**1. Introduction**

The regulation of energy homeostasis and high training stress is dependent on several peripheral factors that communicate the status of body energy stores to the brain [1]. These peripheral factors are also synthesized from adipose, muscle, and bone tissues, which may act as endocrine organs [2]. For example, it has been found that specific adipose-derived factors, including circulating leptin and adiponectin concentrations, can be sensitive to changes in training volume and could be used to characterize physical stress conditions in athletes [1]. In elite female rowers, Kurgan et al. [3] investigated such peripheral markers as tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), insulin-like growth factor-1 (IGF-1), and leptin to assess variations in energy homeostasis and training stress over a training year. It appeared that fluctuations in training load (high vs. low) were accompanied by parallel changes in TNF-α and IL-6, while IGF-1 and leptin remained relatively stable over a training season in this population of young female athletes with suitable energy availability [3]. Similarly, adipokines such as circulating leptin, adiponectin, resistin, and visfatin concentrations have been used to characterize energy homeostasis in highly trained adolescent rhythmic gymnasts (RG), who begin to exercise at an early age and often adopt negative energy balance to retain lean physique [4–6]. Adiponectin was positively associated with weekly training volume in elite young RG participating in World Championships [7], while leptin levels in highly trained adolescent RG can be as low as in anorectic individuals and chronic athletic activity in the presence of prolonged high energy expenditure state decreases leptin concentrations in growing and maturing RG athletes [8].

**Citation:** Jürimäe, J.; Remmel, L.; Tamm, A.-L.; Purge, P.; Maasalu, K.; Tillmann, V. Associations of Circulating Irisin and Fibroblast Growth Factor-21 Levels with Measures of Energy Homeostasis in Highly Trained Adolescent Rhythmic Gymnasts. *J. Clin. Med.* **2022**, *11*, 7450. https://doi.org/10.3390/jcm11247450

Academic Editor: David Rodríguez-Sanz

Received: 15 November 2022 Accepted: 13 December 2022 Published: 15 December 2022

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The importance of tissue crosstalk in energy homeostasis has been highlighted by studies examining the role of different muscle-derived factors in regulating several adipose tissue adaptations to energy metabolism [2,9,10].

Recently, various myokines have been found to mediate training-induced energy and metabolic processes [9,11], besides the most investigated and well-known myokine-IL-6 [1,12]. These myokines include myostatin [13], follistatin [14], irisin [15], and fibroblast growth factor-21 (FGF-21) [16], which have emerged as potential mediators of traininginduced energy metabolism. While myostatin is a negative regulator of muscle mass [2], follistatin is a myostatin-binding peptide that promotes skeletal muscle development and exerts metabolic benefits by improving glucose metabolism [14]. One of the more recently identified myokine, irisin, is primarily secreted by muscle tissue and released into circulation during exercise, resulting in increased energy expenditure and improved glucose metabolism [17]. Irisin levels have been reported to be positively associated with body fat mass (FM) as a surrogate measure of energy availability [18]. However, no differences in serum irisin concentrations were observed between amenorrheic athletes, eumenorrheic athletes and nonathletes aged 14–21 years [19], and between normal weight and overweight young women with a mean age of 18 years [20]. Furthermore, serum irisin concentrations were not related to measures of physical activity and physical fitness in a group of healthy lean women of a wide age range [21]. In addition to irisin, FGF-21 has also emerged as an energy homeostasis hormone that has been implicated in the modulation of energy metabolism in athletes [16,22]. Accordingly, FGF-21 has been proposed as a myokine with metabolic effects on glucose and lipid metabolism that promotes body FM loss [2,23]. It, therefore, appears that irisin and FGF-21 may signal energy status in specific groups of individuals. However, the response of these myokines to chronic exercise training remains to be elucidated in lean adolescent females.

The exact role of circulating irisin and FGF-21 levels in energy homeostasis in female athletes is still not clear. We have previously demonstrated that acute negative energy balance caused by prolonged aerobic exercise elicited the increment in serum irisin and FGF-21 levels and the increase in irisin was related to weekly training volume, while the increase in FGF-21 was associated with exercise energy expenditure in young female rowers with a mean age of 18 years [16]. The present study was undertaken to examine the effect of prolonged athletic activity on serum irisin and FGF-21 concentrations in highly trained adolescent RG athletes. To our best knowledge, whether these myokine levels are related to the measures of energy homeostasis, such as body FM as an index of energy stores, resting energy expenditure (REE), training volume, or other hormones involved in energy homeostasis have not been studied in lean adolescent athletes. We hypothesized that serum irisin and FGF-21 concentrations are higher in highly trained adolescent RG in comparison with nonathletes, and secondly that these circulating myokine levels would be associated with other measures of energy homeostasis in highly trained female athletes with chronically increased energy expenditure state.

#### **2. Materials and Methods**

#### *2.1. Participants and Research Design*

This study included 53 healthy adolescent females with ages ranging from 14 to 18 years. Participants were divided into rhythmic gymnasts (RG; *n* = 33) and untrained controls (UC; *n* = 20). Before entering the study, participants completed medical and training history questionnaires. Athletes were recruited from local training groups and were competing at the international level. Rhythmic gymnasts had trained regularly for the last 10.3 ± 0.9 years with a mean weekly training volume of 17.6 ± 5.3 h/week. The UC group consisted of adolescents, who took part only in compulsory physical education classes and were not involved in any training groups. Information about the age of menarche, changes in the menstrual cycle, past or present diseases, and any kind of medication, vitamin, or mineral supplement, was collected [24]. None of the participants received any medications or had a history of any chronic diseases. No restrictions were placed on dietary intake, and participants consumed their ordinary everyday diet [25]. All UC adolescent females were eumenorrheic, while 22 participants in the RG group were eumenorrheic and 11 were oligomenorrheic or had secondary amenorrhea. Menstruating participants were examined during the follicular phase, where the blood sample was taken between days 7 and 11 from the onset of menstruation [24].

The study design, purpose, and possible risks were explained to the participants and their parents, who gave their written informed consent before entering the study. The study protocol was approved by the Medical Ethics Committee of the University of Tartu, Estonia and was conducted in accordance with the Declaration of Helsinki. Participants underwent an observational cross-sectional examination. Measurements of the current investigation included anthropometry, body composition, energy expenditure, peak oxygen consumption, and blood analyses.
