Increasing Muscle Mass in Elders through Diet and Exercise: A Literature Review of Recent RCTs

This study aimed to review the current evidence on the independent and combined effects of diet and exercise and their impact on skeletal muscle mass in the elderly population. Skeletal muscle makes up approximately 40% of total body weight and is essential for performing daily activities. The combination of exercise and diet is known to be a potent anabolic stimulus through stimulation of muscle protein synthesis from amino acids. Aging is strongly associated with a generalized deterioration of physiological function, including a progressive reduction in skeletal muscle mass and strength, which in turn leads to a gradual functional impairment and an increased rate of disability resulting in falls, frailty, or even death. The term sarcopenia, which is an age-related syndrome, is primarily used to describe the gradual and generalized loss of skeletal muscle mass (mainly in type II muscle fibers) and function. Multimodal training is emerging as a popular training method that combines a wide range of physical dimensions. On the other hand, nutrition and especially protein intake provide amino acids, which are essential for muscle protein synthesis. According to ESPEN, protein intake in older people should be at least 1 g/kgbw/day. Essential amino acids, such as leucine, arginine, cysteine, and glutamine, are of particular importance for the regulation of muscle protein synthesis. For instance, a leucine intake of 3 g administered alongside each main meal has been suggested to prevent muscle loss in the elderly. In addition, studies have shown that vitamin D and other micronutrients can have a protective role and may modulate muscle growth; nevertheless, further research is needed to validate these claims. Resistance-based exercise combined with a higher intake of dietary protein, amino acids, and/or vitamin D are currently recognized as the most effective interventions to promote skeletal muscle growth. However, the results are quite controversial and contradictory, which could be explained by the high heterogeneity among studies. It is therefore necessary to further assess the impact of each individual exercise and nutritional approach, particularly protein and amino acids, on human muscle turnover so that more efficient strategies can be implemented for the augmentation of muscle mass in the elderly.


Introduction
Aging is associated with a generalized deterioration of physiological function, accompanied by a progressive decline in skeletal muscle mass, and strength, gradually leading to functional impairment, increased disability, and dependency [1]. The risks of falls, loss of independence, and premature death have been progressively rising in the last few decades among the elderly population [2,3]. Moreover, these burdens are linked to a higher economic burden on the healthcare system [4]. In addition to the loss of muscle mass due to aging, low muscle mass is observed as a result of diseases such as cancer, chronic obstructive pulmonary disease, heart failure, and renal failure and is a prognostic indicator in a multitude of diseases [5][6][7][8][9][10]. The prevalence of sarcopenia varies according to the population or according to measurement tools [11]. In a meta-analysis by Papadopoulou et al. [11], the prevalence of sarcopenia ranged between 9-11% in women and men, respectively, in community-dwelling older adults. The prevalence of sarcopenia increases dramatically in nursing-home residents and hospitalized individuals, approaching 51% in men and 31% in women, and about 23% in both men and women, respectively [11].
Maintaining muscle mass in populations due to pathophysiological and/or pathoanatomical conditions is difficult without appropriate intervention. The combination of exercise and dietary supplementation with a specific amino acid mixture appears to have similar histopathological, biochemical, and functional changes in elderly patients [12]. A study has gathered evidence that the intake of essential amino acids can improve the exercise capacity of people with chronic heart failure or chronic obstructive pulmonary disease [12]. This improvement is due to the fact that they increase muscle mass and function, improve their aerobic metabolism, and increase their sensitivity to insulin [12]. Their ability to do the above appears to derive from the mechanisms they activate in muscle tissue, namely mitochondrial generation and myofibril growth, as well as glucose and cardiac muscle control [12]. In contrast with healthy adult exercisers and sportsmen, who maintain or even increase muscle mass through exercise and diet following current recommendations, elder people with morbidities and co-morbidities require particular attention, as a pathophysiological decline in muscle mass leads to other health problems [13], perpetuating a vicious cycle.
Muscles play an essential role in performing daily activities. In humans, skeletal muscles comprise approximately 40% of total body weight and contain 50-75% of all body proteins [14]. Maintaining or increasing muscle mass is a key component for improving daily activities as well as sports performance in activities of daily living and sports [15]. On the other hand, accelerated loss of muscle mass and function is associated with increased adverse outcomes, including falls, functional decline, frailty, and mortality [16].
Imaging methods such as X-ray or computed tomography (CT) are the gold standard for measuring muscle mass [17]. However, these methods are not used in daily clinical practice due to several limitations, such as their high cost. Most commonly, dual-energy X-ray absorptiometry (DXA) is used to estimate appendicular skeletal muscle mass/appendicular soft tissue (ASMM) or lean body mass (fat-free mass, total lean soft tissue plus bone mass, or body weight minus fat mass) as a proxy for muscle mass [18]. A simpler and even non-invasive technique of measuring muscle mass is bioelectrical impedance analysis (BIA), which is used for the estimation of fat-free mass (FFM), ASMM, or even total body skeletal muscle mass (SMM) through predictive equations [19]. According to the literature, the range of published cut-off values for ASMM by DXA is 5.86-7.40 kg/m 2 in men and 4.42-5.67 kg/m 2 in women, whereas cut-offs through the BIA analysis method range from 6.75-7.40 kg/m 2 in men and 5.07-5.80 kg/m 2 in women [19]. In healthy populations, cut-off values based on CT or magnetic resonance imaging (MRI) methods ranged between 36.54 and 45.40 cm 2 /m 2 in men and 30.21 and 36.05 cm 2 /m 2 in women [18].
The skeletal muscle groups of the human body consist of muscle fiber bundles. At the level of the whole muscle, its size is mainly determined by the number and size of individual muscle fibers [14]. The two most abundant myofilaments (proteins) are actin and myosin, which together comprise approximately 70-80% of the total protein content of a single fiber [14]. Skeletal muscle fibers are grouped into two categories: type I, called slow twitch, and type II, called fast twitch [20]. Type II muscle fibers can be grouped in terms of how myosin chains are expressed into three classes: IIa, IIx, and IIb, which are not present in our species [14]. A single muscle fiber may simultaneously express more than one type of myosin heavy chain; for example, types I and IIa or IIa and IIx together [14].
Altered expression in the myosin heavy chain may be the result of mRNA transcriptional activity in different nuclear regions [20]. These hybrid fibers appear to have two main roles: i. they function as mediators during fiber-type transitions associated with skeletal muscle development, exercise adaptation, and aging; and ii. they provide a functional continuum of fiber phenotypes, as they possess physiological properties that are intermediate to those of pure fiber types [21].
Muscle mass size is determined by coordinated changes in muscle protein synthesis and breakdown [22]. The combination of exercise and diet is a potent anabolic stimulus through the stimulation of muscle protein synthesis by amino acids [23]. Increased muscle mass is due to the accumulation of small amounts of protein in response to each bout of exercise combined with nutrient intake [23,24]. The effect of dietary supplements on muscle metabolism and muscle loss and regain has also been investigated and will be discussed below.
Metabolism, i.e., the set of reactions of the organism, consists of metabolic pathways in which molecules of chemical compounds are converted into others through processes catalyzed by enzymes and facilitated by other chemical compounds, such as cofactors, coenzymes, and metabolic intermediates [25]. Nutrients are molecules that participate in and influence the reactions of metabolism in general, therefore also muscle metabolism. Nutrients can be obtained through the diet, but in cases of deficiency, especially in the elderly, they can also be taken in the form of supplements.
Nutrition plays a key role in the prevention and treatment of sarcopenia. Energy intake decreases by almost 25% between 40 and 70 years of age, leading to inadequate intake of macro-and micronutrients [26]. Nutritional intake is often compromised in older people, and the risk of malnutrition is increased. According to ESPEN (the European Society for Clinical Nutrition and Metabolism), the presence of either a high unintentional loss of body mass (>5% at six months or >10% beyond six months) or a markedly reduced body mass index (BMI) (<20 kg/m 2 ) or muscle mass should be considered a serious indication of malnutrition that requires clarification of the underlying causes [27].
The total energy expenditure in older individuals is 24-36 kcal/kg body weight (BW) [27]. The basic energy requirements are influenced by lean body mass, gender, and nutritional status. Total energy expenditure is higher for men than for women and increases with decreasing BMI [27]. Maintaining or increasing body weight and muscle mass are central goals of nutritional support. Adequate protein intake of high biological value provides all essential amino acids, whereas both vitamin D and B12 have been studied for the prevention and treatment of sarcopenia [26].
There is a growing body of literature demonstrating how nutritional supplements affect muscle metabolism and muscle mass. Vaughan et al. [28] have shown, in in vitro experiments in human rhabdomyosarcoma cells, that mitochondrial biosynthesis increases within one day when OxyElite Pro and Cellucore HD dietary supplements are administered. These contain, among other ingredients, germanium and caffeine, respectively. The oxidative and glycolytic capacities of the cells also increased [28]. The effect of creatine on muscle metabolism is the subject of various research studies. It has been suggested that it acts as a positive stimulus for protein synthesis pathways, either as a cell stressor or by targeting the mTOR pathway [29]. It even affects the synthesis of muscle fibers, altering the secretion of myokines or regulatory factors that increase the mitotic divisions of cells that will differentiate into myofibrils [29]. L-carnitine supplementation seems to increase muscle mass, but due to increased levels of a potentially atherogenic compound (fasting plasma trimethylamine-N-oxide, TMAO), further research is required before long-term supplementation is recommended [30]. At the amino acid level, in vitro and in vivo animal studies have shown that the amino acids serine and glycine are required for muscle regeneration [31]. In addition, serine levels have been found to decrease in older individuals, so it is hypothesized that serine supplementation may benefit muscle regeneration [31].
Regarding vitamin supplements, meta-analyses show that supplementation with vitamin D has a catalytic effect on muscle strength but not on muscle mass in the elderly and especially in those with initially low serum levels of this vitamin [32]. In contrast, more recent studies do not show that muscle in the elderly population is affected by the intake of vitamin D supplements [33]. More and better-designed studies are needed to accurately determine the effects on the human population. Additionally, the various forms of vitamin K appear to affect energy metabolism in skeletal muscle by increasing ATP production and maintaining the mitochondrial number and satellite cell proliferation in muscle [34].
Exercise is the main method for increasing muscle mass. It is also well-documented that training can promote muscle protein synthesis and activate signaling pathways that regulate muscle fiber metabolism and function [35]. Exercise can be a method to treat muscle atrophy in many of these conditions. Mitochondria play an important role in skeletal muscle homeostasis and bioenergy metabolism. Mitochondria are sensitive to contractile signals, and therefore, exercise can improve mitochondrial function and promote biosynthesis, which ultimately maintains the healthy state of cells and the whole body [36]. Exercise can activate the signaling pathway that stimulates skeletal muscle fiber metabolism and promotes contraction and physiological muscle function [36]. In particular, exercise, regardless of diet, leads to an increase in both muscle mass synthesis and muscle mass breakdown. This continuous turnover of muscle proteins results in the efficient repair and renewal of damaged (either mechanically, through oxidation, or otherwise) proteins [37]. On the other hand, exercise training can improve muscle metabolism and ameliorate abnormalities of muscle function without altering the functional performance of the heart [38]. It has been reported that exercise training can increase mitochondrial volume by up to 40% [37]. During physical exercise, the factors that regulate mitochondrial biogenesis are increased, directly enhancing mitochondrial protein synthesis. In aging skeletal muscle, mitochondria are smaller, with slower metabolism and reduced biosynthesis, resulting in a rapid decline in muscle mass and muscle performance parameters [36]. Moderate exercise training can protect mitochondria from volume and biogenesis-induced reductions in volume and biogenesis from aging and therefore attenuate the age-associated decline in skeletal muscle mass [36].
Progressive muscle-strengthening activities maintain or increase muscle mass and strength [39]. This training method is a well-established treatment for muscle atrophy and has been shown to: (a) shorten hospital stays; (b) enhance muscle cross-sectional area; and (c) improve grip strength in older adults [40]. Progressive resistance training is effective for both strength and muscle mass enhancement in older individuals and has been widely used in this population [41][42][43].
According to current evidence, aerobic training has little effect on skeletal muscle size compared to resistance exercise. However, aerobic activity can help slow down muscle loss with aging [39]. Moderate-intensity aerobic exercise, specifically walking, seems to improve the quality of skeletal muscle in healthy elderly adults who already have a sedentary lifestyle by improving the strength of the quadriceps muscles, without, however, increasing lean muscle mass [44].
For the elderly, multiform therapeutic exercise, which is a combination of resistance exercises, aerobic exercises, and balance and/or walking exercises, has also been proposed to improve muscle mass, strength, and functionality [45]. In addition, engaging in strength exercises at least twice a week has been associated with a reduced risk of sarcopenia, greater muscle mass, and better physical performance in elderly adults who already accumulate 150 min of moderate-intensity aerobic exercise per week [46,47]. Combined resistance training and aerobic programs have shown significant beneficial effects on anthropometric characteristics, including muscle mass and functional parameters [48][49][50].
In contrast, physical inactivity and sedentary behavior contribute to low levels of energy expenditure and result in adverse effects, including loss of aerobic capacity and musculoskeletal and cognitive decline [51].
The aim of this review is to summarize the recent research data on the independent and combined effects of diet and exercise and their effect/impact on skeletal muscle mass in the elderly population/in older adults.

Methods
The PubMed database was searched using the following search strategy: ( Inclusion criteria were: (1) older adults > 65 years old; (2) randomized controlled trials; (3) per os supplementation of nutrients; and (4) English language. Furthermore, muscle mass should be measured by BIA, DXA, MRI, or CT. Participants with diseases such as cancer, kidney disease, mobility issues, osteoarthritis, HIV, transplants, liver diseases, Cushing syndrome, arthritis, and neurological diseases (Parkinson) were excluded. Critically ill patients or patients with comorbidities are also excluded. Trials with home-based exercises are not eligible. Finally, we excluded studies in animal models, in vitro, and in vivo.

Results
A total of 3510 studies were identified through a PubMed search. Among these studies, 3407 were excluded by the title and/or abstract. In total, 103 studies were screened in full text, of which 41 were included in the review.
No changes in qCSA between LITW and WHEY (mean between-group difference,

Discussion
The loss of muscle mass and strength results in a decrease in functionality, which inevitably leads to the inability to perform daily activities, disability, loss of independence, and poor quality of life, leading to the creation of frailty syndrome while at the same time placing a significant burden on the health system. Recent descriptive, synchronic, and prospective studies confirm the importance and magnitude of the problem since the number of falls, fractures, and inability to self-care occur with high frequency in elderly people over 70 years old with sarcopenia [85].
Both older people and elders are at risk of the effects of muscle mass loss with age. Nutritional support for preserving or even enhancing muscle mass is an area of particular interest. Protein intake provides amino acids, which are required for muscle synthesis, as already mentioned. There is also a general concern that older people experience blunted muscle production, which raises the possibility that the overall recommendation for protein intake should be higher for older people [86]. A higher amount of protein (25-30 g) is required to overcome the threshold of maximal activation of muscle protein synthesis [87]. According to ESPEN, protein intake in older people should be at least 1 g/kg/bw/day [27]. The amount should be determined individually and should depend on the nutritional status, the level of physical activity, and the status of any possible existing diseases [27]. The distribution of protein between meals also showed a high frequency of adequate consumption [88]. Data also show that increased protein intake at morning meals led to increased protein intake throughout the day [89] and to the maintenance of skeletal muscle mass [90] in those who consumed increased protein in their evening meal. Timing and frequency of protein consumption are also considered important for muscle synthesis, while it needs to be evenly distributed among the main meals. Based on current evidence, it is observed that to maximize anabolic protein consumption, one should target an intake of 0.4 g/kg/meal, over at least four meals, to reach at least 1.6 g/kg/day [91]. Using the upper daily intake of 2.2 g/kg/day, distributed over the same four meals, a maximum of 0.55 g/kg/meal would be needed [91]. In addition, it has been suggested that muscle protein synthesis is maximized in young adults with an intake of~20-25 g of high-quality protein.
Muscle protein synthesis is attenuated in the older and in the elderly when protein and carbohydrates are co-ingested or when the intake of protein is less than about 20 g per meal. The pivotal role of protein supplementation in muscle mass maintenance and development has been widely investigated. Supplementing regular nutrient-mixed meals with leucine may also improve the muscle protein synthesis rate in the elderly [92] Supplementation of soy protein, whey protein, and their combination in older adults had a neutral effect on the maintenance of the appendicular skeletal mass index (ASMI) for a six-month supplementation, compared to baseline status [61]. Furthermore, fortification with whey or collagen peptides in cases of older people with low energy activity and under energy balance, deficit, and recovery conditions revealed that only the whey supplement protein improved leg lean mass and muscle protein synthesis in the recovery period [62]. An uptake in SMM was observed after 3 months of a supplemented diet with a carbohydrate mixture (fat and low protein) in community-dwelling individuals over 70 years of age [67]. The impact of whey protein supplement intake or dietary intake on muscle strength and mass improvement demonstrated no differences between the above treatments. However, supplement intake was found to improve walking speed in older people under 75 years of age. Whey protein provides a mixture of essential amino acids, with leucine being found in high concentrations. In addition to leucine, other amino acids, such as arginine, cysteine, and glutamine, may be involved in the anabolic effects of whey protein supplementation [93]. Six months of dietary supplementation with beta-hydroxyl, beta-methyl butyric acid (HMB), arginine, and glutamine resulted in significant improvements in total lean body mass among healthy older adults [59]. Casein is also digested slowly and progressively during sleep [94]. Ingestion of 30-40 g of casein (milk protein) 30 min before bedtime or via nasogastric tube increased muscle protein synthesis overnight in both young and elderly men [95]. Moreover, dairy products are a good source of high-quality protein and contain various essential nutrients [96]. Dairy proteins, in an amount of 14-40 g/day, can significantly increase the muscle mass of the limbs in middle-aged and elderly adults without, however, having a significant clinical effect on handgrip and leg press muscle strength [97].
Protein supplementation may enhance the effect of exercise on skeletal muscle signaling, particularly by increasing anabolic pathways and decreasing catabolic pathways. The additional effects of protein supplementation on this signaling can be explained by the increased availability of certain amino acids, such as leucine, and dipeptides, including glutamine dipeptides [93]. Hydrolyzed whey protein is a rich source of these ingredients. The way in which leucine induces these anabolic effects within the muscle cell appears to be through stimulation of the mammalian target of rapamycin complex 1 (mTORc1). This is achieved by inducing two main effects on the mTORc1 pathway. The first takes place through the activation of ribosome S6 kinase p70, and the second occurs through the inactivation of eukaryotic initiation factor 4E binding protein 1 (eIF4B), a protein known to repress protein translation. Other key downstream factors of mTORC1 signaling are ribosomal protein S6 (S6) and eukaryotic elongation factor 2 (eEF2), where activation of these proteins will eventually lead to protein synthesis. Intervention with protein supplementation in individuals working in parallel with resistance training resulted in increased expression of S6 and eEF2, indicating a signal favoring protein synthesis, which may be stimulated by increased leucine supplied to skeletal muscle [91]. The whey protein dose effect has been studied in the elderly and showed greater stimulation of muscle protein synthesis with the consumption of 35 g of whey protein, compared to 20 and 10 g of whey protein [98]. Furthermore, lower values of skeletal muscle index, handgrip strength, and performance were associated with lower blood leucine levels [99]. According to recommendations, 3 g of leucine is needed at each main meal to prevent muscle mass loss in older adults [99]. The richest sources of leucine are lean meat, whey products, dairy products, peanuts, lentils, and black beans [99]. Nevertheless, taking supplements makes it easier for this age group to achieve the nutritional goals for their age [64].
On the other hand, branched amino acids and especially leucine supplements' effects on muscle mass have been studied further. Murphy et al. 2021 [63] observed no effect of leucine supplementation or a combination of leucine with omega-3 fatty acids on muscle protein synthesis, lean mass, and strength in elderly subjects at risk of sarcopenia.
The effect of a combined supplement containing essential amino acids, vitamin D, and a mixture of alanine, resveratrol, CoQ10, and creatine in healthy elderly subjects was evaluated in a 12-week study [65]. The results showed a significant positive effect on muscle mass and strength, covering the age-related loss of years [65], compared to the control group receiving an isothermal maltodextrin supplement. However, this study cannot determine which ingredient or ingredients of the supplement affected the increase in muscle mass, as they were not studied separately.
Another macronutrient compound affecting muscle mass is medium-chain triglycerides (MCTs). MCTs are triglycerides consisting of a glycerol skeleton and three glycerol chains with an aliphatic tail of 6-12 carbon atoms. Abe et al. [68] examined whether intake of medium-chain triglycerides (MCTs) alone is sufficient to increase muscle mass, strength, and function in nursing home elders. MCTs alone or in combination with leucine tended to have a positive effect on the arm muscle area (AMA), increasing compared to the control group. In addition, those effects are reversible within a short period after discontinuation of the intervention. Vitamin D has been linked to the pathogenesis of sarcopenia and other diseases [100]. A study on the combination of vitamin D and protein is adding to the questions about the energy reserves required for the action of vitamin D and protein. In particular, a randomized, controlled, double-blind trial showed a greater increase in muscle mass with supplementation of vitamin D-enriched whey protein and leucine in participants with sarcopenia who had higher initial serum concentrations of 25(OH)D as well as a higher initial dietary protein intake. This suggests that blood vitamin D concentrations greater than 50 nmol/L and a sufficiently high dietary protein intake of more than 1 g per kg bodyweight per day may be required to induce a significant long-term increase in muscle mass from supplemental intake of these macronutrients [69]. The PROVIDE study evaluated the effect of baseline serum 25-hydroxyvitamin D [25(OH)D] and dietary protein concentrations on muscle mass and strength in elderly subjects receiving a dietary intervention. Results showed that basal concentrations of vitamin D and protein were essential for the 3-month dietary intervention to increase muscle mass and muscle strength in the study's participants [69]. Appropriate baseline levels of 25(OH)D and protein intake may be required to improve muscle mass as a result of vitamin D and protein supplementation intervention in sarcopenic older adults. This implies that current levels of vitamin D and protein intake recommendations could be considered the "minimum" for sarcopenic adults to adequately respond to dietary strategies aimed at mitigating muscle loss. Similarly, concomitant administration of a mixture of leucine, cholecalciferol, and medium-chain triglycerides had a positive effect on increasing appendicular muscle mass (AMM), SMI, and SMM in subjects with high levels of vitamin D at baseline [69]. Additionally, in a six-week intervention study combining vitamin D supplementation with whey protein and leucine at breakfast, muscle mass and protein synthesis in healthy elderly subjects were assessed. The results revealed that this dietary intervention stimulated protein synthesis and increased muscle mass in the elderly in the intervention group, statistically significantly compared to the control group consuming a placebo [66]. Many researchers have studied the effect of combining exercise and diet on maintaining or increasing muscle mass in the elderly [72,73,76,77]. The combination of a nutritional supplement and resistance training had no further improvement in muscle mass. The content of each drink had a total energy of 150 kcal and contained 20.7 g protein (whey protein, 3 g leucine, >10 g essential amino acids), 9.3 g carbohydrates, 3 g fat, vitamins (e.g., 800 IU vitamin D, 2.9 mg vitamin B6, 3 µg vitamin B12), and minerals [101]. Similar results were found after 6 weeks of resistance training with concomitant administration of β-hydroxyl-β-methyl-butyrate acid (HMB), a metabolite of leucine. HMB supplementation showed a marginal increase in lean body mass in the thighs compared to placebo, indicating an early increase in protein synthesis compared to the first two weeks of resistance training [71]. A synergistic effect is observed between protein intake, vitamin D, and exercise [83]. A 12-week randomized controlled trial during which elderly subjects with sarcopenia or dynapenia (low body strength) were exercising with bodyweight resistance training while ingesting a supplement with 10 g of whey protein and 20 µg of vitamin D increased muscle mass at the four extremities in those who had sarcopenia but not in those with normal mass [83]. In contrast, the phase angle, an indicator showing the quality of muscle mass related to sarcopenia [102], was increased in the elderly with low physical function and normal muscle mass but not in those with sarcopenia [83]. A significant beneficial effect of supplemental whey protein and vitamin D intake compared to placebo was found in elderly subjects with sarcopenia who participated in controlled resistance training with a total increase of 1.7 kg in body fat-free mass [99]. The beneficial effects observed only in the supplemented group suggest that physical activity is important but not sufficient to achieve a significant effect. However, it should be noted that physical activity was mild and non-intensive, which may explain the lack of increase in free body fat mass in the placebo group [99].
Another component affecting muscle mass and function is exercise. Resistance training programs appear to have maximal effects on the muscle mass of older adults and elders. Training with elastic bands or/and body weight as a workload at a frequency of three times weekly increased lean body mass (LBM) and lean mass of arms and legs also in pre-sarcopenic subjects 70 years of age [103], and an increase in thigh muscle thickness in men and women as well. In addition, an increase in total quadriceps and triceps arm muscle was found with the application of a resistance training program (speed: 3 s in the concentric phase, 3 s in the eccentric phase) in sedentary elderly men [41].
Optimal results in maintaining and regaining muscle mass seem to be achieved through a combination of protein mixtures and exercise types. Particularly, resistance training with a progressive increase in workload and whey protein and leucine supplemen-tation (21 g and 3 g, respectively) led to an augmentation of whole-body lean mass. No change in type I muscle fiber was observed in mass [81]. On the contrary, an elevation of SMI took place after aerobic training in older adults over 65 years old [50] with or without 4 g of n-3 fatty acid intake. Similarly, no statistically significant differences were found in the combined 3 g of n-3 fatty acid intake and lower limb resistance exercise in women over 65 years of age [82]. A combination of 10 g whey protein intake, 800 IU of vitamin D, and resistance training led to an elevation of ALM compared to control subjects [83].
Moreover, no effects on quadriceps cross-sectional area were found when carbohydrate intake was compared with protein and collagen intake under high-intensity resistance exercise conditions in community-dwelling older adults. Nevertheless, a significant elevation of quadriceps cross-sectional area was observed in the high-intensity resistance training group compared to the whey protein group [84].
Taking together all the above, regaining muscle mass is a multidimensional field. Adequate protein intake, along with vitamin D and increased physical activity through exercise, are important interventions to mitigate sarcopenia, which may contribute to prolonged independence and vitality in old age.
Protein supplementation and resistance training seem to be the main factors, but not the only ones, for muscle mass growth. The optimal combination has not been found so far. Global dietary recommendations for older adults and elders have not been established to date.

Conclusions
Changes in body composition associated with aging include a gradual increase in total adipose tissue mass, redistribution of adipose tissue with a preference for muscle fibers and internal organs, and a reduction in peripheral fat. Total skeletal muscle mass reduction is up to 80% from the age of 20 to the age of 80. There is a need to determine the optimal type, amount, timing, and frequency of both the required exercise and dietary intake and supplementation of proteins, amino acids, and vitamins in middle-aged and elderly people and then examine the clinical effectiveness in improving the primary outcomes of sarcopenia.
By better understanding the impact of physical activity and the importance of nutrient sources, particularly protein and amino acids (>1 g/kg bw/day), on human muscle turnover, it will be possible to develop better strategies and new multidisciplinary approaches, combining exercise and nutrition, to address the physiological age-related loss of muscle mass caused by numerous age-related comorbidities.
The intake of nutrient supplements should also be evaluated based on the individual nutrient needs of each person. As a different mixture of molecules is involved in different stages of muscle metabolism, it will be necessary each time to create an individually specific mixture of ingredients according to human needs. Possible future research should be directed in this direction.
Type, frequency, intensity, and timing of exercise combined with an adequate highquality protein and micronutrient intake seem to be reliable ways of facing muscle wasting by aging. However, the results are quite controversial and contradictory in many cases, which is probably due to several limitations and variations from study to study, such as the heterogeneity of the samples studied, the way muscle mass is measured, the time and mode of intervention, and comorbidities in the population concerned. Additionally, there is a dose-response relationship between exercise and muscle mass improvement. Time has been considered one of the main factors for muscle adaptation. Thus, a 12-week-duration exercise program [41,42] has the optimal effects on muscle mass, and detraining after three months of participation in an exercise program is unavoidable and takes place in a short period of three weeks. Studies focusing on exercise velocity demonstrate maximal effects on muscle mass regain in elders, while workload plays an important role in muscle mass increase in older people [41][42][43].
There is a need for larger scale and high-quality randomized controlled trials with longer follow-up and standardized primary results (muscle mass, muscle strength, and physical performance), investigating the role of exercise, protein, amino acids, and vitamins in the prevention and treatment of sarcopenia. More randomized control trials are needed to confirm physical activity and diet recommendations to fight muscle loss.

Data Availability Statement:
The data presented in this study are available on request from the corresponding author.

Conflicts of Interest:
The authors declare no conflict of interest.