Gene-Exercise Interactions in Athletes

Exercise

genetic polymorphism, sport performance, myokines

Athlete status is a heritable trait that could be explained with a number of potentially important DNA polymorphisms contributing to predisposition to success in certain types of sport. The first aim of the study is to determine the genetic profile of Slovenian athletes. The associations of 30 common gene polymorphisms with aerobic and anaerobic athlete status will be investigated as a single and polygenic profile. The second aim is to investigate the impact of the genetic variants contributing to different acute response to low vs. high intensity exercise. Physiological and biochemical measurements will be carried out. Variability in physiological adaptation in response to exercise will provides an opportunity to study the relationship between the molecular response to exercise and the extent of physiological changes in athletes. Currently, it is not yet clear whether different genetic variant associated with exercise responses remains uniform, with different exercise intensities, structure and duration of exercise.

Slovenian athletes (n = 300; 18-40 years) of regional or national competitive standard will be recruited from the endurance oriented and power oriented sports disciplines. Healthy unrelated individuals without any competitive sport experience will serve as controls (n= 200; 18-40 years). After completing the questionnaire (for athletes: covering demographics, geographic ancestry, sports classification, discipline, and history, as well as frequency and volume of training; for control group: demographics, geographic ancestry habits of daily living), genotyping analyses will be performed. The athletes and controls will be genotyped for 30 candidate gene polymorphisms considered likely to influence endurance performance. Molecular genetic analysis will be performed with DNA samples obtained from the capillary whole blood. Genotyping for 30 gene polymorphisms will be performed by Real-Time PCR on LightCycler® 96 Instrument (Roche) and KASP (Kompetitive Allele Specific PCR) genotyping technology. In addition, 40 athletes with endurance genetic variant will be selected to participate in the acute exercise study. In order to investigate the acute effects of low and high intensity exercise the concentration of circulating myokines will be measured. Three distinct experimental sessions scheduled 96 h apart, at the same time of the day and in random order will be applied to this sub-group of athletes. In the first session, aerobic power will be determined using an incremental test to exhaustion on a cycling ergometer (Velodyn, Racermate ™, USA). Oxygen consumption (VO2) will be determined breath by breath using a Quark gas analysis system (Quark, Cosmed, Rim, Italy). The interventions during the other two experimental session will consist of cycling exercise with continuous 60 min cycling at 50% PPO (low intensity), and 8 x 5 min at 80% PPO (between intervals 1.5 min at 75 W) exercise (high intensity). The participants will be asked to follow the prescribed diet regime, to avoid the intake of alcohol and to not perform any physical activity in the 24 hours prior to the experimental sessions. The experimental sessions will be performed from 8 to 11 am, in random order and scheduled 96 h apart. Before each intervention, the individuals will remain seated in the laboratory for a period of 20 minutes with a room temperature between 22-24ºC. During this period, resting blood samples will be collected. Two more samples of blood will be collected, immediately post-exercise intervention and 2h post-exercise. Venous blood will be collected into EDTA tubes, centrifuge at 2500-3000g for 20 minutes, and a separate plasma will be stored at -80 ° C for the further analysis. The quantification of biomarkers will be done using the MAGPIX® system, magnetic bead-based multi-analyte panels and MILLIPLEX® Analyst 5.1 software (MAGPIX®, Merck Millipore). For the myokine analysis a commercial kit HMYOMAG-56K will be used. Genotype distribution and allele frequencies between each of the two groups of athletes (endurance and power) and controls will be compared using χ2 tests. Endurance genotype score (EGS) will be construct. First, each genotype will be scored within each polymorphism. Thus, a genotype score (GS) of 2, 1 and 0 to each individual genotype theoretically associated to highest, medium or lowest potential of endurance phenotypes will be assigned. Second, the GS of each single genotype will be summed ∑(i=1)\^nSNPi. Third, the EGS will be transformed to a 0-100 scale for easier interpretation, as follows: EGS=(100/2n)∑\_(i=1)\^nSNPi. An EGS of 100 represents an 'optimal' polygenic profile for endurance athlete-that is, that all GS are 2. In contrast, an EGS of 0 represents the 'worst' possible profile for endurance athlete, that is, all GS are 0. The mean the EGS obtained in the three study groups will be calculated. The EGS of endurance, power athletes and non-athletes (controls) will be compared with one-way analysis of variance (ANOVA), and Tukey post hoc test will be used for between-group comparisons. We will also performed ANOVA to compare the EGS between elite- and national-level athletes within each group of endurance and power athletes. Data normality was verified through an exploratory analysis using the Shapiro-Wilk test. Two-way mixed ANOVA will be applied to check the main interaction effects of time by exercise session (time\*session) and of time (time) on myokine plasma concentration. For statistically significant effects, a post-hoc Tukey test will be adopted for multiple comparisons. All values will be expressed as mean and standard deviation (SD). P values of \<0.05 will be considered statistically significant. Bonferroni's correction for multiple testing will be performed by multiplying the P value with the number of tests where appropriate. Statistical analyses will be carried out using the SPSS program, version 21 (Chicago, IL).

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