Physical Exercise-Induced FGF-21 to Fight Obesity: An Update Review

: Fibroblast growth factor 21 (FGF-21) is a protein that is involved in the regulation of glucose, lipids, and energy metabolism. To act on target tissues, endocrine FGF-21 binds preferably to FGF receptor 1 (FGFR1) in the presence of the coreceptor named β -klotho (KLB). Some of the effects of FGF-21 include increased fatty acid oxidation, glucose uptake, insulin sensitivity


Introduction
Fibroblast growth factor 21 (FGF-21), a protein with a molecular weight of 23 kDa, is produced in the liver, heart, pancreas, white adipose tissue (WAT), brown adipose tissue (BAT), skeletal muscle, and brain [1], and its action is mediated by the interaction of the transmembrane protein β-Klotho (KLB) with the receptors FGFR1, FGFR2, FGFR3, and FGFR4 [2].
Among other endocrine functions, FGF-21 is known as a potent metabolic modulator, since it stimulates glucose uptake and increases fatty acid oxidation by activating coactivator 1 alpha of the peroxisome proliferator-activated receptor gamma (PGC-1α) [3]. Administration of FGF-21 to rats fed a high-fat diet reduces body weight and visceral fat gain, and increases insulin sensitivity [4]. Apparently, its effect is associated with increased energy expenditure [5], increased fatty acid oxidation, and reduced adiposity and body mass [6]. By exerting such metabolic effects, the therapeutic potential of FGF-21 for the treatment of obesity and diabetes has been the subject of human studies [7].
Obesity has become a significant public health issue worldwide, and strategies to combat the high prevalence are necessary because it causes damage to health and is a financial burden on people and society. Physical exercise has been widely used for the prevention and treatment of obesity. This is because it has the potential to increase lipid oxidation, reduce fat mass, increase insulin sensitivity, and lower blood glucose, as described by some research groups [8,9] and by our [10,11]. We also showed that physical exercise exerts a protective effect diminishing lipid deposition in the kidneys of mice with Obesities 2022, 2 373 insulin resistance [12] and by modifying the skeletal muscle proteins expression to a more oxidative phenotype [13] in an animal model of obesity.
It is known that physical exercise increases both serum concentrations and expression of FGF-21 in the liver [14], increases FGF-21 gene expression in skeletal muscle [15] and increases the gene expression of FGFR1 and KLB in WAT [16] in animals. In humans, the effect of physical exercise on FGF-21 is still contradictory and deserves further investigation. Considering the metabolic role of FGF-21 and the chance of physical exercise to induce FGF-21 secretion, this review aims to explore the potential of physical exercise-induced FGF-21 modulation as a strategy for prevention and treatment of obesity. Therefore, we provided a summary of the research supporting the benefits of physical exercise for the prevention and treatment of obesity and describe the FGF-21 as one of the physiological mechanisms underlying these benefits.

FGF-21 Metabolic Effects
FGF is a large protein family, with 22 members in mammals. Based on gene sequence homology and phylogeny, the FGF family can be divided into seven subfamilies, among which the endocrine FGF-19 subfamily consists of three FGFs, namely FGF-19 (FGF-15 in rodents), FGF-21, and FGF-23 [17]. In order to act on target tissues, FGF-21 binds to FGF receptor 1 (FGFR1) in the presence of the transmembrane protein named β-klotho [18,19]. FGF-21 can also act by other receptors, such as FGFR2, FGFR3, and FGFR4, however it prefers to bind to FGFR1 over these other receptors [20]. Kharitonenkov et al. (2005) were the first to describe FGF-21 as a possible metabolic regulator [5]. They observed that transgenic mice expressing human FGF-21 submitted to high-fat diet for 15 weeks were resistant to obesity, and presented lower levels of fasting glucose and leptin. They also showed that FGF-21 indirectly exerts hypoglycemic effects by inhibiting glucagon secretion. Other studies have reported beneficial effects of FGF-21, including reduction in body weight, liver and circulating triglycerides, fasting plasma insulin and glucose, and increase in energy expenditure [3,21]. Interestingly, obesity and type 2 diabetes have also been associated with increased circulating FGF-21 concentration in humans [22] and animals [23]. The paradoxical effect of FGF-21 was first explained by the FGF-21 resistance associated with lower expression of KLB and FGFR1 [24], but this was contradicted in a recent review by [25].
FGF-21 exerts metabolic effects on several tissues ( Figure 1). In the liver, FGF21 stimulates fatty acid oxidation and reduces lipid flux by increasing peripheral lipoprotein catabolism and reducing adipocyte lipolysis [26]. In the white adipose tissue (WAT), it suppresses lipolysis and enhances insulin sensitivity and adiponectin secretion [26,27], thus ameliorating some harmful effects of obesity, such as hyperglycemia, glucose intolerance, insulin resistance, and dyslipidemia [28,29]. In the brown adipose tissue (BAT), FGF-21 increases glucose uptake, lipolysis, and thermogenesis [30]. Other tissue actions of the FGF-21 can be found in the review published by She et al. (2022) [17].
Since FGF-21 improves hyperglycemia, dyslipidemia, and obesity, the cardioprotective effects of FGF-21 have also been elucidated. In a recent review, it was discussed that FGF-21 prevent endothelial dysfunction and lipid accumulation, inhibiting cardiomyocyte apoptosis and regulating oxidative stress, inflammation, and autophagy [31]. Thus, FGF-21 could be a potential target to prevent and to treat cardiovascular disease.
FGF-21 plays an important role on the metabolism by activating different pathways, usually involving cell differentiation, proliferation, and energy metabolism proteins. One of these proteins is the AMP-activated protein kinase (AMPK), which is recognized as an energy metabolic sensor in cells. The FGF-21 binding to a serine threonine kinase protein called LKB1, which is considered to be a major regulator of AMPK activation [32]. The activation of AMPK pathway leads to rise of PGC1-α, a special regulator of mitochondrial biogenesis, which increases mitochondrial respiratory function and oxidative capacity in adipocytes and consequently higher energy expenditure [33]. Together, FGF-21 and AMPK improve free fatty acid oxidation and energy expenditure [34,35], which can inhibit the accumulation of triglyceride in hepatocytes, and possibly prevent the development of non-alcoholic fatty liver disease [36,37]. Since FGF-21 improves hyperglycemia, dyslipidemia, and obesity, the cardioprotective effects of FGF-21 have also been elucidated. In a recent review, it was discussed that FGF-21 prevent endothelial dysfunction and lipid accumulation, inhibiting cardiomyocyte apoptosis and regulating oxidative stress, inflammation, and autophagy [31]. Thus, FGF-21 could be a potential target to prevent and to treat cardiovascular disease.
FGF-21 plays an important role on the metabolism by activating different pathways, usually involving cell differentiation, proliferation, and energy metabolism proteins. One of these proteins is the AMP-activated protein kinase (AMPK), which is recognized as an energy metabolic sensor in cells. The FGF-21 binding to a serine threonine kinase protein called LKB1, which is considered to be a major regulator of AMPK activation [32]. The activation of AMPK pathway leads to rise of PGC1-α, a special regulator of mitochondrial biogenesis, which increases mitochondrial respiratory function and oxidative capacity in adipocytes and consequently higher energy expenditure [33]. Together, FGF-21 and AMPK improve free fatty acid oxidation and energy expenditure [34,35], which can inhibit the accumulation of triglyceride in hepatocytes, and possibly prevent the development of non-alcoholic fatty liver disease [36,37].
FGF-21 also can act through mitogen-activated protein kinases (MAPK) pathway. When the complex FGFR1 and B-klotho is auto phosphorylated, the cascade signaling for MAPK happens, and leads to the activation of ERK 1/2. Minard et al. (2016) submitted adipocytes cells to stable isotopes to see all the phosphorylation of FGF-21 in these cells [38]. About 15,687 phosphorylation sites occur in 4583 different proteins. FGF-21 signaling was initiated by tyrosine phosphorylation of FGFR1, followed by ERK1 and 2 phosphorylation. In the same study, the authors showed that when the cells were submitted to insulin, FGF-21 could stimulate the MAPK pathway 11-fold more and leads to the activation of mTORC1 resulting in glucose uptake, adiponectin secretion and UCP1 elevation.
Considering that FGF-21 significantly ameliorates metabolic diseases impacts, it has been investigated in pharmacological studies. In this context, six clinical trials in obese subjects have been conducted to test the therapeutic effects of FGF-21, specifically four drugs and one agonist receptor antibody [39]. As reviewed by the authors, the short halflife, the susceptibility to proteolytic inactivation and the obesity-mediated FGF21  [38]. About 15,687 phosphorylation sites occur in 4583 different proteins. FGF-21 signaling was initiated by tyrosine phosphorylation of FGFR1, followed by ERK1 and 2 phosphorylation. In the same study, the authors showed that when the cells were submitted to insulin, FGF-21 could stimulate the MAPK pathway 11-fold more and leads to the activation of mTORC1 resulting in glucose uptake, adiponectin secretion and UCP1 elevation.
Considering that FGF-21 significantly ameliorates metabolic diseases impacts, it has been investigated in pharmacological studies. In this context, six clinical trials in obese subjects have been conducted to test the therapeutic effects of FGF-21, specifically four drugs and one agonist receptor antibody [39]. As reviewed by the authors, the short halflife, the susceptibility to proteolytic inactivation and the obesity-mediated FGF21 resistance are the great challenges for therapies based on FGF-21. Therefore, regular physical exercise may be an alternative to induce the effects of FGF-21 that can contribute to combatting obesity and its damages. In addition, it is a strategy with lower cost and greater access for the population, which can promote more significant effects for individuals and for health systems.

Effect of Physical Exercise on FGF-21
Physical exercise acts in different ways in the body, as it can alter protein secretion, influencing circulating but also locally levels. A continuous aerobic exercise session increases the circulating FGF-21 in both lean and obese individuals immediately after exercise for up to one hour [40]. Interestingly, a high-intensity interval training (HIIT) session appears to prolong the increased levels of FGF-21 induced by the exercise up to 3 h after the end of the session in lean men, while this effect can last for up to 48 h when the resistance exercise with seven types of exercises targeting all the main muscle groups was performed [41]. In obese people submitted to acute moderate-intensity exercise (treadmill exercise, ∼60% peak oxygen uptake), circulating concentrations of FGF21 were elevated after exercise for up to 6 h [42]. Based on research, it seems that resistance exercise session is the type of exercise that keeps the FGF-21 level higher for longer post-acute exercise.
It is important to point out that the acute exercise effect on FGF-21 is not a consensus in the literature. Cuevas  [45] that showed a higher serum concentration of FGF-21 after performing exercise at 90% of maximum heart rate (HR max) in comparison to an exercise performed at 69% of HR max. Furthermore, the correlation between exercise volume and the levels of FGF-21 remains unclear and it is suggested that endogenous FGF-21 is associated with the magnitude of caloric expenditure and oxidative stress caused by acute strenuous physical exercise [46]. This gap remains, and further studies are needed to confirm this hypothesis.
influencing circulating but also locally levels. A continuous aerobic exercise session increases the circulating FGF-21 in both lean and obese individuals immediately after exercise for up to one hour [40]. Interestingly, a high-intensity interval training (HIIT) session appears to prolong the increased levels of FGF-21 induced by the exercise up to 3 h after the end of the session in lean men, while this effect can last for up to 48 h when the resistance exercise with seven types of exercises targeting all the main muscle groups was performed [41]. In obese people submitted to acute moderate-intensity exercise (treadmill exercise, ∼60% peak oxygen uptake), circulating concentrations of FGF21 were elevated after exercise for up to 6 h [42]. Based on research, it seems that resistance exercise session is the type of exercise that keeps the FGF-21 level higher for longer post-acute exercise.
It is important to point out that the acute exercise effect on FGF-21 is not a consensus in the literature. Cuevas-Ramos et al. (2012) [43] and Parmar et al. (2018) [44] observed no significant changes in serum FGF21 of lean individuals after a single session of aerobic exercise and resistance exercise, respectively. A summary of the effects of physical exercise on circulating FGF-21 in lean and obese individuals is demonstrated in the Figure 2. It appears that the intensity of the aerobic exercise affects the FGF-21 secretion response. In fact, He et al. (2019) [45] that showed a higher serum concentration of FGF-21 after performing exercise at 90% of maximum heart rate (HR max) in comparison to an exercise performed at 69% of HR max. Furthermore, the correlation between exercise volume and the levels of FGF-21 remains unclear and it is suggested that endogenous FGF-21 is associated with the magnitude of caloric expenditure and oxidative stress caused by acute strenuous physical exercise [46]. This gap remains, and further studies are needed to confirm this hypothesis. The chronic physical exercise effect on FGF-21 has been extensively investigated. In lean individuals, a significant increase in FGF21 concentration was shown after two weeks of treadmill training [43]. On the other hand, Mendez-Gutierrez et al. (2022) [47] did not find difference in FGF-21 plasma concentration after a 24-week exercise program (continuous aerobic exercise + resistance). As discussed previously, differences in exercise protocols, age, and sex of participants can explain the results [48]. The chronic physical exercise effect on FGF-21 has been extensively investigated. In lean individuals, a significant increase in FGF21 concentration was shown after two weeks of treadmill training [43]. On the other hand, Mendez-Gutierrez et al. (2022) [47] did not find difference in FGF-21 plasma concentration after a 24-week exercise program (continuous aerobic exercise + resistance). As discussed previously, differences in exercise protocols, age, and sex of participants can explain the results [48].
In obese people, both HIIT and resistance training increase FGF-21 levels [49] and the same can be observed in diabetic individuals who performed resistance training and continuous aerobic training [50]. On the other hand, some studies show a reduction in FGF-21 values after resistance training [51] and concurrent training [52], or no changes after continuous aerobic training in cycle ergometer [53] and HIIT [54]. A possible explanation for this phenomenon is the adaptation to subsequent sessions of physical exercise, lowering the production of FGF-21 by the liver.
The liver secretes most of FGF-21 in response to physical exercise. However, FGF-21 is also expressed in heart, adipose tissue, skeletal muscle, and pancreas. Physical exercise has the potential to induce greater tissue expression of FGF-21, as showed by studies with animals. For example, aerobic and resistance training promote greater expression of FGF-21 in the heart [55], while HIIT is able to increase the expression of FGF-21 in skeletal muscle [56]. In another study, continuous aerobic exercise was more effective in increasing FGF-21 in BAT, skeletal muscle, and liver than HIIT. Additionally in this study, there was a drop in the serum amount of trained animals, suggesting that chronic exercise reduces, in the long term, the synthesis of FGF-21 by the liver [57].
Considering that obesity and body adiposity have become a public health problem, FGF-21 has gained notoriety in last years. This attention came after a study that showed that FGF-21 administration promoted weight loss and reduced adiposity in animals fed a high caloric diet and that the effect was dose dependent. This effect was correlated to the higher oxygen consumption, resulting in increased caloric expenditure and proportionally oxidizing more fat at rest [6]. Thus, discovering whether the beneficial effects of physical exercise would be mediated by the action of FGF-21 could give support to understand which intensity and frequency are most appropriate to combat obesity and its damages.
In this context, exercise training reduces the body weight and adiposity of obese animals and restores the concentrations of FGF-21 and β-Klotho in WAT and BAT, ameliorating insulin sensitivity and glucose uptake impaired by obesity [58]. Considering that there are human studies showing that the reduction in fat mass through physical exercise is not necessarily accompanied by changes in FGF-21 levels, perhaps the FGF-21 anti-obesity effects are associated with better crosstalk between liver and BAT. In fact, part of the increase in caloric expenditure induced by FGF-21 has shown to be mediated through BAT and its thermogenic potential and adaptation to exercise training. Evidence shows that adipose tissue sensitivity to FGF-21 appears to be inversely correlated with lower BMI when indirect assessed by measuring the β-Klotho gene expression [59]. In addition, WAT is an endocrine organ, with great capacity to secrete adipokines and other proteins including FGF-21. Taken together, this evidence suggest that next studies should not focus on FGF-21 secreted by the liver through stimulation of exercise training, but by BAT and WAT, increasing their potential for energy expenditure [60].
As physical exercise is the focus of this topic, it is important to highlight its potential to alter the characteristics of adipose tissue, increasing adipokine secretion, heat production, and caloric expenditure. It is known that obese and elderly individuals have less activation of BAT [61], which makes BAT a target of studies in possible treatments. Studies have also already shown that physical exercise in humans has the potential to modify characteristics of BAT, increasing the expression of UCP1 and fatty acid transporters in people with obesity, but not overweight [62,63]. Other studies have already shown that the crosstalk of WAT to BAT is associated with greater tissue expression of FGF-21 [64]. However, more studies are needed to investigate the effect of physical exercise on the synthesis of FGF-21 in adipose tissue, and its potential to alter its characteristics enabling greater heat production and energy expenditure.

Conclusions
In this review, we showed that physical exercise has the potential to modulate circulating FGF21 levels in humans and animals, while tissue effects were primarily observed in animals. By acting in the reduction of blood glucose and lipid concentrations, which improves insulin sensitivity, FGF-21 deserves to be the target of studies that seek to discover ways to prevent and treat obesity. New studies are still needed involving the responses of FGF-21, FGF21 receptors, and co-receptor KLB to different types of physical exercise, which could be useful to support the prescription of physical exercise for the management of obesity.

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