Effects of Aerobic Exercise on Gene Expression of Skeletal Muscle Related to Aging and Prevention of Chronic Diseases in the Elderly

Article information

Asian J Kinesiol. 2025;27(3):65-74

Publication date (electronic) : 2025 July 31

doi : https://doi.org/10.15758/ajk.2025.27.3.65

Tenglong Fan¹, Man-Hsu Lin², Siyun Kim², Kijin Kim^3,

¹Department of Physical Education, Tongling University, Anhui, China

²Department of Sport Marketing, Keimyung University, Daegu, Republic of Korea

³Department of Physical Education, Keimyung University, Daegu, Republic of Korea

^*Correspondence: Kijin Kim, Department of Physical Education, College of Physical Education, Keimyung University, 1095 Dalgubeuldero, Dalseo-gu, Daegu, Republic of Korea; Tel: +82-053-580-5256, Fax: +82-053-580-5314; E-mail: kjk744@kmu.ac.kr

Received 2025 June 16; Revised 2025 July 9; Accepted 2025 July 31.

Abstract

Introduction

Since the trend of population aging has intensified with the development of society, the health problems of the elderly have become a social concern, and elderly fitness is seen as a new research trend in 2023 [1]. Although the process of aging is not easy to explain, the currently recognized signs of aging include as cellular senescence, stem cell exhaustion, altered intercellular communication, and dysregulation of nutrient sensing [2]. Denham Harman’s (1950s) free radical theory described the basic mechanisms of aging, and proposed that aging the result of the accumulation of oxidative damage from reactive oxygen species (ROS), byproducts of various cellular metabolic processes that increase with age and lead to shortened lifespan [3].

Aerobic exercise plays an important role in the aging process of the elderly and geriatric diseases. Aerobic exercise has been recognized to slow the aging process, which in-creases expression of BDNF (brain-derived neurotrophic factor) gene appears to ameliorate hippocampal atrophy in older adults, slowing brain aging [4]. The other genes which might also involve or play critical roles in aerobic exercise in slowing down the physiological process of aging still need to be discovered.

Moreover, an earlier study discovered that voluntary exercise had immunomodulatory effects after a stroke [5]. Both ECS (electroconvulsive seizure) and voluntary exercise experiments commonly induced BDNF, COX-2 (cyclooxygenase-2), SLC1A1, NPY (Neuro-peptide Y), and MIF (Macrophage migration inhibitory factor) [6]. NOTCH3 play a crucial part of inhibiting hepatocellular carcinoma and glioma cell growth [7], ameliorating cardiac fibrosis after myocardial infarction [8], and improving the development of pulmonary arterial hypertension [9]. Dementia patients are at high risk for Alzheimer’s disease [10]. TOMM40 gene expression found to be up-regulated in brains of Alzheimer’s patients after death [11], which often studied as a target gene in Alzheimer’s disease research. Aerobic exercise has been shown to improve cognition and function in Alzheimer’s patients [12], but the target genes for aerobic exercise reducing the risk of Alzheimer’s dis-ease have not yet been identified.

Although aerobic exercise has been studied in slowing down the aging process and preventing various diseases of the elderly, but genomics research in sports medicine is still in its infancy [13], bioinformatics research still few. In genomics, differential gene expression controls the development, function, and pathology of multicellular organisms, and protein interaction networks can study differential gene expression at a system level [14]. Using bioinformatics technology to find differential genes, construct molecular net-works, and find key genes that play an important role in sports has been widely recognized. With the help of bioinformatics technology, some studies have found that the differential genes of yoga exercise mainly affect immunity and circadian rhythm, and also involve apoptosis, angiogenesis, etc. [15] The molecular mechanism by which exercise training is beneficial to pulmonary hypertension has also been explained by bioinformatics techniques [16]. The application of bioinformatics technology is mainly restricted by the popularization of databases and researchers’ mastery of technology [13]. There are genetic data related to aerobic exercise in the GEO database, but few research results on exercise-related bioinformatics. More information about the effect of aerobicexercise-induced differences in skeletal muscle gene expression on aging and geriatric diseases needs to be further explored. Thus, this study investigated the effect of aerobic exercise on skeletal muscle gene expression in healthy elderly, and further provided bioinformatics support on aerobic exercise preventing chronic diseases in the elderly.

Materials and Methods

The research methods are explained as followed; the process is shown in <Figure 1>.

Figure 1.

Research process.

Subjects

The subjects retrieved from GSE9103 dataset were 20 healthy elderly people over the age of 55. The control group included 10 sedentary subjects (4 females, 6 males), and the experimental group were 10 aerobic-exercise trained subjects (4 females, 6 males) in <Table 1>. The characteristics of the subjects were detailed in the previous study [17]. The trained subjects performed at least one hour cycling or running 6 days a week, over the past 4 years or above; while the sedentary subjects exercised less than 30 minutes a day, twice a week.

Table 1.

The control group and experimental group subjects characteristics

Results

Screening for Differential Genes

The value of logFC calculated by “logFC_cutoff <-with (DEG, mean (abs (logFC)) + 2*sd (abs (logFC)))” is 0.437. Up-regulated genes and down-regulated genes were obtained under the threshold of “logFC=0.437, p value=0.05”. The volcano map can display fold-change and t-statistic at the same time, and the -log10 p value in the usual t test of the volcano map is used as the Y axis, log2 fold change as X axis [20]. In <Figure 2>, each dot represented a gene; 229 up-regulated genes were represented by red dots, and 390 down-regulated genes by blue dots.

Figure 2.

Volcano plot of differential genes. The red dots represent up-regulated genes, the blue dots represent down-regulated genes, the gray dots represent genes with no significant changes, and the black dots are formed by several overlapping gray dots.

The heat map in <Figure 3> visualized the gene expression by assigned different colors for better understanding. At the same time, the ‘heatmap’ package in R also added a clustering function to the heat map, and the expression of genes with similar expression values or between different groups can be clearly seen [21]. Among the top 50 differential genes, the distribution of the elderly sedentary group and the aerobic exercise group is shown in <Figure 3>. GSM (genospasm) 230387~GSM230396 represented elderly sedentary subjects, while GSM230397~GSM230406 were the elderly aerobic exercise group. The red parts represented up-regulated genes, blue as down-regulated genes, and the color darkness was related to gene expression. It can be found in <Figure 3> that among the top 50 differential genes, compared with the sedentary elderly group, the expression of 13 genes was down-regulated and the expression of 37 genes was up-regulated in the aerobic exercise group.

Figure 3.

Heatmap of top 50 differential genes. The red squares represent up-regulated genes, and the blue squares represent down-regulated genes. The darker the color, the more regulated of the gene.

GO Enrichment Analysis and KEGG Analysis

The GO enrichment analysis and KEGG analysis were performed on all differential genes, and the results are shown in <Figure 4>. According to the GO enrichment analysis, the biological process mainly affected by differential genes was muscle cell development. The cellular components mainly affected by differential genes were actin cytoskeleton, contractile fiber, sarcomere, myofibril, I band, Z disc. And the main molecular functions affected were actin binding, myosin heavy chain binding, muscle alpha-actinin binding. The KEGG analysis showed differential genes were mainly related to mineral absorption, microRNAs in cancer, estrogen signaling pathway, and pertussis.

Figure 4.

GO enrichment analysis: (a) Biological process, (b) Cellular components, (c) Molecular function; and (d) KEGG analysis. The size of the circle represents the number of enriched genes, and the color shows the enrichment significance p value, blue to red, large to small. In KEGG analysis, the longer the length of the rectangle, the more significant the enrichment of the pathway.

PPI Network Analysis Results of Differential Genes

122 genes were selected with “p value <0.001” according to the p value of the differential genes. Those 122 significantly different genes were imported into String app of Cytoscape, and came up with a PPI (Protein-Protein Interactions) network with a total of 29 nodes and 29 connections. The logFC data corresponding to the differential genes was imported; depending on the logFC value from high to low, the color of the node changed from red to blue (See <Figure 5a>). The degree that calculated via “analyze network” were used to determine the size of the nodes. The key genes were screened based on MCC by cytoHubba app, and the top 10 key genes were found consequently (See <Figure 5b>). Among those key genes, up-regulated genes included ATP2A1 and SLC1A1, while down-regulated were NOTCH3, RPS27A, MLLT4, ACVRL1, TOMM40L, PPIF, ENG, and PAIP2B.

Figure 5.

PPI interaction network and key gene screening: (a) PPI interaction network. Dots in pink represents up-regulated genes, while blue represents down-regulated genes; (b) MCC screening criteria. The key genes in the PPI interaction network constructed by differential genes.

Discussion

Bioinformatics technology was used in this study to mine the gene expression information of the lateral femoral biopsy samples of the elderly aerobic exercise group and the sedentary group, and the distribution of top 50 differential gene expression in two groups. Meanwhile, the screened differential genes were proved by KEGG analysis that aerobic exercise can affect mineral absorption pathways, and may also affect microRNAs in cancer, estrogen signaling pathway, and pertussis. GO enrichment analysis showed the effects of aerobic exercise on biological process, cellular localization and molecular functions in the elderly, mainly on muscle development, actin cytoskeleton and actin binding.

Muscle mass and strength decline with age; as demonstrated in this study, the biological process affected by the aerobic exercise differential gene was muscle development, suggesting that aerobic exercise can slow aging by promoting muscle development. According to Lai’s 2020 study, the actin cytoskeleton was essential for the proper function of somatic cells, stem cells, and gametes, and age-related changes in actin cytoskeleton was consider to be the focus of aging-related research in the next few years [22]. This study found that the cellular localization affected by aerobic exercise was mainly actin cytoskeleton, which provides a new explanation for why aerobic exercise slows aging.

The importance of actin binding in coronary atherosclerotic disease has been verified, and the decrease of actin binding level was found in the medial layer of atherosclerotic coronary arteries, signs of actin disorganization [23]. The main molecular functions affected by aerobic exercise in this study are actin binding, indicating that aerobic exercise also plays a certain role in the adjunctive treatment of atherosclerotic diseases.

Lean body mass loss and sedentary lifestyles that come with aging reduce elderly energy needs, but nutritional status has an impact on how quickly their physiological and functional abilities decline as they age [24]. Studies have shown differences in the metabolism of the trace minerals chromium, zinc and copper between sedentary and aerobic exercise groups. Trained athletes have lower resting urinary chromium losses, lower resting serum zinc levels and differences in copper nutrition compared with sedentary populations [25]. Most of the subhealth status and poor behavioral conditions are related to the lack of essential minerals or the excess of toxic minerals. Survey showed that the most deficient minerals are chromium, magnesium, zinc and calcium, and poor eating habits and dangerous lifestyles precede the development of most epidemic diseases [26]. Differential genes are enriched in the mineral absorption pathway, providing a new explanation for aerobic exercise slowing down the aging process: aerobic exercise slows down the aging process by interfering with the absorption of minerals.

The role of aerobic exercise in the treatment and prevention of cancer has been confirmed by many studies [27]. Among them, circulating microRNAs have been shown as potential markers of aerobic exercise [28] and physical activity to reduce cancer risk [29]. This study proved that one of the pathways mainly affected by aerobic exercise is microRNA in cancer, so it can be inferred that aerobic exercise can reduce the risk of cancer through the expression of circulating microRNAs.

The estrogen signaling mainly includes activation of intracellular estrogen receptor, and the use of estrogen receptortargeted drugs is a routine approach in the treatment of breast cancer patients. However, initial or acquired resistance to these treatments frequently occurs, leading to a recurrence of metastatic tumors [30]. This study proved that aerobic exercise differential genes affect estrogen signaling pathway, which provides a new idea for the treatment of breast cancer.

Pertussis is an acute respiratory infectious disease that poses a serious threat to infants, was once considered a childhood disease. Pertussis can also affect the lung function of adolescents and adults; vaccination has become the main method for preventing [31]. A follow-up survey found that the forced vital capacity value of adults with pertussis history in infant phase was lower than that of those born in the same period without a history of pertussis [32]. However, aerobic exercise has found effective in reducing lung symptoms in the elderly. In a study on elderly women with pertussis, body weight supported treadmill training effectively improved the lung function of this group of people [33]. This study also proved from the approach of bioinformatics that aerobic exercise-induced enrichment of differentially expressed genes in the pertussis pathway in skeletal muscle of the elderly.

PPI interaction network screened out 10 key genes (NOTCH3, RPS27A, MLLT4, ACVRL1, ATP2A1, TOMM40L, PPIF, ENG, SLC1A1, and PAIP2B) and found that ATP2A1 and SLC1A1 were up-regulated. ATP2A1 is mainly responsible for the transport of sodium ions and potassium ions [26], and also a key enzyme in calcium ion metabolism in muscle cells. The up-regulation of ATP2A1 gene expression helps reducing muscle relaxation [34]. The growth suppressor miR-126 is dysregulated with age, and there is a correlation between miR-126 inhibition and increased ATP2A1 expression in skeletal muscle [35]. Although this study cannot prove that the upregulation of ATP2A1 expression caused by aerobic exercise can slow down the miR-126 dysregulation associated with aging, the up-regulation of ATP2A1 expression induced by aerobic exercise provides an idea for regulating miR-126 dysregulation. Also, ATP2A1 might be the next target gene of slowing down aging. SLC1A1 was also found up-regulated in this study, which indicating that aerobic exercise helps the formation of voluntary movement awareness. SLC1A1 is a candidate gene for obsessive-compulsive disorder, mainly related to glutamate trans port [36]. Some researchers also found the increased expression of SLC1A1 is often accompanied by increased awareness of voluntary movement [37].

The other 8 key genes screened out via PPI interaction network (NOTCH3, TOMM40L, PPIF, RPS27A, MLLT4, ACVRL1, ENG, PAIP2B) were down-regulated. The gene expression and function of those down-regulated genes are discussed as follow:

NOTCH3 down-regulation inhibits the growth of hepatocellular carcinoma and glioma cell, ameliorates cardiac fibrosis after myocardial infarction, and improves the development of pulmonary arterial hypertension [9-12]. As an important paralog of TOMM40L, TOMM40 was found up-regulated in the post-mortem human brain of Alzheimer’s disease [38], and also been more applied in the research of Alzheimer’s disease. However, it has not been proved whether down-regulation of TOMM40L gene expression will lead to down-regulation of TOMM40, while a large number of studies have proved that aerobic exercise can improve the cognition and function of Alzheimer’s patients [39]. Thus, TOMM40L may be another target gene since aerobic exercise improves cognition and function in Alzheimer’s patients. PPIF produces Cyclophilin D, and studies showed Cyclophilin D sensitizes brain mitochondria to permeability transition. The loss of Cyclophilin D or inhibition of PPIF gene expression increased stability of mitochondria to calcium ion stress [40]. Mitochondria were also target organelles for exercise-induced cardio protection [41]. Cyclophilin D is necessary for oxidative damage inducing cell death, and down-regulation of PPIF gene expression also largely protects the body from calcium ion overload and oxidative stress-induced cell death [42].

The down-regulation of NOTCH3, TOMM40L, and PPIF genes can help the elderly to prevent Alzheimer’s disease, prevent calcium ion overload and oxidative stress-induced cell death, while the effect of the down-regulation of RPS27A, ACVRL1, ENG, PAIP2B, and MLLT4 still need to be more studied. RPS27A can promote the proliferation of leukemia cells, and regulates cell cycle arrest, promotes cell proliferation and inhibits apoptosis through p53, Raf/MEK/ERK, P21 and BCL-2 signaling pathways [43]. And, it is also an important gene for mesenchymal stem cells to treat diabetic nephropathy. ACVRL1 (ALK1), which controls angiogenesis, is a transforming growth factor-β (TGF-β) type I receptor mainly expressed in endothelial cells and plays a key role in vascular remodeling and angiogenesis [44]. Diseases associated with ACVRL1 include telangiectasia, hereditary bleeding disorder type 2, and primary pulmonary hypertension [45]. The ENG gene is expressed in proliferating vascular endothelial cells and other cell types associated with the cardiovascular system, and controls a variety of cellular processes, including cell differentiation, proliferation, angiogenesis, inflammation, and wound healing [46]. PAIP2B is a protein-coding gene, and its related diseases include retinitis pigmentosa 63 and optic atrophy 8 [47]. PAIP2B acts similarly with PAIP2A, inhibiting the translation of capped and polyadenylated mRNAs in vitro and in vivo by displacing PABP from the poly (A) tail. Besides, like PAIP2A, PAIP2B does not affect translation mediated by the internal ribosome entry site of hepatitis C virus [48].

ot only the down-regulation of the genes mentioned above need more attention of follow-up research; related researches on MLLT4, an important gene that constitutes a collection of conserved exercise regulators [49], still focus on the fact of MLLT4 as an important gene that constitutes the germ cell-Sertoli cell junction signaling, leukocyte extravasation signaling, and tight junction signaling [50]. Overall, the importance of MLLT4 gene expression has not been thoroughly studied, and no research has proven the impact of MLLT4 down-regulation. Even in people over 65 years of age, there may be some differences in body composition and physiological ability, so it is possible that aerobic exercise may show gender differences in the response process of molecular biological factors in skeletal muscles. In this regard, it has been reported that women have greater basal skeletal muscle TLR expression and differential response to unaccustomed exercise than men [51]. In addition, it was reported that miRNAs play an important role in programming sex differences in the results of analyzing the immune function index for preventing skeletal muscle diseases due to aerobic exercise in the aging process [52]. However, the results on the sex difference in gene expression of skeletal muscle due to aerobic exercise in the aging process are insufficient and it is thought that continuous research is needed in the future.

In this study, bioinformatics analysis was performed on skeletal muscle gene expression of sedentary and aerobic exercise elderly people. It proposed the gene expression mechanism of aerobic exercise to delay aging and prevent various diseases from the perspective of bioinformatics, as long as provided target genes and new research ideas for future research in this field. However, because of the limitations of the bioinformatics method itself and the limited sample size of elderly people from GSE9103, further verification is suggested. For instance, utilizing bioinformatics analysis with larger sample size, western blotting and quantitative real-time polymerase chain reaction experiments on target genes, in order to verify whether genes are highly expressed and to implement related survival analysis, etc.

Conclusions

This research found that aerobic exercise-induced differential genes affect muscle development, actin cytoskeleton, mineral absorption pathways, and down-regulation of the key gene PPIF may slow down the aging process. Also, it proposed the relationship between ATP2A1 up-regulation and agingaccompanied miR-126 dysregulation an important research direction for follow-up studies on delaying aging.

In addition, the enrichment of differential genes in actin binding molecular function, microRNAs in cancer, estrogen signaling pathway showed that aerobic exercise may help the elderly to prevent atherosclerosis, cancer, and breast cancer. And down-regulation of the key gene NOTCH3 also showed that aerobic exercise can help the elderly to prevent Alzheimer’s disease, liver cancer, and pulmonary hypertension. Follow-up studies are suggested to pay attention on the negative effects of down-regulation of key genes RPS27A, ACVRL1, ENG, PAIP2B and MLLT4.

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