loss of muscle mass in catabolic illnesses. At least 3 signals have been identified as causing loss of muscle mass: acidosis, defective insulin action, and glucocorticoids. The influence of inflammatory cytokines on this system in muscle is more complicated because cytokines can suppress the system unless glucocorticoids are present. An initial reaction that breaks down muscle appears to involve caspases. Such information could lead to therapies that successfully prevent the loss of muscle mass in catabolic illnesses.     

Managing sarcopenia with progressive resistance exercise training

Advancing age appears to alter the chemical and physical properties of skeletal muscle proteins. Alterations include: reduced contractile, mitochondrial, and enzyme protein synthesis rates, altered expression and post-translational modifications to muscle proteins, reduced maximum voluntary muscle strength, reduced muscle strength per unit muscle mass and muscle power. These age-associated impairments in muscle protein quantity and quality contribute to physical disability and frailty, a loss of independent function, the risk of falling and fractures, and contribute to escalating health care costs. Progressive resistance exercise training is a potent, non-pharmacologic, effective therapy that opposes the impairments in muscle protein quantity and quality in middle age and physically frail adults. In the absence of contraindications to exercise, muscle proteins adapt to an exercise training stimulus despite the depredation of age. The proposed pathogenesis for some of these impairments is briefly reviewed. Evidence that supports the use of progressive resistance exercise training to restore muscle quality and quantity in elderly adults is reviewed.     

Tracers to investigate protein and amino acid metabolism in human subjects

Three tracer methods have been used to measure protein synthesis, protein breakdown and protein oxidation at whole-body level. The method using L-[1-(13)C]leucine is considered the method of reference. These methods have contributed greatly to the existing knowledge on whole-body protein turnover and its regulation by feeding, fasting, hormones and disease. How exercise and ingestion of mixed protein-containing meals affect whole-body protein metabolism is still open to debate, as there are discrepancies in results obtained with different tracers. The contribution of whole-body methods to the future gain of knowledge is expected to be limited due to the fact that most physiological disturbances have been investigated extensively, and due to the lack of information on the relative contribution of various tissues and proteins to whole-body changes. Tracer amino acid-incorporation methods are most suited to investigate these latter aspects of protein metabolism. These methods have shown that some tissues (liver and gut) have much higher turnover rates and deposit much more protein than others (muscle). Massive differences also exist between the fractional synthesis rates of individual proteins. The incorporation methods have been properly validated, although minor disagreements remain on the identity of the true precursor pool (the enrichment of which should be used in the calculations). Arterio-venous organ balance studies have shown that little protein is deposited in skeletal muscle following a protein-containing meal, while much more protein is deposited in liver and gut. The amount deposited in the feeding period in each of these tissues is released again during overnight fasting. The addition of tracers to organ balance studies allows the simultaneous estimation of protein synthesis and protein breakdown, and provides information on whether changes in net protein balance are caused primarily by a change in protein synthesis or in protein breakdown. In the case of a small arterio-venous difference in a tissue with a high blood flow, estimates of protein synthesis and breakdown become very uncertain, limiting the value of using the tracer. An additional measurement of the intracellular free amino acid pool enrichment allows a correction for amino acid recycling and quantification of the inward and outward transmembrane transport. However, in order to obtain reliable estimates of the intramuscular amino acid enrichment and, therefore, of muscle protein synthesis and breakdown in this so-called three-pool model, the muscle should be freeze-dried and the resulting fibres should be freed from connective tissue and small blood clots under a dissection microscope. Even when optimal precautions are taken, the calculations in these tracer balance methods use multiple variables and, therefore, are bound to lead to more variability in estimates of protein synthesis than the tracer amino acid incorporation methods. In the future, most studies should focus on the measurement of protein synthesis and breakdown in specific proteins in order to understand the mechanisms behind tissue adaptation in response to various stimuli (feeding, fasting, exercise, trauma, sepsis, disuse and disease). The tracer laboratories, therefore, should improve the methodology to allow the measurement of low tracer amino acid enrichments in small amounts of protein.     

The utility of resistance exercise training and amino acid supplementation for reversing age-associated decrements in muscle protein mass and function

Advancing age is associated with reduced skeletal muscle protein synthesis, altered expression of and chemical modifications to muscle proteins, reduced muscle strength, muscle strength per unit muscle mass and muscle power (sarcopenia). These age-associated impairments in the quantity and quality of contractile protein contribute to physical disability and frailty, a loss of independent function, the risk of falling and fractures, and escalating health-care costs. Progressive resistance exercise training is a potent, non-pharmacologic, efficacious therapy for the impairment in muscle quantity and quality in middle age and physically frail adults. Evidence is accumulating that dietary amino acid supplementation may also improve muscle protein balance in the elderly. Several potential cellular mechanisms for the loss of muscle protein and resistance exercise-induced improvements in muscle quantity and quality in elderly adults are reviewed.     

Tissue antioxidant status differs in spontaneously hypertensive rats fed fish protein or casein

The present study was designed to determine whether changes in dietary protein source are related to changes in antioxidant status determined by enzyme activities of catalase, superoxide dismutase (SOD), gluthatione peroxidase (GSH-Px) and gluthatione reductase (GSSG-Red) and lipid peroxidation levels in various tissues. Spontaneously hypertensive rats (SHR; 5 wk old) were fed diets containing 20% casein or fish protein for 2 mo. Feeding the fish protein diet lowered blood pressure and reduced plasma total cholesterol levels and SOD activity in all tissues except muscle compared with the casein diet. Feeding fish protein also enhanced GSH level and GSH-Px activity in liver and heart, accompanied by lower lipid peroxidation. In kidney, however, the lower catalase activity in rats fed fish protein was associated with an enhancement in lipid peroxidation. Plasma and VLDL + LDL lipid peroxidation was unaffected by dietary proteins. In conclusion, the fish protein diet did not play a relevant role in plasma antioxidative defense status but increased it in liver and heart compared with the casein diet. Fish protein attenuated the development of hypertension and also decreased plasma total cholesterol concentration. Thus, it enhances protection against cardiovascular diseases.     

Polyphenols and their potential role in preventing skeletal muscle atrophy

Skeletal muscle atrophy is the consequence of various conditions, such as disuse, denervation, fasting, aging, and disease. Even if the underlying molecular mechanisms are still not fully understood, an elevated oxidative stress related to mitochondrial dysfunction has been proposed as one of the major contributors to skeletal muscle atrophy. Researchers have described various forms of nutritional supplementation to prevent oxidative stress-induced muscle wasting. Among a variety of nutrients, attention has also focused on polyphenols, a wide range of plant-based compounds with antioxidant and inflammatory properties, many of which have beneficial effects on human health and might retard skeletal muscle loss and function impairment. The purpose of this review is to describe polyphenol actions in skeletal muscle atrophy prevention. Published articles from the last 10 years were searched on PubMed and other databases. Polyphenols are important molecules that should be considered when discussing possible strategies against muscle atrophy. In particular, the collected studies describe, for each polyphenol subclass, the beneficial effect on muscle mass preservation in various skeletal muscle disorders. In these examples, the polyphenol compounds appear to mainly act by reversing mitochondrial dysfunction. Given that the current information on polyphenols is mostly restricted to basic studies, more comprehensive research and additional studies should be performed to clarify their mechanisms of action in improving skeletal muscle functions during atrophy.     

Insulin stimulated glucose disposal in peripheral tissues studied with microdialysis and stable isotope tracers

BACKGROUND & AIMS: Methods to study glucose kinetics in vivo in specific tissues or tissue beds in humans are often not feasible due to invasiveness or costs of equipment needed. Here we investigate whether the loss (fractional extraction) of 2H7-glucose infused via a microdialysis catheter can be used to study glucose disposal in skeletal muscle and subcutaneous adipose tissue. METHODS AND RESULTS: A perfusion period of 2 h was needed to ensure an isotopic steady state in the microdialysis catheters in skeletal muscle and adipose tissue. In six healthy volunteers the fractional extraction increased during a hyperinsulinemic euglycemic clamp in both skeletal muscle and adipose tissue. Following 48 h of starvation in the same subjects, insulin was not able to increase the fractional extraction of 2H7-glucose from the microdialysis in comparison with a baseline measurement. CONCLUSIONS: In response to insulin infusion, the fractional extraction of 2H7-glucose from a microdialysis catheter increases in skeletal muscle and subcutaneous adipose tissue and this increase is blunted during insulin resistance induced by starvation. These results validate that the fractional extraction of a glucose tracers infused via microdialysis can be used as an index of glucose disposal in peripheral tissues or tissue beds.     

Endocannabinoid signaling and energy metabolism: a target for dietary intervention

The endocannabinoid (EC) signaling (ECS) system involves the activation of receptors targeted by endogenously produced ligands called endocannabinoids that trigger specific physiologic events in various organs and tissues throughout the body. ECs are lipid mediators that bind to specific receptors and elicit cell signaling. The focus of this review is to discuss the responses that direct pathways of systemic energy metabolism. Recent findings have indicated that an imbalance of the ECS contributes to visceral fat accumulation and disrupts energy homeostasis, which are characteristics of the metabolic syndrome. Constant activation of ECS has been linked to metabolic processes that are associated with the hypothalamus and peripheral tissues of obese patients. In contrast, inhibition of ECS results in weight loss in animal and human subjects. Despite these findings, the mechanism involved in the dysregulation of ECS is unclear. Interestingly, the level of endogenous ligands, derived from arachidonic acid, can be directly manipulated by nutrient intervention, in that a diet rich in long-chain ω-3 polyunsaturated fatty acids will decrease the production of ligands to modulate the activation of target receptors. In contrast, a diet that is high in ω-6 polyunsaturated fatty acids will cause an increase in ECS activation and stimulate tissue specific activities that decrease insulin sensitivity in muscle and promote fat accumulation in the adipose tissue. The purpose of this review is to explain the components of ECS, its role in adipose and muscle energy metabolism, and how nutritional approaches with dietary ω-3 polyunsaturated fatty acids may reverse the dysregulation of this system to improve insulin sensitivity and control body fat.     

β-Sitosterol Attenuates Dexamethasone-Induced Muscle Atrophy via Regulating FoxO1-Dependent Signaling in C2C12 Cell and Mice Model

Sarcopenia refers to a decline in muscle mass and strength with age, causing significant impairment in the ability to carry out normal daily functions and increased risk of falls and fractures, eventually leading to loss of independence. Maintaining protein homeostasis is an important factor in preventing muscle loss, and the decrease in muscle mass is caused by an imbalance between anabolism and catabolism of muscle proteins. Although β-sitosterol has various effects such as anti-inflammatory, protective effect against nonalcoholic fatty liver disease (NAFLD), antioxidant, and antidiabetic activity, the mechanism of β-sitosterol effect on the catabolic pathway was not well known. β-sitosterol was assessed in vitro and in vivo using a dexamethasone-induced muscle atrophy mice model and C2C12 myoblasts. β-sitosterol protected mice from dexamethasone-induced muscle mass loss. The thickness of gastrocnemius muscle myofibers was increased in dexamethasone with the β-sitosterol treatment group (DS). Grip strength and creatine kinase (CK) activity were also recovered when β-sitosterol was treated. The muscle loss inhibitory efficacy of β-sitosterol in dexamethasone-induced muscle atrophy in C2C12 myotube was also verified in C2C12 myoblast. β-sitosterol also recovered the width of myotubes. The protein expression of muscle atrophy F-box (MAFbx) was increased in dexamethasone-treated animal models and C2C12 myoblast, but it was reduced when β-sitosterol was treated. MuRF1 also showed similar results to MAFbx in the mRNA level of C2C12 myotubes. In addition, in the gastrocnemius and tibialis anterior muscles of mouse models, Forkhead Box O1 (FoxO1) protein was increased in the dexamethasone-treated group (Dexa) compared with the control group and reduced in the DS group. Therefore, β-sitosterol would be a potential treatment agent for aging sarcopenia.     

Ornithine alpha-ketoglutarate: Could it be a new therapeutic option for sarcopenia?

Our current knowledge on the causes of sarcopenia is still fragmentary. One of the most evident candidates to explain muscle loss in elderly includes imbalance in protein turnover, i.e. decreased muscle protein synthesis rate, notably in the post-prandial state. Nutritional strategies such as leucine supplementation, use of fast digested proteins or a pulse protein intake have been show to enhance the synthesis rate of muscle proteins in older individuals. Ornithine alpha-ketoglutarate (OKG) is a precursor of amino acids such as glutamine, arginine and proline, and increases the secretion of anabolic hormones, i.e. insulin and growth hormone. A beneficial anabolic action of OKG has been demonstrate in several pathological conditions associated with muscle loss. Therefore, OKG may be of a potential interest to modulate muscle protein metabolism and to maintain muscle mass during aging.