This review summarizes the literature describing the significance of various conditions, such as hypoxia, oxidative stress, and, above all, physical exercise, in the hypoxia-inducible factor-1 (HIF-1) and vascular endothelial growth factor (VEGF) signaling pathway mainly in skeletal muscle. HIF-1α acts as a master regulator for the expression of genes involved in the hypoxia response of most mammalian cells. Namely, HIF-1α initiates transcription of various hypoxia-adaptive genes, such as angiogenesis, glycolysis, and erythropoiesis, after the formation of heterodimer with HIF-1β. Among them, VEGF is the most potent endothelial specific mitogen, which recruits endothelial cells into hypoxic foci and avascular area and stimulates their proliferation. The study on acute exercise shows that several components of the HIF-1 pathway, involving VEGF and erythropoietin, are activated in response to acute changes in oxygen demand in human skeletal muscle, suggesting that oxygen sensitive pathways could be relevant for adaptaion to physical activity by increasing capillary growth. Also, the effects of endurance training on the activity of the HIF pathway in human skeletal muscle under hypoxic conditions appear to be definitely higher than those under normoxic conditions, indicating that combining hypoxia with exercise training appears to improve some aspects of muscle O2 transport and/or metabolism. On the other hand, increased levels of reactive oxygen species (ROS) due to physical exercise induce the expression of peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α), which regulates mitochondrial biogenesis in multiple cell types, resulting in increases in VEGF expression and subsequent angiogenesis, strongly suggesting HIF-1α-independent regulation of VEGF and angiogenesis. Thus, the precise relationship among exercise, the HIF-1 pathway including VEGF, PGC-1α, and ROS needs further study.
The skeletal muscles in animal models with metabolic syndrome and lifestyle-related diseases, e.g., type 2 diabetes, have decreased oxidative capacity, which is associated with reduced expression levels of the gene for peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α). Running restores decreased oxidative capacity in the skeletal muscles of rats with metabolic syndrome and type 2 diabetes, and hypertriglyceridemia and hyperglycemia are improved, respectively. Longer running distances are associated with increased oxidative capacity and PGC-1α mRNA levels in the skeletal muscles of rats with metabolic syndrome and type 2 diabetes. In contrast, a high-fat diet reduces the muscle oxidative capacity and PGC-1α mRNA levels of rats with metabolic syndrome, and weight gain, hypertriglyceridemia, and hyperglycemia are accelerated. Recently, mild hyperbaric oxygen at 1.25 atmospheres absolute with 36% oxygen has been proven effective for hypertension and type 2 diabetes. Mild hyperbaric oxygen inhibits a growth-related increase in glucose levels and further decreases high glucose levels in adult rats with type 2 diabetes—changes that are associated with improvements in muscle oxidative capacity and PGC-1α mRNA levels. This paper is a review of the fiber type distribution, oxidative enzyme activity, mRNA levels, and capillary architecture of skeletal muscles in animal models with metabolic syndrome and lifestyle-related diseases. In addition, the effects of a high-fat diet, exercise, and mild hyperbaric oxygen have been summarized.
The responses of soleus muscle and/or muscle fibers to gravitation loading and/or unloading and the roles of mechanical stress and/or neural activity in the regulation of morphological properties in the soleus muscle were briefly reviewed. The soleus muscle, one of the ankle plantar flexors, shows tonic activity during body weight support against gravity (weight bearing exercise?). Passive shortening of muscle length and inhibition of the electromyogram (EMG) activity level are noted if rats are exposed to microgravity or hindlimb-unloading by tail suspension, which lead to severe atrophy in soleus muscle fibers. Previous studies reported that the unloading-related inhibition of mechanical stress and/or neural activity caused down-regulation of muscle mass. On the contrary, functional overload by amputation of the peripheral tendons or ablation of the synergists (plantaris and gastrocnemius) increased force production and sensory nerve activity in the soleus muscle, resulting in fiber hypertrophy. However, deafferentation at the L4-5 segmental level of the spinal cord down-regulated muscle mass, although the loss of muscle mass was not induced if the functional overload was performed in addition to deafferentation. These observations indicate that both mechanical stress and neural activity play important roles in the regulation of soleus muscle mass. It is well known that the level of protein synthesis and degradation strongly affects muscle mass. The mechanical stress-induced recruitment of satellite cells is also one of the important pathways in up- or down-regulation of fiber size. The essential roles of mechanical load applied to muscle fibers and neural activity of the muscle itself were clearly suggested by the reports cited in the present review.
For many locomotor behaviors, such as walking or running, we count on subliminal somatosensory information to smoothly maintain on-going movement and avoid falling down when disturbances are presented to the stability of the body. Reflex responses induced by disturbances to stability play important roles in generating quick corrective responses. Reflex outputs to the arm and leg muscles generated by muscle and cutaneous afferents during locomotor movement show quite different features compared to those generated by simple voluntary contraction during sitting or standing, irrespective of the similar background activity of motoneurons. In particular, the excitability of cutaneous reflex pathways elicited by the electrical stimulation of low threshold mechanoreceptors on the skin is strongly modulated in a phase-, nerve-, task-dependent manner during locomotor movement. The pattern generating system in the spinal cord, which has been studied intensively in quadrupedal animals, may be responsible for both generating locomotor movement and reflex modulation even in humans. However, due to methodological difficulties, the accumulated evidence derived from human experiments is indirect. In this review, we will outline these unique features of the cutaneous reflexes during locomotor and rhythmic movements in humans.
Skeletal muscle has only recently been considered a secretory organ. Muscle-derived proteins are now termed myokines. Until date, about 20 proteins known as cytokines, growth factors, and adipokines have been reported as myokines. However, only a few studies have been able to demonstrate secretion from the skeletal muscle. Furthermore, many reports are still uncertain of whether proteins are secreted from skeletal muscle cells or from the surrounding tissue, because some studies have measured myokine concentration in blood taken from human and animal subjects, which also contains other organ-derived proteins. Secretion of some myokines is promoted by muscle contraction or insulin stimulation, whereas others seem to be constitutively secreted. The mechanisms of action and roles of myokines are also complicated. Some are believed to affect distant organs through endocrine and paracrine mechanisms, while others affect organs through an autocrine mechanism. In this article, we review updates of myokines, including their history. Furthermore, the article discusses the need to re-define myokines in order to avoid possible misunderstandings because of insufficient data.
Activation of 5′-AMP-activated protein kinase (AMPK) in skeletal muscle is implicated in the multiple health benefits of exercise, including its anti-obesity, anti-diabetic, and anti-aging effects. AMPK consists of three subunits (α, β, and γ), among which the catalytic α subunits (α1 or α2) display distinct activation patterns in response to various stimuli in skeletal muscle. It was widely accepted that α2-containing AMPK (AMPKα2) is more dependent on AMP than α1-containing AMPK (AMPKα1); and that, as muscle energy status decreases, AMPK is activated in the order of AMPKα2 at lower intensities and AMPKα1 at higher intensities. On the other hand, AMPKα1 can be activated in the absence of apparent energy deprivation, indicating that AMPKα1 is a metabolic sensor that responds to low-intensity muscle contraction. Preferential activation of AMPKα1 is found in rat skeletal muscles stimulated ex vivo not only by low-intensity electrical contraction, but also by H2O2 and caffeine, which are well-known metabolic stimulators of skeletal muscle. Although further research is required to verify this hypothesis, AMPKα1 may be one of the signaling intermediaries that lead to the health-promoting effects of low-intensity exercise in daily life.
Physical inactivity contributes to type II diabetes, cardiovascular disease, depression, dementia, and cancer, defined as the “diseasome of physical inactivity”; however, there is no exercise training regimen broadly available in the field to prevent such diseasome. The reasons are that there is no database on the effects of exercise training according to inter-individual variations in physical fitness, disease, and genetic background. The authors have developed interval walking training, a portable calorimeter, and the e-Health Promotion System, which enables one to develop a database to provide the most appropriate exercise prescription for individuals to prevent diseasome. Also, it will enable exercise prescriptions to evolve to the level of the current nutritional prescription system which is broadly used in hospitals and health centers by dieticians and nurses and supported by national health insurance.
Humans have physiological, intellectual, and cultural capabilities to maintain viable body temperatures under several conditions. We do exercise in daily living for labor, health, and just fun. However, exercise is one of the strong factors disturbing the maintenance of body temperature. Some conditions, such as heavy exercise in thermal extremes, could rapidly lead to dangerous internal temperatures. Body temperature constancy is achieved by two major processes: i) behavioral processes of maintaining or searching for a preferable environment and ii) autonomic processes, e.g. vasodilation of the skin, sweating and shivering. The thermal load posed by the environment or by heavy exercise may be too great for the capacity of the regulators. Or, the regulator could be deranged due to extreme temperatures. Accidents during exercise often involve a compromise of many aspects of thermal balance; and an altered body temperature is very likely. In the present review, recent knowledge is looked at about the thermoregulation system, focusing on its defense mechanisms against the thermal load due to exercise and pathophysiological reasons for incidences of abnormal body temperature during exercise.
To learn the mechanisms underlying resistance exercise-induced muscle hypertrophy, recent studies on muscle protein metabolism and myogenic progenitor cells were reviewed. Numerous studies have suggested that activation of the translation process plays a major role in a resistance exercise-induced increase in muscle protein synthesis, and also in muscle hypertrophy after a prolonged period of training. Among regulators of the translational activity, the mTORC1 signaling pathway has been shown to be important, although the relation between its upstream regulation and exercise regimen remains unclear. In addition, the muscle satellite cells play a part, even if not indispensable, in exercise-induced muscle hypertrophy, by supplying muscle fibers with new myonuclei. Middle to high exercise intensity has been regarded as essential for gaining muscle mass, because it causes the recruitment of large motor units with fast, type II muscle fibers, which are readily hypertrophied through activation of mTORC1 signaling. However, several studies have shown that low-intensity resistance exercises with either large exercise volume or prolonged contraction time effectively activate protein synthesis and induce muscle hypertrophy. These findings suggest that various strategies are possible in exercise regimens, and exercise intensity is not necessarily a primary factor for gaining muscular size.
Skeletal muscle, fascia, tendon, cartilage, and bone are all of mesodermal origin and constitute a continuous musculoskeletal system. It is necessary that this locomotive system be sufficiently robust to support mechanical impacts and body weight, and to allow friction-free joint movement. As a strategy to fulfill these apparently conflicting requirements, tissues of locomotive systems are proposed here to utilize differential water states and their transitions, on the basis of analyses of structure and function of skeletal muscle and cartilage. Although many studies on physical fitness and sports medicine are now focusing on functional aspects of cytoskeletons and solutes such as proteins and signaling molecules, another breakthrough may be possible from conscious awareness of the role of water molecules supplying a universal field for functioning molecules.
In the present review, how to measure motor imagery ability, brain activity during motor imagery, the benefits of motor imagery practice, and the influence of sensory inputs on motor imagery, are summarized. First, the classification of motor imagery is explained. Many methods have been utilized to evaluate motor imagery ability. For example, questionnaires, mental chronometry, and mental rotation tasks have been used in the psychological approach. Brain activity has been measured utilizing transcranial magnetic stimulation (TMS), functional magnetic resonance imaging (fMRI), positron emission tomography (PET), and electroencephalography (EEG). Some brain regions are activated motor execution in both and motor imagery, including the supplementary motor area (SMA), the premotor cortex (PM) and the parietal cortex. Although motor imagery is done without movement or muscle contraction, sensory input from the periphery interacts with motor imagery. Brain activation during imagery of an action, as assessed by TMS, is stronger when sensory inputs resemble those present during the actual execution of the action. Many studies have provided evidence of the effects of motor imagery practice on basic motor skills and sport performance. Most elite athletes (70-90%) report that they use motor imagery to improve performance, and professional players, as compared to amateurs, utilize imagery practice more often. Many studies have confirmed that motor imagery practice can also be useful not only in sports, but also for improving performance in patient rehabilitation programs.
Innate immunity is our first line of defense against infectious pathogens. One of the major constituents of the innate immune system is macrophages that perform important phagocytic, regulatory and antigen presentation functions. Physically active individuals are reportedly less susceptible than sedentary individuals to viral and bacterial infections, suggesting that exercise improves immune function. Exercise increases the release of catecholamines, glucocorticoids and other factors that have immunomodulatory effects. For example, proinflammatory cytokine production from macrophages was inhibited by β2-adrenergic receptor (β2AR) agonist catecholamines, but exercise training down-regulated the expression of β2AR in macrophages, improving innate immune functions. In addition, the generation of suppressor macrophages by glucocorticoids that are normally induced during acute cold stress was inhibited in swimming-trained mice. Also, an inactive lifestyle leads to the accumulation of visceral fat, which is accompanied by adipose tissue infiltration by pro-inflammatory immune cells, mainly macrophages, and the development of a low-grade systemic inflammatory state. Exercise decreases chronic low-grade systemic inflammation and improves insulin sensitivity in obese individuals. Although the mechanisms behind these favorable effects of exercise are not fully understood, recent studies have shown that exercise training prevents adipose tissue infiltration by pro-inflammatory macrophages and that ghrelin expressed in macrophages functions as a mediator of the anti-inflammatory effects of exercise training. In this Review, the authors focus on the known mechanisms by which exercise exerts its effects on macrophages, and discuss the implications of these effects for the prevention and treatment of disease.
Many researchers have been interested in the effects of heat stress on skeletal muscle. Recently, it has been demonstrated that heat stress on skeletal muscle results in an increase in muscle protein mass and subsequent muscle hypertrophy, and attenuation of muscle atrophy. Although the cellular mechanism remains unclear, it is thought that heat shock proteins (HSPs), which are highly conserved proteins induced by heat stress, play a major role in these phenomena. However, new insights suggest that heat stress-induced muscle hypertrophy and the prevention of muscle atrophy may be regulated not only by elevated HSP expression but also by multiple signaling pathways associated with protein synthesis and breakdown. Additionally, heat stress seems to cause various changes in other muscle functions. Although further studies are required to reveal the molecular biological mechanisms involved in the heat stress-induced changes in skeletal muscle, heat stress may be a useful tool for increasing muscle mass, attenuating disuse skeletal muscle atrophy, facilitating an early recovery from muscle damage, and improving glucose metabolism. This paper reviews studies of these effects of heat stress on skeletal muscle.
Iron deficiency is still a problem in developing countries as well as developed countries. Moreover, an athletic population has a high rate of iron deficiency anemia; most of whom participate in aerobic exercise. Therefore, most studies which have been conducted to investigate the effect of exercise on iron status have used aerobic exercise. A considerable number of studies suggest that aerobic exercise has harmful effects on the iron levels in the body. Therefore, most of the studies on improving iron deficiency focus on the effect of nutrition. However, mild resistance exercise improves iron status in the body. These results suggest the possibility of a difference in the effect of different types of exercise on iron status. Exercise would be economically advantageous as a measure to improve iron deficiency in developing countries that have a high rate of iron deficiency. Future studies might thus be needed to clarify the relationship between iron status and physical activity.
In this review, we discuss the anabolic and metabolic responses of skeletal muscles to β2-agonists and exercise. β2-agonists increase muscle mass, particularly in fast-twitch muscles. Exercise positively regulates glucose homeostasis, mitochondrial biogenesis, and metabolic enzyme levels in skeletal muscles; whereas treatment with β2-agonists attenuates these beneficial effects. This review also describes the role of β2-adrenergic receptor (β2-AR) signaling molecules, such as cyclic adenosine monophosphate response element-binding protein, mitogen-activated protein kinase, and Akt/protein kinase B, in the response of skeletal muscles to β2-agonist treatment and exercise. For example, β2-agonists and exercise increase the phosphorylation of p38 mitogen-activated protein kinase in slow-twitch muscles. Our interpretation of these findings is that β2-adrenergic receptor signaling plays a functional role in the anabolic and metabolic responses of skeletal muscles to β2-agonists and exercise.
Minor amounts of glucose incorporated into the cells is metabolized via the hexosamine biosynthetic pathway, resulting in the production of uridine-5’-diphosphate-N-acetylglucosamine (UDP-GlcNAc), which is utilized as a donor substrate for the N- and O-linked glycosylation of extracellular and membrane proteins in the Golgi apparatus, or for the O-linked GlcNAc modification (O-GlcNAcylation) of intracellular proteins in the cytosol. In particular, O-GlcNAcylation, the addition of GlcNAc to serine/threonine residues, is reversibly regulated by the enzymatic activities of O-GlcNAc transferase (OGT) and O-GlcNAcase. Since OGT activity is sensitive to the intracellular UDP-GlcNAc level, flux via the hexosamine pathway influences O-GlcNAcylation. Actually, excess flux via the hexosamine pathway and resultant increased O-GlcNAcylation are associated with several pathophysiological conditions, including insulin resistance. To date, there are several studies addressing the effects of exercise on the hexosamine pathway and O-GlcNAcylation. Thus, this short review focuses on the relationships among exercise, the hexosamine pathway, O-GlcNAcylation, and insulin resistance.
Skeletal muscle is the most abundant tissue in the mammalian body and is composed of multinucleated fibers that contract to generate force and movement. In addition, skeletal muscle has the ability to regenerate following severe damage by exercise, toxins or disease. Regeneration is possible because of the presence of mononucleated precursor cells called satellite cells. After injury, satellite cells are activated, proliferate, and fuse with the damaged fibers or fuse together to form new myofibers. A fraction of satellite cells self-renew and behave as muscle stem cells. Although satellite cells are the main players in muscle regeneration, a number of other cell types are also recruited to form new fibers or to modulate the behavior of satellite cells. Here we present an overview of current knowledge of regeneration focusing on muscle satellite cells and other stem cells and discussing promising stem cell therapy for diseases such as muscular dystrophy.
Warm-up routine prior to sports activities is a customary practice among sporting people and athletes. Although warm-up is done with the expectation that it will enhance subsequent performance, little is known about how one should warm-up. In addition, scientific evidence is lacking on the best procedure that would allow for optimal preparation for a given event, especially for explosive sports activities such as jumping and sprinting. Recently, there has been considerable research on the functional significance of activity-dependent potentiation in dynamic muscular performance. Interest has evolved around whether high-intensity, short-duration muscle contraction as a warm-up procedure can improve such performance, although it remains inconclusive since muscle contractions induce fatigue as well as potentiative effects. In this review, based on evidence from well-controlled studies, proper warm-up procedures for enhancing voluntary dynamic muscular performance are proposed from the perspective of both fatigue and potentiative effects.
Autophagy is a process in which lysosomes participate in the degradation of cellular proteins and organelles, and is essential for cell survival. Recent studies of autophagy have focused on its role in skeletal muscle homeostasis. Autophagic flux in skeletal muscle is induced not only by atrophic stimuli such as fasting and denervation, but also by physical exercise. Excessive activation of autophagy promotes the progression of muscle atrophy. In contrast, impaired or deficient autophagy appears to promote muscle weakness, myopathy, and age-related muscle atrophy due to the accumulation of denatured proteins and damaged organelles. The induction of autophagy during exercise is believed to play a role in physiological adaptations to exercise, such as increased oxidative capacity and insulin sensitivity. Thus, it is important for skeletal muscle homeostasis to appropriately regulate autophagy.
Protein synthesis is suppressed in working skeletal muscle. Teleologically, the skeletal muscle halts an ATP-consuming anabolic process such as protein synthesis to spare ATP for contractile activity during an emergency. So far, 2 mechanisms have been proposed for halting protein synthesis in working muscle. One of these mechanisms suggests that AMPK inhibits mTORC1, which is arguably a master regulator of the initiation step in protein translation. Another theory suggests Ca2+-dependent inactivation of eukaryotic elongation factor 2 (eEF2), which regulates the elongation step in protein translation. Previous reports in the literature suggest that factors other than AMPK and/or eEF2 are involved in the suppression of protein synthesis. We have recently shown that REDD1 might also be involved in blunting protein synthesis in working muscle. Understanding these mechanisms might lead to the development of new strategies and treatments, not only for athletes but also for individuals with muscle-wasting conditions such as sarcopenia.
A physically active lifestyle is important across the entire life span. However, little is known about life-long participation in regular exercise among older people. The purpose of the present study was to describe regular exercise throughout a person's lifetime and evaluate the impact of exercise earlier in life on participation in exercise at age 60 and over. The participants were 984 community-dwelling older people aged 60 to 86 years. Each participant's life was divided into five age categories: 12-19, 20-29, 30-39, 40-59, and 60 years and over. The association between exercise at an earlier age and that at 60 years and over was assessed using logistic regression analysis adjusted for potential confounders. Men had exercised throughout their lives more than women. Among women, participation in exercise during their 20s and 30s showed a sharp decline. The preference for exercise differed according to age and gender. Among men, the most common patterns of exercise throughout life were exercise during all the age categories, and starting exercise at age 60 and over; whereas in women the most common pattern was no exercise at all. The adjusted odds ratio of exercise at 40-59 years for exercise at age 60 and over was 5.85 (95% confidence interval: 3.82-8.96) among men and 6.89 (4.23-11.23) among women. Regular exercise in the younger age categories affected exercise at age 60 and over among men, but not among women. Regular exercise at 40-59 years was a strong predictor of exercise at 60 years and over in both men and women.
The aim of the present study was to examine the relationship between human β-defensin-2 (HBD-2), cathelicidin (LL-37) and upper respiratory tract infections (URTI). In addition, the possible association between salivary cortisol and the salivary antimicrobial peptides was also examined. We hypothesized that the saliva levels of HBD-2 and LL-37 are lower in elite marathon runners; and that saliva cortisol levels might have a negative association with saliva HBD-2 and LL-37. Twenty elite male marathon runners were studied, and twenty additional male subjects were used as sedentary controls. Saliva samples were collected between 12:00 and 14:00 in the afternoon. We selected the cotton swab method of saliva collection. Elite marathon runners tend to have lower concentrations of salivary antimicrobial peptides (HBD-2 and LL-37) than sedentary subjects. Saliva cortisol levels in the elite marathon runners were significantly higher than those in the sedentary subjects. Concentration of saliva cortisol levels showed a negative correlation with saliva HBD-2 and saliva LL-37 concentration levels. Number of URTI in the elite marathon runners was significantly higher than in the sedentary subjects. Number of URTI was negatively correlated with saliva HBD-2 concentration and saliva LL-37 concentration levels. The present findings suggest the relationship between antimicrobial peptides and URTI in elite marathon runners and sedentary subjects. In addition, salivary antimicrobial peptides in the elite marathon runners were significantly lower than sedentary control subjects. It is possible that, while strenuous exercise in elite athletes could partly enhance oral innate immunity, the physical stress could simultaneously restrict the immunological enhancement due to HPA axis activity.
The purpose of this study was to investigate the effects of three different intensities of exercise on oxidative stress and antioxidant capacity. Eight healthy male subjects performed three different intensities of exercise for 20 minutes: (1) 70% anaerobic threshold (AT) (light intensity, LI), (2) 100% AT (moderate intensity, MI), and (3) 130% AT (high intensity, HI) on a cycle ergometer. A control (C) trial was conducted under resting conditions. Blood samples were taken pre-exercise, immediately and 30 min after exercise. In the HI trial only, the concentrations of plasma derivatives of reactive oxygen metabolites (d-ROMs) were significantly higher immediately (P<0.01) and 30 min after exercise (P<0.05) than at pre-exercise. Furthermore, plasma trolox equivalent antioxidant capacity was significantly higher immediately after exercise in the HI trial (P<0.01). And, plasma glutathione peroxidase (GPX) activity was significantly higher immediately after exercise than at pre-exercise in the MI (P<0.05) and HI trials (P<0.01). These data suggest that 20 min of pedaling exercise above the AT level may increase ROS production, while endogenous enzyme activity (especially GPX activity) or non-enzymatic antioxidants may modulate exercise-induced ROS generation. Thus, exercise under the AT level induces very little oxidative stress damage in young people.
Habitual exercise is important for improving or maintaining arterial function with age. However, the role of physical activity on arterial stiffness in adolescents is unclear. This study evaluated the influence of physical activity on the brachial-ankle pulse wave velocity (baPWV) in high school students. The baPWV, brachial systolic blood pressure (SBP), diastolic blood pressure (DBP), and body composition were assessed in 221 healthy male high school students (16.4 ± 1.5 yrs). Self-reported physical activity (PA) was evaluated by the short form of the International Physical Activity Questionnaire (IPAQ-SF). Measurement variables were calculated for four age stages (15, 16, 17, and 18 yrs). The values for baPWV, SBP and DBP increased from 15 to 18 yrs. Stepwise regression analysis showed that SBP (β = 0.496), PA (β = -0.170), age (β = 0.186), and weight (β = -0.133) were independent contributors to baPWV, accounting for 36.6% of the variability. These results support the concept that physical activity lowers baPWV, which indicates an improvement in the arterial function of adolescents as well as middle-aged to elderly people.