Overactivation of the renin–angiotensin (Ang) system (RAS) increases the classical arm (Ang-converting enzyme (ACE)/Ang II/Ang type 1 receptor (AT1R)) to the detriment of the protective arm (ACE2/Ang 1-7/Mas receptor (MasR)). The components of the RAS are present locally in white adipose tissue (WAT) and skeletal muscle, which act co-operatively, through specific mediators, in response to pathophysiological changes. In WAT, up-regulation of the classical arm promotes lipogenesis and reduces lipolysis and adipogenesis, leading to adipocyte hypertrophy and lipid storage, which are related to insulin resistance and increased inflammation. In skeletal muscle, the classical arm promotes protein degradation and increases the inflammatory status and oxidative stress, leading to muscle wasting. Conversely, the protective arm plays a counter-regulatory role by opposing the effect of Ang II. The accumulation of adipose tissue and muscle mass loss is associated with a higher risk of morbidity and mortality, which could be related, in part, to overactivation of the RAS. On the other hand, exercise training (ExT) shifts the balance of the RAS towards the protective arm, promoting the inhibition of the classical arm in parallel with the stimulation of the protective arm. Thus, fat mobilization and maintenance of muscle mass and function are facilitated. However, the mechanisms underlying exercise-induced changes in the RAS remain unclear. In this review, we present the RAS as a key mechanism of WAT and skeletal muscle metabolic dysfunction. Furthermore, we discuss the interaction between the RAS and exercise and the possible underlying mechanisms of the health-related aspects of ExT.
Overweight and obesity are major global health problems, affecting approximately 39% of the world’s adult population. The World Health Organization classifies excessive fat accumulation according to body mass index (BMI), with overweight defined as BMI greater than or equal to 25 kg/m2 and obese as BMI greater than or equal to 30 kg/m2 [1,2]. Obesity even during the first years of life can cause morbidity in adult life and lead to premature mortality . One study with a pooled analysis of more than 129 million people showed that the prevalence of overweight and obesity among children and adolescents has been steadily increasing worldwide over the past four decades more than the prevalence in adults, increasing from 0.7 to 5.6% obese girls and from 0.9 to 7.8% obese boys . This increasing trend in obesity prevalence is critical for healthcare and increases the economic burden on the healthcare system [5,6]. Obesity can reduce the quality of life and has been classically associated with metabolic syndrome , cardiovascular disease , type 2 diabetes  and increased mortality .
Many factors favour the occurrence of overweight and obesity, including environmental, genetic and functional factors. Two major risk factors that lead to the burden of obesity are unhealthy diet habits and physical inactivity – high amounts of daily screen time (e.g. computer, television, mobile phones and video-game usage) and sedentary behaviour, which are common in the current way of life [11,12]. Furthermore, regional deposits of adipose tissue may facilitate the occurrence of certain conditions that contribute to the increased cardiovascular and metabolic risk. Both subcutaneous white adipose tissue (WAT) (i.e. adipose tissue between the dermis and aponeuroses and fasciae of the muscles) and visceral WAT (i.e. within the chest, abdomen and pelvis) are associated with an increased cardiometabolic risk, although this association is more pronounced with the latter .
Excess WAT in the obese can contribute to ectopic lipid accumulation in non-adipose tissues such as the skeletal muscle. This accumulation may be toxic for the muscle (lipotoxicity), impairing its function, reducing its density and releasing pro-inflammatory cytokines, which may lead to the loss of muscle force and insulin resistance observed in the skeletal muscles of obese individuals . In addition to the increase in adiposity, aging is also associated with skeletal muscle atrophy (sarcopenia), a cause of disability and morbidity . This association between obesity and sarcopenia is called sarcopenic obesity (SO), which imposes a greater risk of adverse events than these conditions alone and cannot be detected by simple BMI analyses .
One common link between obesity and sarcopenia might be the renin–angiotensin (Ang) system (RAS). The RAS typically modulates blood pressure control and hydroelectrolyte balance  and consists of a classical arm (Ang-converting enzyme (ACE), Ang II and Ang type 1 receptor (AT1R)) and protective arm (comprising ACE2, Ang 1-7 and Mas receptor (MasR)). The activation of the RAS classical arm positively correlates with fat accumulation and muscle atrophy in both human and animal studies and the protective arm may have evolved to counterbalance the pathophysiological effects of overactivation of the classical arm [18–21]. Interestingly, all components of the RAS are present locally in both WAT and skeletal muscle, and the most likely configuration of the local RAS is a combination of local synthesis and uptake from the systemic RAS components [22,23]. These findings have important clinical relevance because patients with chronic diseases such as heart failure, chronic kidney disease, or obesity often have increased local and systemic Ang II levels, which independently worsen the outcome and negatively impact mortality and morbidity [18,24].
Therefore, preventing SO is of paramount importance on public-health scale worldwide. Lifestyle interventions such as a switch to low-calorie diets and increased physical activity are the first line of choice to prevent and treat SO . Exercise training (ExT) has been used to improve performance for centuries, and the beneficial effect of ExT to prevent and treat several pathologies is also evident . Currently, the health benefits achieved with aerobic ExT are well recognized worldwide. For decades, ExT was considered the main lifestyle intervention to prevent risk factors, treat cardiovascular diseases, metabolic dysfunction and improve the quality of life [27–29]. The ideal dose–response relationship between ExT and SO improvement, as well as the mechanisms involved in leading the physiological adaptations, remain unclear.
In this regard, this review presents the current evidence for the relevance of the RAS as a key mechanism of metabolic dysfunction in WAT and skeletal muscle and its relation to the deleterious effects of obesity and SO.
Furthermore, we discuss the interaction between the RAS and exercise and the mechanisms underlying the health-related effects of ExT.
Initially, the RAS was considered a sequence of many enzymatic steps that culminate in the production of a single biologically active metabolite, Ang II . The RAS has been studied for more than 100 years, and, in the last two decades, the identification of novel enzymes, receptors and local mediators has amplified the classical view of the RAS . One of the most important recent findings is the proposal of the RAS components as a system of two counter-regulatory arms in addition to the control of blood pressure and hydroelectrolyte balance [30,31]. A simplified view of the RAS is shown in Figure 1.
RAS overview and ExT effects
The first point in both arms of the RAS is the glycoprotein angiotensinogen (AGT), mainly produced by the liver. The enzymatic signalling starts with kidneys releasing renin, which converts AGT into a decapeptide Ang I (Ang 1–10). The renin/pro-renin receptor, upon binding to renin, induces the non-proteolytic activation of prorenin and increases the efficiency of AGT cleavage to Ang I . The next step is the Ang I conversion into an octapeptide Ang II (Ang 1–8), the main effector peptide of the classical arm, through the action of ACE . Alternatively, other enzymes can be involved in Ang II production, such as cathepsin, chymase and tonin (Figure 1) . Additionally, ACE can inactivate bradykinin, a potent vasodilator peptide with additional beneficial effects on insulin-dependent glucose transport activity .
The physiological effects of Ang II are mediated by receptors that are assigned to the G-protein-coupled receptor family, type 1 (AT1R) and type 2 (Ang type 2 receptor (AT2R)), with 33–34% homology between them [17,30]. Most of the physiological Ang II effects are mediated by AT1R, which generates second messengers such as diacylglycerol, inositol trisphosphate, NADPH oxidase and reactive oxygen species (ROS) signalling [22,35]. Activation of the classical arm (ACE/Ang II/AT1R) stimulates vasoconstriction, sodium retention, sympathetic nervous activation and ROS generation, as well as causes deleterious effects, including endothelial dysfunction and induction of inflammatory, thrombotic, proliferative and fibrotic processes [35,36]. In contrast with AT1R, AT2R in most cases seems to counteract AT1R, exerting tissue-protective effects, including vasodilation, natriuresis, anti-inflammatory and antiproliferative effects .
In the RAS counter-regulatory arm, Ang 1-7 is a direct product of Ang II degradation through the action of the ACE-homologue enzyme ACE2 or other peptidases and directly from Ang I by peptidases . Alternatively, ACE2 generates Ang 1-9 from Ang I hydrolysis and then Ang 1-9 can be cleaved by ACE, resulting in Ang 1-7 heptapeptide . However, Ang 1-7 is mostly produced through the action of ACE2 on Ang II (Figure 1). Ang 1-7 binds to a specific receptor, MasR, which is also a G-protein-coupled receptor and triggers anti-inflammatory, antifibrotic and antiproliferative actions, in general, opposing the actions triggered by Ang II [39,40].
The more recent view of the RAS shows new peptides and receptors (Figure 1), but their biological effects have not been fully explored. For instance, Ang III (Ang 2-8), formed by the action of aminopeptidase A on Ang II, is related to cardioprotective effects against ischaemic injury via AT2R . Another isoform, Ang IV (Ang 3-8) can be generated from Ang III by the action of aminopeptidase N and binds to AT4R, which is associated with insulin-regulated aminopeptidase (IRAP). Activation of AT4R/IRAP seems to be involved in cognition, memory and glucose homoeostasis because IRAP is linked to the glucose transporter (GLUT) 4 (GLUT4) [22,42]. In addition, Ang A exerts a similar vasoconstrictive effect via AT1R . Lautner et al. (2013)  discovered a new heptapeptide, alamandine, which can be formed directly from Ang 1-7 or by the catalysis of Ang A via ACE2. Alamandine acts on the Mas-related G-protein-coupled receptor (Mrg) D (MrgD) and exerts similar effects to Ang 1-7, such as vasodilation, antihypertensive and antifibrosis effects (Figure 1) [31,44]. Another metabolite of Ang 1-7 is Ang 1-5, catalysed by ACE stimulating atrial natriuretic peptide secretion via MasR .
These discoveries bring into perspective new possibilities of an interaction among RAS components. Furthermore, local RAS components have been found in almost every tissue, including the heart, blood vessels, kidney, brain, pancreas, liver, WAT and skeletal muscle, functioning as independent mechanisms, such as in endo-, para- and autocrine signalling . Below, we briefly review some recently identified aspects related to the actions of both RAS arms, especially in WAT and skeletal muscle.
RAS and WAT
Adipose tissue is widely distributed throughout the body and can be grouped according to anatomical, functional and radiological characteristics. The type of adipose tissue plays an important role in the pathogenesis of obesity. WAT, abundant during adulthood and proportionally larger throughout the aging process, stores mainly triglycerides in a single droplet (unilocular cells with a decentralized nucleus and few mitochondria). While brown adipose tissue (BAT), abundant in newborns, reduces with aging, and stores fat in multiple small vesicles (multilocular cells and rich in mitochondria) .
Based on their characteristics, the different types of adipose tissues present distinct functions. Briefly, WAT is responsible for stocking energy in triglyceride form and releasing them when necessary; in contrast, BAT plays an important role in energy dissipation through heat production (i.e. thermogenesis) . Recently, beige adipocytes were described in rodents and humans as white cells that, under stimuli (e.g. cold exposure, adrenergic stimulation and exercise), acquire a brown fat-like phenotype in a process called browning . In obesity, the accumulation of visceral WAT appears to play a more significant pathogenic role, while subcutaneous WAT appears to be protective and is the preferable site for browning [48,49].
WAT is currently considered an endocrine organ that plays an important role in energy metabolism. The two primary metabolic functions of WAT are lipogenesis and lipolysis, although this tissue is known to produce and secrete a wide variety of signalling substances, including leptin, adiponectin, visfatin, resistin, interleukin (IL)-6 and tumour necrosis factor (TNF) α (TNF-α). Combined, these substances are called adipokines .
In agreement with the endocrine function of WAT, this tissue has been reported to express several components of the RAS. AGT is expressed in adipose tissue [51–53], and in rodents, approximately 30% of circulating AGT levels is provided by WAT . Renin expression in adipocytes was observed by some studies  but not others , while ACE expression was also detected [52,53]. Finally, AT1R has been identified in adipocytes , as well as AT2R, especially in differentiating adipocytes [55,56]. More recently, the presence of the Ang 1-7/MasR arm in WAT was also described , as well as the (pro)renin receptor . In addition, adipocytes and preadipocytes contain AT4R-IRAP , and, in adipocytes, alamandine acts via MrgD, supressing leptin expression and secretion, thereby exhibiting opposing actions to Ang 1-7/MasR .
As endo-, para- and autocrine mediators, the local RAS regulates both WAT and systemic metabolism, as well as blood pressure, energy balance and whole-body glucose homoeostasis . Studies have shown that both AGT and Ang II are adipokines released in high concentrations into the bloodstream in obesity (particularly by excess visceral fat), contributing to high systemic levels and being associated with hypertension and other abnormalities . In the next topics, the effects of WAT RAS on lipid storage, adipogenesis, inflammation and insulin resistance will be discussed. Recent literature on the interaction between adipose tissue RAS and browning will also be presented.
Lipid storage: lipogenesis and lipolysis
Briefly, lipogenesis consists of fatty acid synthesis and storage in the form of triglycerides, a task that is routinely performed by WAT. By contrast, lipolysis consists of the breakdown of triglycerides to release fatty acids in the bloodstream to be used as energy sources. The RAS classical arm plays a pivotal role in regulating lipid storage, hence promoting adipocyte hypertrophy (Figure 2). In both 3T3-L1 and human adipocytes, Ang II increases the cell lipid content, via elevated lipogenesis, enhancing fatty acid synthase (FAS) and glycerol phosphate dehydrogenase (GPDH) activity. These effects are mediated by AT2R . Transgenic mice overexpressing AGT showed adipocyte hypertrophy with increased FAS gene expression, and these effects were reversed with AT2R gene deletion (Figure 2), confirming the role of this receptor in mediating lipogenesis . On the other hand, AGT gene silencing in 3T3-L1 adipocytes reduced the expression of AGT and Ang II, culminating in reduced lipid accumulation. This gene silencing decreased GPDH and fatty acid-binding protein (FABP) 4 gene expression . Supporting these findings, AGT-deficient mice showed adipocyte hypotrophy with reduced FAS activity , and high-fat-fed mice with AGT inactivation showed reduction in lipid metabolism marker gene expression, including lipoprotein lipase (LPL) and FAS . Furthermore, Ang II infusion inhibited lipolysis in human subcutaneous adipocytes via AT1R (Figure 2). Although this effect can be partially attributed to reduced blood flow, the full mechanism remains to be elucidated .
RAS effects on WAT metabolism
The Ang 1-7/MasR arm inhibits lipogenesis and stimulates lipolysis (Figure 2). In mice treated with an ACE2 activator (diminazene aceturate), there was increased ACE2 expression, as well as decreased ACE and AGT expression, in visceral WAT. These results were associated with decreased gene expression of acetyl-CoA carboxylase (ACC) and FAS, culminating in reduced fat pad mass . Additionally, rats transgenically modified to express Ang 1-7 showed decreased LPL expression and activity associated with reduced lipid synthesis in WAT . Primary adipocytes treated with Ang 1-7 showed increased lipolysis, and rats treated with either captopril or Ang 1-7 showed increased hormone-sensitive lipase (HSL) phosphorylation. These effects were downplayed with a MasR blocker, indicating that the effects of Ang 1-7 occur via MasR .
Aside from lipogenesis and lipolysis, adipose tissue expansion also involves the formation of new adipocytes (i.e. adipogenesis), thereby generating WAT hyperplasia . While adipocyte expansion through hypertrophy generates metabolically dysregulated cells, hyperplasia generates new small adipocytes that present a better metabolic profile . AGT appears to stimulate adipogenesis, contributing to a healthier WAT. Studies have demonstrated reduced differentiation in human preadipocytes  and in murine cells, where it was associated with reduced expression of genes involved in adipogenesis, including peroxisome proliferator-activated receptor (PPAR) γ (PPARγ) . The contribution of Ang II to adipogenesis depends on the specific receptor. Ang II through AT1R suppresses adipogenesis via extracellular signal-regulated kinase (ERK) 1/2 (ERK1/2) pathway activation and PPARγ inactivation . In primary human culture preadipocytes, Ang II adipogenesis inhibition was reversed by AT1R blockade . By contrast, Ang II via AT2R promotes adipocyte differentiation (Figure 2). In primary preadipocytes, selective AT2R activation in the initial steps of differentiation increased PPARγ expression . Likewise, Ang II promoted adipogenic differentiation in mesenchymal stem cells (the main source of adipocytes in adipose tissue) from human adipose tissue through AT2R .
Similar to AT2R, Ang 1-7 via MasR promotes adipogenesis (Figure 2). In human and 3T3-L1 preadipocytes, the RAS protective arm promoted adipogenesis via ERK1/2 pathway inhibition and activation of phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt) pathway, together leading to increased PPARγ expression and activity .
The hypertrophied adipocytes observed in obesity, quickly become deregulated. This deregulation includes an imbalance in the inflammatory status, with increased pro-inflammatory mediators, stimulation of macrophage infiltration and decreased anti-inflammatory mediators . The effects of RAS on adipose tissue inflammation may be both direct and indirect. Indirect effects are related to WAT expansion (Figure 2). In one study with mice and humans, obesity was accompanied by increased RAS components and inflammation in WAT, while malnourishment reduced these parameters . In agreement, Azushima et al. (2017)  developed transgenic mice that showed internalization of AT1R along with suppressed overactivation of tissue AT1R signalling (AT1R-associated protein (ATRAP) transgenic mice). They found that these mice, when fed a high-fat diet, were protected from adipocyte hypertrophy and, consequently, cell dysregulation. These effects included reduced gene expression of inflammatory markers (leptin, TNF-α, IL-6) and macrophage infiltration (F4/80, cluster of differentiation (CD) 68 (CD68) and monocyte chemoattractant protein (MCP) 1 (MCP1)) and increased gene expression of adiponectin . In a study with fructose-induced adipocyte abnormalities in rats, aliskiren (a renin inhibitor) improved adipocyte hypertrophy and dysregulation with concurrent reduction in visceral WAT Ang II levels. The authors observed that aliskiren promoted decreased expression of leptin, visfatin and resistin and increased the expression of adiponectin . In humans, valsartan treatment, an AT1R blocker (Ang receptor blocker (ARB)), reduced subcutaneous adipocyte size, which was associated with reduced markers of macrophage infiltration, including CD68 . Following the same logic, the protective arm, while reducing adipocyte hypertrophy, also promotes attenuation of macrophage infiltration . Similarly, Ang 1-7 treatment in adipocytes induced adiponectin secretion in a MasR-dependent way .
Studies have also shown the direct effects of RAS on inflammation. In mice with stress-induced WAT inflammation, irbesartan (an ARB) reduced pro-inflammatory markers (TNF-α and IL-6) and macrophage infiltration (F4/80, MCP1 and CD68) and restored adiponectin expression . Tsuchiya et al. (2006)  demonstrated that Ang II increased MCP1 in an AT1R-dependent manner and that the mechanism involved the nuclear factor κ B (NFκB) pathway. Accordingly, in adipocytes treated with Ang II, there was increased secretion of MCP1 and resistin, which was prevented when an NFκB inhibitor was used . Finally, Ang II-stimulated IL-6 production and secretion was blocked by an NFκB inhibitor and candesartan (an ARB). Ang II acts on NFκB through phosphorylation of the p65 subunit and, consequently, increases translocation to the nucleus (Figure 2). In the present study, an AT2R inhibitor could not prevent these effects, hence demonstrating the essential participation of AT1R in Ang II-induced inflammation . Regarding the protective arm, the effects of Ang 1-7 on adipocyte inflammation may also be due to reduced activation of NFκB  (Figure 2).
Dysfunctional adipocytes also facilitate the development of local insulin resistance, which is, at least in part, driven by the inflammatory state . Whether the WAT RAS directly modulates local insulin resistance remains a matter of debate. Some evidence suggests that, in the absence of an adipocyte abnormality, local RAS has beneficial effects on WAT insulin signalling . In adipocytes, Ang II increased insulin-mediated glucose uptake via phosphorylation of the insulin receptor (IR) subunit β, Akt phosphorylation and GLUT4 translocation to the cell membrane (Figure 2), these effects seem to be AT1R dependent . In another study, Ang II could not alter pAkt expression in adipose tissue, while Ang 1-7 increased pAkt expression . On the other hand, recent evidence has also demonstrated that Ang II increased pAkt expression in the absence of insulin, and inhibited Akt phosphorylation when insulin was present. Interestingly, this inhibitory effect of Ang II on pAkt in the presence of insulin was abolished by Ang 1-7 in a MasR-dependent mechanism . However, this issue remains to be further elucidated, but Ang 1-7 appears to be a potential insulin sensitizer (Figure 2).
In a different approach, when considering dysfunctional adipocytes, RAS promotes local insulin resistance in an indirect pathway, through actions in the inflammatory response (Figure 2). In this sense, AGT inactivation could maintain GLUT4 gene expression and improve IR substrate (IRS) gene expression in high-fat-fed mice, and these results were preceded by reduced WAT inflammation and macrophage infiltration . Similarly, in ATRAP transgenic mice fed with a high-fat diet, the reduction in adipose tissue inflammation and macrophage infiltration was accompanied by increased pAkt and GLUT4 expression . Finally, diabetic rats treated with an ARB showed amelioration of WAT insulin resistance, which was achieved by improvement in adipocyte abnormalities .
Similarly, chronic treatment with Ang 1-7 in fructose-fed rats decreased IRS-1 inhibitory phosphorylation and increased pAkt expression in a MasR-dependent way, thus restoring insulin signalling through the IR/IRS-1/PI3K/Akt pathway in WAT [87,89]. Although inflammation was not measured in this study, fructose is well described to induce WAT inflammation ; therefore, we can speculate that the results seen may be due to the effects of Ang 1-7 on inflammation (Figure 2).
The main pathway that activates browning is through the β3-adrenergic receptor, whose downstream effects lead to activation of p38 mitogen-activated protein kinase (MAPK) that, in turn, activates PPARγ co-activator 1 α (PGC-1α) . PGC-1α is a key regulator in brown adipogenesis, stimulating mitochondrial biogenesis, ultimately inducing uncoupling protein 1 (UCP1) expression . Aside from PGC-1α, other important factors in the development of beige cells are PR (PRD1-BF1-RIZ1 homologous)-domain containing 16 (PRDM16), which is indispensable for browning induction . Finally, a newly described mediator called irisin was shown to be produced by adipocytes and stimulate browning .
The studies of RAS on browning are recent, and the results are scarce. AT1R deletion was able to induce the brown fat-like phenotype with increased gene expression of thermogenic factors, including PPARγ and UCP1, in subcutaneous WAT. Moreover, AT1R deletion during adipocyte differentiation reduced genes involved in brown phenotype development such as UCP1, PGC-1α and PRDM16 (Figure 2). Meanwhile, AT2R deletion showed no alterations in these markers . Similarly, in ATRAP transgenic mice, obese animals showed up-regulation of genes involved in thermogenesis—PGC-1α and UCP1—in subcutaneous and visceral WAT compared with obese wild-type animals . Additionally, losartan (an ARB) increased browning in the subcutaneous WAT of diet-induced obese mice, with elevated gene expression of β3-adrenergic receptor, PGC-1α, PRDM16 and UCP1 . Than et al. (2017) , aside from confirming that an AT1R antagonist promoted browning, demonstrated that AT2R agonist could also increase this phenomenon. These authors showed that Ang II promoted browning in vivo and in vitro through the Ang II/AT2R axis (Figure 2), which is associated with increased PPARγ, PRDM16 and UCP1 expression. The mechanism proposed involved AT2R inhibition of ERK1/2, thus increasing PI3K/Akt and AMP-activated protein kinase (AMPK) pathways that, in turn, activated PPARγ , in a mechanism similar to that described for adipogenesis.
Regarding the RAS protective arm, Ang 1-7 was shown to counteract diet-induced obesity in mice through increased thermogenesis in subcutaneous WAT (Figure 2), but these effects appeared independent of UCP1 expression . In another study, although Ang 1-7 was able to induce UCP1 expression in white adipocytes, this effect appeared to be dependent on Ang 1-7 activation of AT2R and not on the ACE2/Ang 1-7 pathway .
In summary, the local RAS classical arm is elevated in obesity, thus contributing to the disruption of WAT . Moreover, the defective capacity to store lipids in WAT promotes ectopic accumulation of fat, mainly in the skeletal muscle. In this context, SO might hold an important pathophysiological cross-talk between the RAS in WAT and skeletal muscle .
RAS and skeletal muscle
Skeletal muscle is one of the most dynamic and plastic tissues in the human body, accounting for approximately 50% of the total body mass and taking up 85% of insulin-stimulated glucose . The main roles of skeletal muscle are force and movement generation. From a metabolic perspective, skeletal muscle is the main site for glucose and amino acid storage, contributes to maintain body temperature and energy metabolism, and is the main site of ATP resynthesis during physical activity or exercise . Furthermore, with the detection of myokines (myostatin, IL–6, insulin-like growth factor (IGF) 1 (IGF1), fibroblast growth factor (FGF) 21 (FGF21) and irisin), skeletal muscle has also been defined as endocrine organ exerting auto-, para- and endocrine effects, allowing communication of the skeletal muscle with other tissues .
The skeletal muscle fibre subtypes are classified according to the myosin heavy chain (MHC) isoforms and metabolic activity. In rodents, skeletal muscle fibres are characterized as type I (slow contraction, highly oxidative and resistant to fatigue); type II and its intermediary subtypes IIx/d and IIb (fast contraction, uses mainly glucose as the energy source and highly fatigable). In humans, the spectrum of muscle fibre types is restricted to type I, IIa and IIx . Additionally, in association with muscle fibres, satellite cells contribute to the repair of damaged myofibres and participate in muscle hypertrophy .
The balance between protein synthesis and degradation is a finely regulated process influenced by factors such as aging, nutritional status, hormonal balance, physical activity/exercise and disease . Skeletal muscle also expresses several components of RAS. AGT was shown to be expressed in the microcirculation and in myoblasts; and AGT production was observed in response to muscle stretch in both proliferating and differentiated C2C12 cells . Moreover, renin receptor and prorenin expression were observed in skeletal muscle . ACE activity and expression were observed in the sarcolemma and endothelial cells of local capillaries in skeletal muscle . Interestingly, genetic studies support the hypothesis that the ACE system is involved in skeletal muscle function and physical performance [23,108]. ACE expression seems to be unevenly distributed into different muscle fibre types, likely related to differences in muscle function . Finally, studies have shown Ang II receptors (i.e. AT1R and AT2R) throughout foetal and adult skeletal muscle fibres, satellite cells and microcirculation, in humans and rodents [110,111].
In recent years, many studies have proposed that the RAS protective arm exerts beneficial effects in skeletal muscle [109,112], such as improvement in insulin sensitivity [112–114] and anti-atrophic factor [115,116]. Additionally, Ang 1-7/MasR seems to be pivotal for muscle hypertrophy induced by ExT because ACE2 knockout mice did not show an increase in muscle diameter after ExT . Next, we will review the main pathways involved in muscle biosynthesis, degradation and homoeostasis, and Figure 3 summarizes the cross-talk among these pathways in a muscle fibre.
Effects of RAS on protein biosynthesis and degradation in skeletal muscle
Muscle physiology signalling pathways and the interaction with RAS
Insulin and IGF1 are two hormones involved in the control of skeletal muscle mass and insulin sensitivity via Akt signalling. Akt promotes the activation of two independents pathways of protein biosynthesis and degradation via the mammalian target of rapamycin (mTOR) and glycogen synthase kinase 3β (GSK3β) .
mTOR phosphorylation forms the mTORC1 and mTORC2 complex. mTORC1 activates P70 ribosomal protein s6 kinase (S6K)1 and phosphorylates 4E binding protein 1 (4EBP1) [119,120]. Both mechanisms exert a key role in promoting muscle hypertrophy  because these complexes phosphorylate Akt at Ser473, with the phosphorylation required for maximum activation of Akt in addition to phosphorylation at Thr308 by phosphoinositide dependent kinase 1 (PDK1) .
Reduced levels of IGF1 are observed in experimental models after systemic and local (e.g. skeletal muscle) Ang II infusion, and increased circulating Ang II stimulates Ser307 pIRS1 via protein kinase C (PKC) (Figure 3). In addition, the infusion of Ang II can induce the activation of caspase-3 and protein ubiquitin stimulation [122,123]. These mechanisms impair the insulin-IGF1/mTOR signalling pathway and, consequently, contribute to muscle atrophy . On the other hand, recent studies have shown that the RAS protective arm can contribute to stimulation of insulin-IGF1/mTOR signalling (Figure 3). In an in vivo disuse muscle atrophy model, the Ang 1-7/MasR arm can activate the IGF1/Akt pathway, thereby increasing IGF1 expression in addition to Akt phosphorylation . Additionally, in the skeletal muscle cell line C2C12, Ang 1-7 could decrease skeletal muscle atrophy induced by Ang II through MasR Akt-dependent phosphorylation .
Akt also phosphorylates GSK3β, which normally activates ubiquitination via up-regulation of ubiquitin ligases atrogin-I/muscle atrophy F-box (MAFBx) and muscle RING-finger protein-1 (MuRF1). Both compounds of the ubiquitin-proteasome pathway (UPP) are common denominators for the loss of skeletal muscle mass in almost all atrophic conditions .
Several studies have shown that blocking Ang II signalling results in skeletal muscle improvement (Figure 3). Losartan could restore skeletal muscle remodelling and protect against disuse in sarcopenia, through blocking the cross-talk between the classical arm and Akt . The age-related increase in Ang II levels contributes to the development of sarcopenia (Figure 3), and losartan could improve inflammation in the skeletal muscle of older mice . On the other hand, experimental evidence has shown that infusion of Ang 1-7 can increase the GSK3β phosphorylation at Ser9, which inactivates GSK3β (Figure 3) and shows opposite effect to Ang II .
The Akt signalling pathway facilitates GLUT4 translocation to the myocyte cellular membrane and glucose uptake by the skeletal muscle . Ang II abolishes insulin signalling in skeletal muscle by the inhibition of IRS1 and 2/Akt activation, as well as by GLUT4 translocation (Figure 3), which is reversed by ARB treatment [126,129]. These potential mechanisms involve the activation of ERK1/2 and NFκB signalling, leading to marked skeletal muscle atrophy [130,131]. On the other hand, infusion of Ang 1-7 in rats stimulates an improvement in insulin signalling and glucose tolerance via the Akt/GLUT4 pathway in skeletal muscle [112–114].
Forkhead Box O as an inhibitor of protein biosynthesis
Forkhead Box O (FoxO) transcription factors consist of a large family of proteins (i.e. FoxO1, FoxO3 and FoxO4) involved in skeletal muscle physiology. The FoxO isoforms are predominantly located in the nucleus; when phosphorylated, mainly by Akt, FoxO isoforms leave the nucleus and become unable to transcribe genes (e.g. atrogin-I/MAFBx and MurF1) for muscle atrophy [101,132,133]. mTORC2 phosphorylates FoxO to prevent protein degradation , and PGC-1α physically interacts with FoxO3 to suppress the expression of atrogin-1/MAFBx and MurF1 in skeletal muscle .
Ang II causes dephosphorylation of Akt, which maintains FoxO1 in the nucleus (Figure 3) to transcribe atrogin-1/MAFBx and MurF1 . Therefore, after 24 h of Ang II infusion, the atrogin-1/MAFBx and MuRF-1 mRNA expression levels are increased in the mouse diaphragm , and the Ang 1-7/MasR arm decreases Ang II-induced skeletal muscle atrophy (Figure 3) by decreasing atrogin-1/MAFBx and MurF1 levels [115,137].
Inflammation in muscle homoeostasis
Inflammation plays an important role in muscle homoeostasis. Chronic diseases contribute to increased pro-inflammatory cytokines, leading to the loss of muscle mass [138,139]. NFκB and its target genes play a pivotal role in increasing inflammation, and the activation of this transcription factor is under the influence of the ACE/AT1R arm. The Ang II/AT1R arm activates NFκB in a ROS-dependent manner (Figure 3), through phosphorylation and ubiquitination of inhibitor of κB (IκB) α . Although the Ang 1-7/MasR arm often counterbalances the Ang II/AT1R arm, the direct or indirect influence on inflammation in skeletal muscle remains unknown. Next, we briefly discuss three inflammatory cytokines whose functions are well studied in studies on skeletal muscle.
IL-6 can exert both anti- and pro-inflammatory effects. Local IL-6 production impacts skeletal muscle hypertrophy and muscle regeneration after acute stimuli. Nevertheless, persistent inflammation, as found in some chronic disease states, is associated with long-lasting elevated systemic IL-6 levels, increasing muscle wasting, very often combined with elevated Ang II levels . Thus, Ang II actions in skeletal muscle could be mediated by IL-6 (Figure 3) and include enhanced expression of atrogin-1/MAFBx and MurF-1, decreased levels of IRS-1 and pAkt, and increased suppressor of cytokine signalling (SOCS) 3 (SOCS3) expression .
In the maintenance of homoeostasis, TNF-α influences satellite cell proliferation ; however, in high levels, it induces atrophy and ROS formation . A recent study demonstrated that the increase in circulating Ang II activates the TNF-α/TNF receptor (TNFR) 1 (TNFR1) complex, which up-regulates cholesterol 25-hydroxylase (Ch25h), an enzyme that produces 25-hydroxycholesterol (Figure 3). The latter inactivates Akt, favouring GSK3β activation and, consequently, protein degradation .
Ang II/AT1R binding can directly activate transforming growth factor β (TGFβ) signalling, leading to an inflammatory state through ERK1/2 and Jun N-terminal kinase (JNK1/2) pathway activation . Furthermore, experimental evidence has already shown that Ang II induces the expression of TGFβ via AT1R (Figure 3). This process is dependent on p38 MAPK phosphorylation and an increase in NADPH oxidase-dependent ROS . Conversely, the Ang 1-7/MasR arm shows opposite effects on TGFβ signalling, also decreasing TGFβ expression in skeletal muscle .
Mitochondria and skeletal muscle health
Mitochondria contribute to muscle physiology because they are the sites of ATP production in myocytes but also produce ROS through many chemical reactions . Evidence has suggested that PGC-1α expression, a master regulator of mitochondrial biogenesis, decreases during muscle aging in different species, including humans [149,150]. Enhanced ROS production is observed in age-associated mitochondrial changes. This increase is linked to a lower mitochondrial content, down-regulation of PGC-1α and increased mitochondrial apoptotic susceptibility, which may all be involved in age-related sarcopenia .
Chronic infusion of Ang II produces mitochondrial abnormalities in the skeletal muscle of healthy mice with an increase in ROS production . Furthermore, Ang II could inhibit the expression of mitochondrial fission and fusion proteins, impairing mitochondrial dynamics in mouse skeletal muscle (Figure 3). The same study showed that Ang II reduced PGC-1α expression, as well as that of mitochondrial transcription factor A mitochondrial (TFAM), both essential to mitochondrial biogenesis . These data strongly suggest that blockade of Ang II signalling is a promising target for the treatment of muscle wasting in pathological conditions such as SO.
As seen, imbalances of the two arms of the RAS in skeletal muscle and WAT favouring the classical arm are involved in the pathophysiology of chronic diseases (i.e. metabolic syndrome and SO). Strategies to rescue the RAS balance, increasing the activation of the protective arm, can be used to ameliorate the pathological state of chronic diseases. There is strong evidence that ExT can shift the balance towards the protective arm of the RAS, thereby being the first line of choice as a non-pharmacological therapeutic strategy to improve metabolic dysfunction, muscle wasting and fat accumulation.
RAS and ExT
The metabolic and molecular responses to an acute excise load are distinct; therefore, chronic ExT generates different physiological and functional adaptations. The ExT adaptations depend on a combination of type (i.e. strength and aerobic), frequency, intensity (i.e. low, moderate and high) and volume (i.e. low, moderate and high) of the exercise performed. The ideal combination for training relies on which physical capacity should be improved.
In general, strength or resistance training is recommended to improve muscle strength and muscle resistance and to stimulate muscle hypertrophy [154,155]. In sarcopenic patients, high intensity and moderate volume of strength exercise is recommended, with appropriate progression [28,154]. Recently, women diagnosed with SO submitted to a strength training protocol showed improved strength, increased fat-free mass and enhanced physical performance in daily life tasks after 12 weeks of training . The effects of strength training for adipose tissue loss are debatable; in young healthy men, strength training does not seem to change body fat [157,158]. In obese, sedentary, middle-aged men, strength training three times a week increased leg strength and decreased blood pressure levels but did not change the body percentage of fat mass, cholesterol or inflammatory status . Therefore, strength training is considered a good strategy to improve muscle health, but there is insufficient evidence regarding fat weight loss.
Aerobic exercise (i.e. cycling, running and swimming) training is related to cardiorespiratory and metabolic improvements (Figure 4) [160,161]. Low-intensity exercise, which metabolizes predominantly fatty acids to supply the metabolic demand, when performed for a long period of time with a volume of more than 300 min a week, is known to induce fat weight loss and improve metabolism (i.e. plasma cholesterol and glucose levels, insulin sensitivity and inflammatory status), even without weight loss . Continuous high-intensity exercise, in which carbohydrate (i.e. glucose) metabolism predominates as an energy source, besides weight loss and metabolic improvements, is related to enhanced aerobic capacity, marked by increased maximal oxygen consumption . Furthermore, high-intensity aerobic exercise improves thermoregulation and the metabolic rate, likely driven, in part, by the browning of WAT in experimental models . However, these adaptations are not very clear in humans .
Schematic representation of RAS modulation following ExT
Recently, more attention has been given to high-intensity interval training . This type of training consists of performing short bouts of near maximal effort . The meta-analysis by Maillard et al. (2017)  provided information regarding the effect of high-intensity interval training in normal weight and overweight/obese populations. Their results showed that high-intensity protocols can decrease total fat mass only in patients with excess adiposity. Furthermore, low-intensity ExT seems to be a better approach for reducing abdominal fat mass in lean subjects because higher intensity ExT only reduced abdominal and visceral fat in overweight/obese patients . Aerobic ExT combined with strength training might improve adipose tissue metabolism, reduce abdominal fat and prevent muscle wasting or even induce skeletal muscle hypertrophy in SO patients, as observed in older obese population with age-related loss of muscle mass [168,169].
ExT adaptations are mediated by a network of molecular and metabolic pathways that are activated by muscle contraction. Exercise-induced remodelling could be mainly associated with myokine and adipokine secretion, mitochondrial biogenesis, and protein turnover in a cross-talk between the skeletal muscle and WAT (Figure 4). Interestingly, studies focusing on muscle contraction-induced myokine secretion show contradictory effects [141,170]. For example, the exercise-induced increase in IL-6 levels in skeletal muscle improves adipocyte insulin signalling, stimulating IL-10 production and AMPK activity, which have been related to small-sized adipocytes and browning in obese animals [171,172]. However, also in obesity, muscle and plasma IL-6 and TNF-α levels are reduced after training, probably mediated by the anti-inflammatory effects of ExT [173,174]. Thus, exercise appears to be capable of generating anti-inflammatory effects mediated, at least in part, via increased IL-6 production in skeletal muscle and diminished IL-6 production in WAT .
The secretion of catecholamine stimulates triglyceride breakdown within the adipocytes, promoting the loss of WAT mass, and small adipocytes affect the expression of adipokines . ExT reduces the secretion of leptin, TNF-α, MCP-1, and IL-6, and increases the expression of adiponectin and FGF21 in WAT in obesity [175,177]. A complex cross-talk between the adipokine and myokine possibly mediates the improvement of whole-body insulin sensitivity and glucose tolerance related to ExT, mainly by increasing GLUT4 translocation to the cell membrane [173,175,178].
The increase in mitochondrial biogenesis is one of the most relevant exercise-induced adaptations, leading to metabolic and energy efficiency and closing the adipocyte–myocyte loop. ExT increases thermogenic mediators (UCP1, FGF21 and PRDM16) through PGC-1α and PPARγ activation, and the increase in sympathetic activity up-regulates mitochondrial metabolism, promoting the browning of WAT [177,179]. Interestingly, ExT also induces irisin secretion , and irisin is not only a myokine but also an adipokine with myogenic and browning effects that induces positive self-regulation in both skeletal muscle and WAT . In this regard, exercise-induced PGC-1α leads to the activation of β-oxidative genes in type I muscle fibres that culminate in the increase in maximum oxygen consumption [94,180].
Moreover, mitochondrial biogenesis protects against muscle atrophy and decreases lipid storage, leading to protein turnover, which is another cellular process involved with exercise-induced remodelling . Therefore, muscle wasting might be counteracted by exercise through enhancement of the secretion of IGF-1 . In addition, endurance exercise contributes to satellite cell differentiation through p38 MAPK and Akt signalling, reducing the FoxO3-associated muscle atrophy and stimulating autophagy protection from sarcopenia and increasing lifespan . On the other hand, aerobic exercise can inhibit skeletal muscle protein degradation through changes in atrogin-1 expression in the muscles of high-fat-fed ovariectomized rats .
The cross-talk between adipose tissue and skeletal muscle is of considerable interest, especially from the RAS perspective (Figure 4), which we proposed in this review. Overall, the overactivation of the RAS classical arm is one of the mechanisms driving fat weight gain and muscle wasting.
A plethora of studies has shown that ExT shifts the actions of the RAS towards the protective arm (ACE2/Ang 1-7/MasR) in several disease models [182–187], as shown in Figure 4. Chronic heart failure induces severe skeletal muscle atrophy, and ExT seems to minimize this effect via the RAS . Therefore, the overactivation of the ACE/Ang II/AT1R arm in chronic heart failure rats is normalized by aerobic training, observed by an increased plasma Ang 1-7/Ang II ratio, decreased AT1R expression and increased MasR gene expression in the soleus muscle . Similar results were observed in diet-induced obese rats, where obesity increased the local ACE/AT1R axis and reduced the ACE2/MasR axis in skeletal muscle . Low-volume (5 days/week; 30 min/day) and high-volume (5 days/week; 60 min/day) ExT, at moderate intensity (60% of maximal velocity), were able to ameliorate blood pressure, body composition, the blood lipid profile, inflammatory status and insulin resistance . However, only high-volume ExT shifted the balance of the local RAS towards the protective arm in skeletal muscle, with a decrease in ACE and AT1R protein expression and increase in ACE2 and MasR protein .
In addition to the beneficial effects that ExT exerts on the RAS (Figure 1), the opposite idea that RAS directly influences exercise (Figure 4) has been described by Motta-Santos et al. (2016) , demonstrating that ACE2 knockout mice might have lower physical capacity. During free wheel running training for 6 weeks, knockout mice ran 30% less than wild-type mice. Moreover, the ACE2 knockout mice did not exhibit beneficial physiological adaptation induced by exercise in body composition , likely driven by the lower volume of exercise spontaneously performed by these animals, and not necessarily by a lower physical capacity. However, there are no studies on ExT with alamandine/MrgD, Ang IV/AT4R and other new RAS components in skeletal muscle and adipose tissue.
In humans, the RAS might also modulate the effect of ExT [190,191]. The ACE gene insertion (I allele: 287 bp) polymorphism is related to lower RAS activity and seems to be related to a better response to physical training in skeletal muscle . Furthermore, the ACE genotype probably alters substrate mobilization after ExT, leading to improved metabolic efficiency and better performance . Folland et al. (2000) , although showing no association between ACE genotype and strength in untrained subjects, showed that participants with the ACE deletion (D allele: 287 bp) genotype produced higher strength levels after 9 weeks of isometric training . Hamada et al. (2011)  suggested that AT2R genotype may modulate the effect of ExT on blood pressure because subjects with the AT2R insertion genotype did not show improvements in blood pressure, regardless of weight loss after ExT. The mechanisms leading to exercise-induced changes in the RAS and its consequent physiological adaptations remain to be investigated.
Obesity leads to major changes in body composition, such as excessive accumulation of adipose tissue and skeletal muscle wasting. The coexisting adipose tissue gain and loss of muscle mass recognized as SO is associated with a higher risk of mortality than obesity alone. Recent studies have shown that SO is associated with chronic low-grade inflammation, increased oxidative stress, increased plasma concentration of various hormones (e.g., insulin and leptin), and lipotoxicity. The current management of SO includes lifestyle changes, with ExT as the first-line choice. In this review, we present the RAS as a key mechanism of WAT and skeletal muscle metabolic dysfunction. Furthermore, we discuss the interaction between the RAS and exercise and the possible underlying mechanisms of the health-related aspects of ExT.
The RAS exists as both a circulating hormone system and as a local endo-, para- and autocrine signalling mechanism within various tissues, including the adipose tissue and skeletal muscle. Therefore, the RAS has emerged as a key regulator of metabolic homoeostasis and has become the target of therapeutic intervention for tackling obesity and its comorbidities, not only in animals but also in clinical trials. In adipose tissue, the up-regulation of the classical arm promotes adipocyte hypertrophy through inhibition of lipolysis, activation of lipogenesis, dysregulation of adipokine secretory patterns, decreased glucose uptake, and impaired insulin signalling and induces ROS generation and the expression of NADPH oxidase subunits. Altogether, these mechanisms are linked to insulin resistance, reduced adiponectin release, increased macrophages recruitment and raised inflammatory markers. Therefore, as previously proposed, Ang II acts as a novel ‘adiposity signal’. In skeletal muscle, the RAS classical arm has been implicated in skeletal muscle wasting, reduction in aerobic capacity and atrophy by the UPP overactivation, decreased muscle strength and fibre diameter, down-regulation of the MHC levels, and induction of ROS and insulin resistance.
Recent studies have indicated the importance of the balance between ACE/Ang II/AT1R (classical arm) and ACE2/Ang 1-7/MasR (protective arm) to treat the pathophysiological cascade of SO. Ang 1-7, through MasR, promotes vasodilation, inhibits cell proliferation, inflammation and antihypertensive effects, improves insulin sensitivity, increases glucose uptake and prevents ROS generation. In addition, Ang III, Ang IV, AT4R, Ang A, alamandine, and MrgD have emerged as new components of the RAS that could be targets to treat obesity.
Growing evidence also suggests that ExT can shift the balance of the RAS towards the protective arm, supporting the concept of bidirectional interaction. Additionally, the training protocol, intensity and duration may interfere with the effects of ExT on the RAS components. Several studies have examined the impact of the activation or inhibition of RAS components on energy and cardiovascular homoeostasis.
Important progress has been made in our understanding of the underlying metabolic and molecular mechanisms of the RAS, and potential perspectives have emerged as targets for tackling SO. A recent approach of the modulation of the RAS components by ExT may be even more promising. ExT can stimulate the ACE2/Ang 1-7/MasR arm in parallel with the inhibition of ACE/Ang II/AT1R arm. This change shifts the balance of the RAS towards the protective arm, supporting the concept of bidirectional interaction between ExT and the RAS, which may be revealed as routes for therapeutic intervention for SO. The present study also highlights the complexity and importance of RAS regulation in adipose tissue and skeletal muscle pathophysiology. However, few prospective trials in humans and some animal data have assessed the efficacy of this strategy. Many questions persist regarding the complex mechanisms whereby ExT modulates the RAS in the setting of obesity. Future studies are required to clarify whether ExT directly modulates the RAS and/or whether this modulation is secondary to reduced adiposity, improved metabolic and muscle health, and indirect effects from intermediate molecules in adipose tissue and skeletal muscle. Finally, interventions involving RAS modulation have clear potential as clinical treatments for SO-induced metabolic dysfunction.
The authors declare that there are no competing interests associated with the manuscript.
phosphorylates 4E binding protein 1
protein kinase B
AMP-activated protein kinase
angiotensin receptor blocker
Ang type 1 receptor
Ang type 2 receptor
brown adipose tissue
body mass index
cluster of differentiation 68
extracellular signal-regulated kinase
fatty acid synthase
fibroblast growth factor 21
forkhead box O
glucose transporter 4
glycerol phosphate dehydrogenase
glycogen synthase kinase 3β
insulin-like growth factor 1
Jun N-terminal kinase
muscle atrophy F-box
mitogen-activated protein kinase
monocyte chemoattractant protein 1
myosin heavy chain
Mas-related G-protein-coupled receptor
mammalian target of rapamycin
nuclear factor κ B
PPARγ co-activator 1 α
peroxisome proliferator-activated receptor
PR (PRD1-BF1-RIZ1 homologous)-domain containing 16
reactive oxygen species
transforming growth factor β
tumour necrosis factor α
uncoupling protein 1
white adipose tissue
These authors contributed equally to this work.