Protein supplementation under resistance training conditions in elderly patients with sarcopenia: A meta-analysis of randomized controlled trials

Background: The efficacy of interventions for elderly patients with sarcopenia has received increasing attention. Exercise and nutrition have been recognized as effective treatments for sarcopenia in many studies. However, evidence-based support from relevant studies is still lacking. Methods: The PubMed, Embase, Cochrane Library, VIP, CNKI, and SinoMed databases were searched. The basis for the diagnosis of sarcopenia, general condition of the subjects, duration and methods of exercise/nutritional therapy, outcome indicators, and quality of evidence were evaluated, and a meta-analysis of differences in treatment outcomes between the groups from baseline to the end of each study was conducted. Results: A total of 1860 papers were screened, including six randomized controlled trials, and the effects of protein intervention under exercise conditions on muscle mass, strength, and function in elderly patients with sarcopenia were investigated. The results showed that protein supplementation under resistance exercise conditions had the following effects on elderly patients with sarcopenia: there was a significant difference in knee flexion and extension strength ( p = 0.02), grip strength ( p = 0.02), fat mass ( p = 0.04), and normal pace and pace ( p = 0.0008 and p = 0.0010, respectively) between the intervention group and baseline data. Conclusion: The meta-analysis revealed some positive effects of protein intervention treatment under exercise conditions on elderly patients with sarcopenia. However, the quality of evidence is low. High-quality randomized controlled trials should be conducted in the future to provide a better clinical basis.


Introduction
Sarcopenia is a progressive disease associated with advancing age in which muscle mass and/or muscle strength or physiological function, is decreased [1]. Early symptoms of sarcopenia are not typical. However, this latent and easily overlooked disease frequently causes adverse outcomes such as fractures, disability, and loss of activity in the elderly, while it increases the risk of disability and seriously that affects the quality of life [2]. Therefore, reasonable interventions should be provided for elderly patients with sarcopenia based on early detection and diagnosis, which is significant for improving the quality of life [3].
Currently, the primary treatment of sarcopenia includes drug treatment [4], nutritional intervention [5,6], and exercise therapy [7,8]. Elderly patients should consume 1.0-1.5 g/kg body weight of protein per day. Branched-chain amino acids are important amino acids that constitute muscle proteins. Whey protein is rich in branched-chain amino acids and can increase muscle mass and some muscle functions [9]. Studies have reported that exercise therapy is based primarily on resistance, rapid strength, and whole-body vibration training. Further studies have reported primary interventions such as tai chi and massage [10].
Although the clinical understanding of sarcopenia has improved, and exercise and nutrition have been found to be effective treatments for sarcopenia [11], research on the therapeutic effect of protein supplementation on senile sarcopenia patients under the condition of resistance training is lacking. This study used meta-analysis to investigate the effects of protein supplementation under resistance training conditions on the symptoms of sarcopenia in elderly patients under randomized controlled trial conditions, to provide reliable evidence-based support for the development of nutrition and exercise programs for elderly patients with sarcopenia.

Search strategy
The PubMed, Embase, Cochrane Library, VIP, CNKI, and SinoMed databases were searched. Date limits were from database establishment to April 2019. The key words the literature retrieved included "sarcopenia", "exercise", "sports" ,"physical education" ,"resistance exercise" ,"resistance training" ,"proteins" ,"lactalbumin" ,"albumin" ,"amino acids", "peptides" ,"milk proteins" and "dietary proteins" .Search strategy：Sarcopenia AND(Exercise OR sports OR physical education OR resistance exercise OR resistance training) AND (proteins OR lactalbumin OR albumins OR amino acids OR peptides OR milk proteins OR dietary proteins).

Literature inclusion and exclusion criteria
The inclusion criteria were the following: (1) elderly people aged ≥60 years; (2) participants diagnosed with sarcopenia based on the criteria established by the European Working Group on Sarcopenia in Older People, Asia Working Group for Sarcopenia, International Working Group on Sarcopenia, and Foundation for the National Institutes of Health [12]; (3) randomized controlled trial study design; and (4) the intervention was for protein supplementation under regular resistance training conditions.
The exclusion criteria were the following: (1) repeated publications; (2) diagnostic criteria for sarcopenia were experimental tools or scale assessment.
Primary outcome indices were muscle strength (including grip strength and knee flexion and extension strength), pace (including normal and maximum pace), and skeletal muscle mass.
Secondary outcome indices were limb muscle mass index, body weight, body mass index, and fat mass.

Data extraction
Two researchers independently extracted the data before table extraction and performed a crosscheck. Extracted information primarily included the author of the paper and year of publication, basis for sarcopenia diagnosis, basic characteristics of the research subjects, duration of the exercise/nutrition therapeutic intervention, details of the exercise/nutrition intervention, and outcome indices.

Literature screening and quality assessment
Two researchers independently assessed the included papers based on literature quality assessment standards in the Cochrane Handbook version 5.1.0 [13], and disagreements were resolved through consensus or expert consultation.

Statistics
Data were quantitatively combined using RevMan version 5.3 software. If there was heterogeneity among the studies (I 2 ≥ 50%), a random-effects model was used for the meta-analysis, and if the heterogeneity among the studies was small (I 2 < 50%), a fixed-effects model was used. Continuous variable data were expressed as weighted (WMD) or standardized (SMD) mean difference and 95% confidence intervals (95% CI). In this study, the final value after intervention was used as the main effect parameter. If the difference in baseline data between the intervention and control groups was too large, the difference between the baseline measurement and the final value after intervention was used as the effect parameter. The p < 0.05 standard was used for overall response, and Z-test was used for statistical analysis of the meta-analysis results. Publication bias was analyzed using Egger's or Begg's test, with p > 0.10 indicating no publication bias.

Literature search results
In the initial screening, 1781 papers satisfying the search requirements were selected, of which six were finally included after layer-by-layer screening [14][15][16][17][18][19]. The process for paper inclusion is shown in Figure 1. Basic characteristics of the included papers are shown in Table 1.  Table 1 Basic characteristics FFM, fat-free mass; DEXA, dual-energy X-ray absorptiometry; PASE, Physical Activity Scale for the Elderly.

Methodological quality assessment of the included papers
Two researchers independently performed quality assessments of the included papers based on the literature quality assessment standards in the Cochrane Handbook version 5.1.0. The results showed that the quality of the six papers was grade B. The risk of bias for individual studies is shown in Figure 2, and the risk proportion for study inclusion is shown in Figure 3.

Meta-analysis results
The included papers had intervention durations ranging from 12 to 24 weeks. The six included papers all employed a matched randomized design, of which five employed a random number table for grouping and one did not report the method of randomization. Allocation concealment was used in two papers, and specific allocation concealment was not reported or mentioned in four. A double-blind methodology was used in four papers, and two did not report the method of blinding.
All data in the results of the included paper were complete, description of the selected reports was clear, and there were no other factors that led to bias in the implementation of the study.
This meta-analysis continued to analyze whether the difference between the intervention group and baseline data was significant when data between the control and experimental groups after intervention was not significant. If the difference between the intervention group and baseline data was not significant, the analysis would not be performed. The following are the significant different of the data between the control group and the experimental group and the significant differences between the intervention group and baseline data.       (Figure 9). A reanalysis of the intervention group with baseline data also revealed no significant difference in body weight between the two groups. A reanalysis of the intervention group with baseline data also revealed no significant difference in body mass index between the two groups.

Publication bias analysis
Egger's test reported no significant publication bias existed for the included studies (p > 0.10).

Methodological quality assessment of included studies
Our meta-analysis, which aimed to evaluate the effect of protein supplementation under resistance exercise conditions in elderly patients with sarcopenia, demonstrated a significant effect of protein supplementation on increasing muscle strength and pace in these patients. Protein inherently promotes the synthesis of muscle proteins [22], especially of whey protein that is rich in branched-chain amino acids, which prevents muscle breakdown and promotes muscle synthesis [23]. A number of meta-analyses and systematic retrospective studies [24][25][26][27][28][29][30][31][32] have concluded that active strength training, including progressive resistance training, can significantly increase muscle strength in the elderly or patients with chronic ill. A systematic review [7] showed that comprehensive exercise for 3-18 months increased muscle strength and improved physical function but had no significant effect on muscle mass, which is consistent with the findings of the meta-analysis in the present study. This may be related to nerve recruitment ability [33]. As the number of motor units activated during muscle contraction reflects the state of muscle recruitment, the greater the number of motor units involved in contraction, the greater is the muscle strength. Regular, long-term exercise training continuously strengthened certain muscles in the subjects, resulting in a large enhancement in muscle recruitment and thereby an increase in muscle strength.

Effect of protein supplementation under resistance training conditions on fat mass
in elderly patients with sarcopenia compared to conventional therapy The forest plot results showed that protein supplementation under resistance exercise conditions can increase fat mass in elderly patients with sarcopenia. All subjects were aged ≥60 years and diagnosed with sarcopenia. They have lower protein absorption capacity and protein conversion rate than normal elderly individuals.
When compared at the same basal metabolic rate, increased moderate exercise combined with complete diet structure in addition to sufficient daily protein intake may cause excessive protein intake, resulting in a surplus of calories and an increase in fat mass.
4.4 Effect of protein supplementation under resistance training conditions on pace in elderly patients with sarcopenia compared to conventional therapy The results of both normal and maximum pace forest plots showed that exercise and nutritional therapy can improve the pace of elderly patients with sarcopenia. The pace of all subjects increased through exercise and nutrition, and with adequate nutrition, their ability to perform continuous muscle work also increased. Studies have shown significant improvements in the lung capacity, minute ventilation, cardiac output, and ejection fraction of elderly subjects after long-term resistance training [34].
We believe that resistance training continuously improved lung and heart function, which in turn increased the exercise ability of the subjects, resulting in increased pace.
However, the study also had some limitations, such as the inclusion of only a few studies due to language restrictions, inconsistent quality of research methods, possibility of publication bias, small sample size, and short exercise duration.
Discovering the effects of other outcome indices, such as muscle mass, may be difficult.

Conclusions
Protein supplementation under resistance training conditions has a positive effect on the treatment of elderly patients with sarcopenia. However, more high-quality, large-sample, multicenter randomized controlled trial studies are needed to further evaluate the therapeutic effects. The 6 articles included in this study all adopted intervention measures of nutrition and exercise, but the article did not mention whether an informed agreement was signed with the observation group before the intervention.

Data Availability
The data sets generated during and/or analyzed during the current study are not publicly available but are available from the corresponding author on reasonable request.

Conflicts of Interest
The authors declare no conflicts of interest.

Funding Statement
This work was supported by the Funding of the First Affiliated Hospital of the Air Force Medical University.