Effects and individual response of continuous and interval training on adiponectin concentration, cardiometabolic risk factors, and physical fitness in overweight adolescents

This study aimed to evaluate the effect and individual responsiveness after 12 weeks of high-intensity interval training (HIIT) and moderate-intensity of continuous training (MICT) on adiponectin, cardiometabolic risk factors and physical fitness in overweight adolescents. This study was participated by 52 adolescents, both sexes, 11 and 16 years old, separated into HIIT (n = 13), MICT (n = 15), and control group (CG, n = 24). Body mass, height, waist circumference (WC), fat mass (FM), fat-free mass (FFM), blood pressure, high-density lipoprotein (HDL-c), low-density lipoprotein (LDL-c), triglycerides, glucose, insulin, adiponectin, and C-reactive protein (CRP) were evaluated. Body mass index z-score (BMI-z), waist-to-height ratio (WHtR), insulin resistance, and insulin sensitivity were calculated. Resting heart rate (HRrest), peak oxygen consumption (VO2peak), right handgrip strength (HGS-right), left handgrip strength (HGS-left), and abdominal resistance (ABD) was evaluated. HIIT session lasted around 35 min and MICT of 60 min of exercises on stationary bicycle, three times a weekday for 12 weeks. ANOVA, effect size, and prevalence of responders were used for statistical analysis. HIIT reduced BMI-z, WHtR, LDL-c, and CRP, while increased of physical fitness. MICT reduced HDL-c, while increased of physical fitness. CG reduced FM, HDL-c, and CRP, while increased FFM and HRrest. Frequencies of respondents in HIIT were observed for CRP, VO2peak, HGS-right, and HGS-left. Frequencies of respondents in MICT were observed for CRP and HGS-right. Frequencies of no-respondents in CG were observed for WC, WHtR, CRP, HRrest, and ABD. Conclusion: Interventions with exercises were effective to adiposity, metabolic health, and physical fitness improvements. Individual responses were observed in inflammatory process and physical fitness, important changes in overweight adolescent’s therapy. Trial registration number and date of registration: This study was registered with the Brazilian Registry of Clinical Trials (REBEC), the number RBR-6343y7, date of registration May 3, 2017. What is Known: • Effect of regular physical exercise positively affects overweight, comorbidities, and metabolic diseases, recommended mainly for children and adolescents. What is New: • Due to the great inter-individual variability, the same stimulus can provide different responses; adolescents who benefit from the stimulus are considered responsive. • Intervention of HIIT and MICT did not alter the concentrations of adiponectin; however, the adolescents presented responsiveness to the inflammatory process and physical fitness. What is Known: • Effect of regular physical exercise positively affects overweight, comorbidities, and metabolic diseases, recommended mainly for children and adolescents. What is New: • Due to the great inter-individual variability, the same stimulus can provide different responses; adolescents who benefit from the stimulus are considered responsive. • Intervention of HIIT and MICT did not alter the concentrations of adiponectin; however, the adolescents presented responsiveness to the inflammatory process and physical fitness.


Introduction
Insufficient levels of physical activity have been considered a major cause of morbidity and mortality [1], and the effect of regular exercise positively affects overweight, and metabolic diseases, recommended mainly for adolescents [2,3]. Exercises have been indicated as an effective strategy in the treatment and prevention of obesity, metabolic disorders, and increased physical fitness [2][3][4]. In adolescents, regular practice promotes beneficial effects on adiposity, lipid profile, insulin sensitivity, and metabolic markers [3,4].
However, the magnitude of the benefits can vary according to frequency, intensity, duration, volume, and type of exercise [5]; thus, it has been suggested that changes in adiposity come from moderate-intensity continuous training (MICT) [6], and changes in vascular function from high-intensity interval training (HIIT) [7]. In this context, it has been verified the effect of exercise at adiponectin concentration in obese children and adolescents, but they present divergent results according to the type of exercise practiced. García-Hermoso et al. [2] found that interventions increase adiponectin concentration, suggesting that exercise was effective in those who had the greatest reduction in body composition.
Benefits of exercise on the health of children and adolescents have been widely demonstrated, and it is focused only on the mean values. However, due to the great interindividual variability, the same stimulus can provide different responses according to each individual, and those who benefit from the stimulus after interventions are considered responsive, while those who are not responsive show no change or have worsening condition [8]. In the pediatric population, few studies have analyzed individual responsiveness after exercise interventions on health outcomes and cardiometabolic markers [9][10][11][12] and, to our knowledge, no research has demonstrated adiponectin concentration results in obese adolescents.
Therefore, in view of the scarcity and the divergent results, the analyses of different intensities of exercises, as well as the individual variability of the concentration of adiponectin, are aspects to be investigated. Thus, the objectives of the study were to investigate the effect and determine the prevalence of respondents of HIIT and MICT intervention on adiponectin, cardiometabolic risk factors, and physical fitness in overweight adolescents.

Study design and participants
Study of a quasi-experimental longitudinal design was approved by the Ethics Committee of the UniDBSCO University Center (CAAE 62,963,916.0.0000.5223/2017) and registered in the Brazilian Registry of Clinical Trials (RBR-6343y7). Sample size was calculated a priori using the G*Power software, analysis of variance (ANOVA) of repeated measures, two measures, and three groups. Power of 0.95, α of 0.05, and effect size (f) of 0.25 were assigned. Based on these criteria, minimum sample size was 13 adolescents for each group.
Study population consisted of adolescents, both sexes, aged between 11 and 16 years, recruited from schools, residents of Curitiba, and metropolitan region, Paraná State, Southern Brazil. Recruitment was carried out in a non-probabilistic sampling process, for convenience. Parents and/or guardians were informed about the procedures, signed the informed consent form, while the adolescents signed the assent form. Fiftytwo adolescents participated: HIIT (n = 13), MICT (n = 15), and control group (CG, n = 24). Inclusion criteria were (a) classification of BMI-z as overweight (score ≥ 1 and < 2) or obesity (score ≥ 2) [13], (b) participate in all assessments, (c) not present medical contraindication, (d) not participate in regular exercise in the last 6 months, (e) not participate in weight loss program, (f) not use medication that interfere, and (g) frequency of participation above 75%.

Somatic maturation
Somatic maturation was estimated by determining the distance in years from peak height velocity (PHV) by the mathematical model based on height, age, and sex. The prediction of age at peak height velocity (APHV) was determined by subtracting from the chronological age [14].

Anthropometry and body composition
Height was measured using a portable stadiometer (Avanutri ® ), and body mass was measured on a digital platform scale (Welmy ® ). Body mass index z-score (BMI-z) were calculated in the WHO Anthro Plus ® [13]. Waist circumference (WC) measurement was evaluated with a flexible and inextensible tape (Sanny ® ), applied to the skin at the level of the iliac crests. Waist-to-height ratio (WHtR) was calculated on the ratio between WC and height. Body composition was assessed by tetrapolar bioelectrical impedance analysis (Biodynamics ® model 450). We were instructed to follow the recommendations: (a) avoid vigorous efforts the past 12 h, (b) abstain from food and drinks the past 12 h, (c) abstain from alcohol and caffeinated drinks over the past 48 h, (d) not using diuretics over the past 7 days, (e) urinate about 30 min before the exam, and (f) do not use metallic objects during the exam. Fat mass (FM) and fat-free mass (FFM) were calculated using the equations proposed by Houtkooper et al. [15].

Clinical and metabolic variables
All individuals were evaluated by pediatrician for physical examinations and authorization to participate. Systolic blood pressure (SBP) and diastolic (DBP) were measured using mercury sphygmomanometer with the appropriate cuff size. Two measurements were performed at a 1-min interval, with the lowest value being considered. Blood samples were collected in the morning after 12 h of fasting. Colorimetric enzymatic method was used to measure high-density lipoprotein (HDL-c), low-density lipoprotein (LDL-c), triglycerides (TG), and insulin. Chemiluminescence method was used to determine glucose. C-reactive protein (CRP) was determined by the turbidimetry technique. Total adiponectin concentration was determined by the ELISA method. Homeostasis model was used to assess the insulin resistance (HOMA-IR) [16], and quantitative insulin sensitivity check index (QUICKI) [17] was determined.

Cardiorespiratory fitness
Peak oxygen consumption (VO 2peak ) was measured with maximum incremental test, on a treadmill, using a K4b 2 metabolic analyzer (Cosmed ® ). Protocol started at a speed of 4.0 km·h −1 , a progressive increase of 0.6 km·h −1 every minute, and a constant and fixed inclination of 1%. Heart rate monitor (Polar ® ) was used to determine the resting (HR rest ) and maximum heart rate (HR max ). Test was considered maximum when two of the criteria were observed: (a) exhaustion or inability to maintain the required speed; (b) respiratory exchange ratio ≥ 1.0; (c) achieve the HR max predicted. Highest value obtained was determined in L·min −1 .

Muscular fitness
Muscle strength was assessed by the handgrip strength test, with hydraulic dynamometer (Saehan ® ). Subject standing, shoulders in a neutral position, elbow flexed at a right angle at 90°, and forearm in a neutral position instructed the handle with maximum force. Measurements were performed on each right limb (HGS-right) and left limb (HGS-left) with a 1-min recovery. Muscular resistance was measured by the sit-up test; adolescent positioned in the supine position, with knees flexed at 45°, feet flat on the floor, and arms crossed and at chest height. The signal started the trunk flexion movements until touching the elbows on the thighs, returning to the initial position, and performing the maximum repetitions in 1-min [18].

Intervention
Training sessions consisted of exercises on stationary bicycle (Schwinn ® ), three times a week on alternate days during for 12 weeks, performed in groups of ten adolescents, in an indoor cycling room of a gym, conducted by physical education teachers specialized and supervised by researchers. Exercises intensity was determined from the HR max obtained in the cardiorespiratory fitness test, by the equation of reserve heart rate (HR res ). Heart rate was measured by a heart monitor (Polar ® ) to ensure the pre-established target intensity and the caloric expenditure. Session's intensity was stimulated by the rhythm of music, using a specific playlist for each stage, according to pre-established target heart rate. Caloric expenditure was equivalent between the exercises (t = 0.86; p = 0.388).
HIIT session lasted around 35 min (5 min of warm-up, 20-25 min of training, 5 min of resting activities), being 3 sets of 4 repetitions of 30 s of maximum effort, with 60 s of active recovery with self-selected speed, and 4 min of passive rest between sets, with intensity of 80-95% of the HR res for 8 weeks and 90-100% of the HR res for the last 4 weeks. MICT session lasted 60 min (5 min of warm-up, 35-40 min of training, 5 min of resting activities) with intensity progression every 4 weeks, established at 35-55%, 45-65%, and 55-75% of the HR res . CG was instructed to maintain their usual activities during the study period.

Statistical analysis
Data normality was verified by the Shapiro-Wilk test and homogeneity of variance by the Levene's test. Descriptive statistical of means and standard deviations were used. For comparisons between groups, analysis of covariance (ANCOVA) with Bonferroni's post hoc adjusted for sex, age, and APHV was used. Differences of each group at baseline and post were verified by ANOVA of repeated measures with Bonferroni's post hoc, and Friedman's ANOVA for non-parametric variables. Effect size (ES) was calculated to compare magnitude of the effect. Clinical inference (CI) was realized according to the magnitude of the standardized ES, considering trivial (|0.20|), possibly beneficial/harmful (|0.20-0.39|), beneficial/harmful (|0.40-0.79|), and very beneficial/harmful (|> 0.80|) [19].
Frequencies of individual responsiveness were obtained according to the theoretic model considering the magnitude of the individual effect through the division of delta values (final value-initial value) by the grouped standard deviation (SD), calculated by the formula "√((SD 1 2 + SD 2 2 )/2)." Cutoff point used the responsiveness to present an ES ≥ 0.20 or ≤ −0.20 [8]. Difference in the frequency between groups were analyzed using logistic regression. Data analysis was performed with the SPSS v.24.0, and significance level was established at p ≤ 0.05.

Results
Fifty-two adolescents participated in the study, three groups: HIIT (boys = 9); MICT (boys = 4), and CG (boys = 13) (p = 0.069). Descriptive characteristics and comparisons at baseline and after 12 weeks, as well as effect size results and clinical inference for the groups are presented in Table 1 Figure 1 shows the frequency of respondents in the cardiometabolic risk after the 12-week intervention for the groups. Frequencies of respondents were found at reduction of CRP on HIIT and MICT in relation to the GC. The adiponectin no differences were observed in the frequency of responders and non-responders. CG had frequencies of nonresponders in the WC and WHtR. Figure 2 shows the frequency of respondents in the physical fitness after 12 weeks for the groups. HIIT presented frequency of respondents in the increase of VO 2peak , HGS-right, and HGS-left. MICT presented frequency of respondents in the increase of HGSright. CG had frequencies of non-responders in the increase of HR rest and reduction of abdominal.

Discussion
The main findings of the study suggest that (a) no mean changes were found in adiponectin, effect suggests a trivial change in the HIIT and possibly beneficial in the MICT, and no difference was in the frequency of responsiveness; (b) in the adiposity, HIIT showed mean changes and trivial effect; however, CG showed frequency of non-respondents; (c) in the cardiometabolic risk, beneficial changes were observed in the HIIT, and harmful in the CG. CRP there was a frequency of respondents in the HIIT and MICT and nonresponsiveness for the CG, and (d) in physical fitness, HIIT and MICT showed mean changes, beneficial effects, and frequency of responsiveness, while CG of non-responsiveness.
Our results confirm the importance of exercises be considered as non-pharmacological therapy in adolescents with overweight and in the prevention of associated. Benefits of exercise include adaptations in metabolic parameters, reduction of inflammatory adipocytokines, and increase in antiinflammatory, crucial factors considered in the treatment of obesity-related complications [2][3][4]20]. Due to the importance of inter-individual variability, it verified the frequency of responsiveness, considering each intervention and specificity of each adolescent's response to exercises. Evidence has observed the frequency of responsiveness post-exercise on physical and cardiometabolic health [9][10][11][12]21], and to our knowledge, no published research has analyzed the responsiveness of adiponectin post-exercise in overweight adolescents.
Studies have verified the effect of exercises on adiponectin in different age groups and clinical conditions. In adults, metanalysis found that aerobic exercise demonstrated an influence on adiponectin in overweight individuals [22]. Studies in the pediatric population have shown an increase in the adiponectin [6,20], while others did not report changes [4,23,24]. Differences are related to changes in body composition [2], indicating that exercise was more effective in influencing adiponectin in those with the greatest reduction in body fat.
We did not find differences in adiponectin in the groups, and effect of HIIT was trivial, while in the MICT, an effect was found possibly beneficial, even in the MICT that did not promote reduction in FM. Results similar to those found in girls demonstrated that no increase in adiponectin was observed, even with reduction in body fat after combined  [4], and after lifestyle intervention and moderate intensity of continuous [24]. School intervention of moderate to vigorous exercise did not find an increase in the adiponectin, while a reduction in the percentage of trunk fat was observed, but not the percentage of total fat [23].
Evidencing the probable influence of the body fat distribution as a determinant of adiponectin concentration in obese. Conversely, adiponectin is more related to abdominal fat than to other fat deposits, whereas gluteofemoral fat appears to exert a protective effect [25].
Furthermore, exercise groups did not show differences in frequency of respondents and non-responders in relation to adiponectin and adiposity, but chances of responsiveness were observed for adiponectin in HIIT and MICT compared to the CG, as well as a prevalence of non-responders was observed for WC and WHtR in the CG. Our results confirm that the lack of regular exercise can be harmful to overweight adolescents, changes that negatively impact of cardiometabolic health, consequently, on the reduced physical fitness in those who do not benefit from exercise [11,12,21]. Studies have shown that the regular practice of exercises can promote beneficial effects in the adiposity and body fat [4,12,20].
Different from our results in MICT, it was observed reduction on central adiposity after multidisciplinary intervention with higher volume aerobic exercise [12]. Reinforcing the importance of the time of weekly practice to provide favorable effects in the reduction of adiposity, since the accumulated volume was double what was used in our study, which totaled 150-min weeks. Therefore, it seems that the volume of training may be a determining factor in interventions that seek to reduce adiposity. Thus, higher volume exercise can lead to temporary appetite suppression, helping to regulate energy balance and caloric deficit [26], as well as greater reduction in adiposity in obese children for programs long term (> 24 weeks) [20].
We found in the HIIT reduction in CRP a protein associated with low-grade chronic inflammation, probably associated with a reduction in adiposity. Frequency of responsiveness was observed in HIIT and MICT, demonstrating that adolescents responded favorably in reducing the subclinical inflammatory process associated with obesity, even though the mean value of the MICT did not show a notable change. Also, in the CG, a frequency of CRP non-responsiveness was observed. A trivial effect on HIIT was observed in the increase of adiponectin, and possibly beneficial effect on MICT. However, no differences were found in the adiponectin in the mean values and the prevalence of responders for groups. Results suggest that the anti-inflammatory effect of adiponectin may be crucial according to metabolic demand, and possibly, its concentration presents stable levels in adolescents who do not need its protection, since HIIT promoted changes in adiposity and in the inflammatory process.
Therefore, after intervention with exercises, obese adolescents who present reductions in adiposity and/or in the inflammatory process, there is no need to increase the secretion of adiponectin for metabolic protection, demonstrating that exercise can play a decisive role in the relationship with adiposity [11]. Changes that occur in the microenvironment of adipocytes and their anti-inflammatory response can inhibit the expression and secretion of adiponectin, which seems to be mediated by the secretion and expression of pro-inflammatory cytokines [27]. Thus, inflammation is considered an important risk factor in the pathophysiology of obesity, insulin resistance, and hypertension, contributing to the hyperactivation of the sympathetic nervous system and the renin-angiotensin-aldosterone system [28].
Mechanisms by which exercises can influence the adiponectin have not yet been fully elucidated. In addition to reducing FM [2], exercise may lead to upregulation of AdipoR1 and AdipoR2 receptors, which could result in a reduced need for adiponectin, or could alter its proportions of its isoforms, decreasing the medium molecular weight and increasing the high molecular weight, indicating that it may be more important to measure its isoforms [29]. Nielsen et al. [30] contribute the hypothesis that adiponectin is secreted as a compensatory mechanism for low levels of exercise. We demonstrate that adolescents who participated in the exercise showed increase and responsiveness in cardiorespiratory fitness. This increase could influence so that there was no increase in adiponectin, demonstrating that cardiorespiratory fitness can play a moderating role in these relationship.
Recently, it has been shown that adiponectin is also a myokine, produced and released by skeletal muscle [31], stimulating the autophagic flow, and in case of autophagy activation, affect muscle function, playing a key role in muscle fibers and skeletal muscle dysfunction [32]. The effect of adiponectin on skeletal muscles is like the action signaled by insulin, decreasing insulin secretion and hepatic glucose production [33]. Reduction in adiposity and the increase in physical fitness can increase in adiponectin. Our results demonstrate increase in the muscular fitness, as well as frequencies of responders. Thus, we can consider that muscular fitness could also influence so that there was no increase in adiponectin.
We emphasize that in our study, the measurement of adiponectin was performed in its entirety, and not its isoforms, which may influence the results. Evidence has suggested that high molecular weight adiponectin better reflects obesityrelated metabolic abnormalities than total concentration [29,34]. However, studies also show divergences, while Ciccone et al. [34] found that high molecular weight adiponectin was related to cardiovascular risk in overweight children, Nascimento et al. [23] even after 8 months of intervention with exercise in obese children and adolescents, they did not observe differences in the adiponectin and its isoforms.
The behavior observed in the adiponectin can vary in a non-linear way, according to the degree of adiposity, inflammatory process, and physical fitness. We observed that there are higher concentrations of adiponectin when the levels of adiposity and physical fitness are adequate, indicating that the regular practice of exercises can play a decisive role in the relationship with adiposity and inflammatory process. Obese adolescents probably show lower concentrations of adiponectin because they have an altered cardiometabolic profile, and for there to be an increase in its secretion, we found that there is a need to establish threshold points for levels of physical activity and physical fitness.
We highlight as strengths of this study the effects of different intensity of aerobic exercise and the control group with mean values and interindividual variability, which has been little investigated in physical health and cardiometabolic outcomes in the population of children and adolescents. Furthermore, we used measurements of adiposity, body composition, cardiometabolic risk, and physical fitness, with valid, reliable, and viable methods for health monitoring, as well as we used direct measurement for the determination of CRF. This study has some limitations that must be considered, such as the selection of adolescents and the sample size; the generalization of the results must be carried out with caution. Measurement of adiponectin was the total, and we considered a single measurement, and it is possible that it does not accurately reflect its state, as it results from several tissues; we question to what extent skeletal muscle can contribute. Nutritional variables and other variables on the adolescents' lifestyle were not included, which could influence. Genetic factors that may be determinants of inter-individual variability were not verified. We suggest that interventions aimed at reducing and controlling obesity in adolescents should be encouraged to redirect future adults towards a better health profile.

Conclusions
In conclusion, we found that exercise intervention programs of different intensities did not change adiponectin concentrations and adolescents did not show frequency of individual responsiveness. However, in adolescents who did not benefit from the practice of exercises in the intervention, harmful effects, and greater chances of non-responsiveness were observed, highlighting that the lack of regular exercise practice can be harmful, negatively impacting cardiometabolic risk factors and physical fitness. We emphasize that prevention and early detection and search for therapeutic alternatives to control and treat obesity in adolescents should be indicated for the prevention of secondary complications and indicators of morbidity and mortality in adulthood, since the phase is crucial for the establishment of healthy habits in adult life. In this way, intervention with exercises lead to improvements in physical fitness, in the reduction of obesity, and cardiometabolic risk factors in children and adolescents.