Energy Metabolism, Mitochondria, Mitochondrial Function, Bioenergetics, Physical Fitness
Conditions
Keywords
Aerobic Fitness, Metabolic Flexibility, Metabolic Fuel Utilization, Substrate Oxidation, Fasted Exercise, Acute Exercise, Peripheral Blood Mononuclear Cells, Mitochondrial Respiration, Indirect Calorimetry, Maximal Oxygen Uptake, Fat Oxidation, Carbohydrate Oxidation
Brief summary
The aim of this study is to investigate the effects of aerobic fitness level, feeding status, and acute exercise on whole-body metabolic fuel utilization and mitochondrial function in peripheral blood mononuclear cells (PBMCs) in healthy young men. The main questions it aims to answer are: 1. Do individuals with high and low aerobic fitness levels differ in fat and carbohydrate oxidation at rest and during exercise following an overnight fast, and do their PBMC mitochondrial responses to acute exercise differ? 2. Does feeding alter fat and carbohydrate oxidation at rest and during exercise compared with the fasted state in individuals with high aerobic fitness? 3. Are PBMC mitochondrial respiration parameters and their exercise-induced changes associated with fat and carbohydrate oxidation during fasted exercise?
Detailed description
This experimental study uses a mixed design incorporating both between- and within-subject comparisons to investigate the influence of aerobic fitness level, feeding status, and acute exercise on substrate oxidation and peripheral blood mononuclear cell (PBMC) mitochondrial function. Between-group comparisons will examine differences between individuals with high and low aerobic fitness under standardized fasted conditions, whereas within-subject comparisons of fasted and fed conditions will be performed in the high-fitness group. In addition, the repeatability of substrate oxidation measurements will be assessed in the high-fitness group. A total of 30 healthy men aged 18-25 years with a body mass index (BMI) of 18.5-24.9 kg·m-² will be recruited. Participants will be classified into two groups according to maximal oxygen uptake (V̇O₂max). The high-fitness group will comprise 15 endurance-trained individuals with a V̇O₂max \>55 mL·kg-¹·min-¹, whereas the low-fitness group will comprise 15 sedentary or physically inactive individuals with a V̇O₂max \<45 mL·kg-¹·min-¹. During the first laboratory visit, participants in both groups will undergo body composition assessment using dual-energy X-ray absorptiometry (DXA) and complete an incremental cycling test to determine V̇O₂max. Following sufficient recovery, participants will complete a 20-min familiarization cycling session at an intensity corresponding to 65% of V̇O₂max. Participants in the high-fitness group will complete four laboratory visits in total. During the second and third visits, participants will arrive at the laboratory following an overnight fast. Resting metabolic rate (RMR) will be assessed, after which participants will complete 30 min of cycling exercise at 65% of V̇O₂max. These visits will be performed under identical standardized conditions to determine the repeatability of substrate oxidation measurements. During the fourth visit, participants in the high-fitness group will arrive at the laboratory following an approximately 12-h overnight fast. RMR will first be assessed, followed by the collection of a venous blood sample immediately before exercise. Participants will then complete 30 min of cycling at 65% of V̇O₂max, and a second venous blood sample will be collected immediately after exercise. These samples will be used to assess exercise-induced changes in PBMC mitochondrial function under fasted conditions. One hour after completing the fasted exercise trial, participants will consume an ad libitum meal consisting of pasta and meatballs. One hour after completion of the meal, RMR assessment and the 30-min cycling exercise trial at 65% of V̇O₂max will be repeated under fed conditions. No blood samples will be collected during the fed condition. Accordingly, comparisons between fasted and fed conditions will be restricted to outcomes related to whole-body substrate metabolism. Participants in the low-fitness group will complete two laboratory visits. The first visit will include DXA assessment, determination of V̇O₂max, and the familiarization cycling session, as described above. During the second visit, participants will complete the fasted experimental condition. Following an approximately 12-h overnight fast, RMR will be assessed and a venous blood sample will be collected immediately before exercise. Participants will then complete 30 min of cycling at 65% of V̇O₂max, followed immediately by collection of a second venous blood sample. The low-fitness group will not complete the repeatability trials or the fed condition. Therefore, between-group comparisons of substrate oxidation and PBMC mitochondrial function will be performed using data obtained under the standardized fasted condition. Respiratory gas exchange will be continuously measured during resting and exercise assessments to determine oxygen consumption (V̇O₂) and carbon dioxide production (V̇CO₂). These measurements will be used to calculate rates of fat and carbohydrate oxidation. Rating of perceived exertion (RPE) will be recorded every 5 min during exercise using the Borg 6-20 scale. PBMCs will be isolated from venous blood samples collected immediately before and after the fasted exercise test. Mitochondrial oxygen consumption rate and proton efflux rate will be measured using a Seahorse XFe96 extracellular flux analyzer. These measurements will be used to evaluate exercise-induced changes in PBMC mitochondrial function and their relationship with whole-body fat and carbohydrate oxidation during exercise. Participants will also complete a 24-hour dietary intake record before the main experimental visit.
Interventions
BEHAVIORALAcute Moderate-Intensity Cycling Exercise
Participants will perform 30 minutes of continuous cycling exercise on a cycle ergometer at an intensity corresponding to 65% of their individual maximal oxygen uptake (VO2max). Both aerobic fitness groups will complete the exercise following an overnight fast. Participants in the high aerobic fitness group will also repeat the exercise one hour after consuming an ad libitum meal. Respiratory gas exchange will be measured throughout exercise to calculate fat and carbohydrate oxidation, and rating of perceived exertion will be recorded every five minutes using the Borg 6-20 scale.
OTHERAd Libitum Mixed Meal
One hour after completing the fasted exercise test, participants in the high aerobic fitness group will be provided with an ad libitum mixed meal consisting of pasta and meatballs and will be allowed to eat until comfortably full. One hour after the meal, resting metabolic rate measurement and the 30-minute cycling exercise test at 65% of VO2max will be repeated under the fed condition. No blood samples will be collected during the fed condition.
Sponsors
Hacettepe University
Study design
Allocation
NON_RANDOMIZED
Intervention model
PARALLEL
Primary purpose
BASIC_SCIENCE
Masking
NONE
Intervention model description
Participants will be non-randomly classified into two parallel groups according to measured maximal oxygen uptake (VO2max) and habitual training status. The high aerobic fitness group will complete repeated fasted assessments and a main experimental visit including fasted and fed exercise conditions in a fixed order. The low aerobic fitness group will complete the fasted experimental condition only. Both groups will perform 30 minutes of cycling exercise at 65% of their individual VO2max.
Eligibility
Sex/Gender
MALE
Age
18 Years to 25 Years
Healthy volunteers
Yes
Inclusion criteria
* Healthy male volunteers aged 18-25 years * Body mass index between 18.5 and 24.9 kg/m² * Residing on the Hacettepe University Beytepe Campus * Eligibility for one of the following aerobic fitness groups * High aerobic fitness group: maximal oxygen uptake (VO2max) greater than 55 mL/kg/min and regular participation in endurance training. * Low aerobic fitness group: VO2max lower than 45 mL/kg/min and sedentary or not regularly physically active
Exclusion criteria
* Current smoking * Regular or excessive alcohol consumption * Any musculoskeletal condition that may limit or prevent exercise * Allergy to any of the foods provided as part of the study meal * Currently following a weight-loss or other restrictive diet
Design outcomes
Primary
| Measure | Time frame | Description |
|---|---|---|
| Change in PBMC Mitochondrial Oxygen Consumption Rate From Pre- to Post-Exercise | Immediately before and immediately after the fasted 30-minute cycling test at 65% of VO2max. | Mitochondrial oxygen consumption rate (OCR) will be measured in peripheral blood mononuclear cells (PBMCs) using a Seahorse XFe96 extracellular flux analyzer. PBMCs will be isolated from venous blood samples collected immediately before and immediately after the fasted 30-minute submaximal cycling test. Pre- to post-exercise changes in OCR will be evaluated between the high and low aerobic fitness groups and examined in relation to whole-body fat and carbohydrate oxidation during exercise. |
| Change in PBMC Proton Efflux Rate From Pre- to Post-Exercise | Immediately before and immediately after the fasted 30-minute cycling test at 65% of VO2max. | Proton efflux rate (PER) will be measured in peripheral blood mononuclear cells (PBMCs) using a Seahorse XFe96 extracellular flux analyzer. PBMCs will be isolated from venous blood samples collected immediately before and immediately after the fasted 30-minute submaximal cycling test. Pre- to post-exercise changes in PER will be evaluated between the high and low aerobic fitness groups and examined in relation to whole-body substrate oxidation during exercise. |
| Fat Oxidation Rate During Submaximal Exercise | During each 30-minute submaximal cycling test performed at 65% of VO2max. | Whole-body fat oxidation rate, expressed in grams per minute (g/min), will be calculated using the Frayn stoichiometric equation from oxygen consumption (VO2) and carbon dioxide production (VCO2) measured by indirect calorimetry during 30 minutes of cycling at 65% of the participant's individual VO2max. Fat oxidation will be compared between the high and low aerobic fitness groups under fasted conditions and between the fasted and fed conditions within the high aerobic fitness group. |
| Carbohydrate Oxidation Rate During Submaximal Exercise | During each 30-minute submaximal cycling test performed at 65% of VO2max. | Whole-body carbohydrate oxidation rate, expressed in grams per minute (g/min), will be calculated using the Frayn stoichiometric equation from oxygen consumption (VO2) and carbon dioxide production (VCO2) measured by indirect calorimetry during 30 minutes of cycling at 65% of the participant's individual VO2max. Carbohydrate oxidation will be compared between the high and low aerobic fitness groups under fasted conditions and between the fasted and fed conditions within the high aerobic fitness group. |
Secondary
| Measure | Time frame | Description |
|---|---|---|
| Resting Fat Oxidation Rate | During the resting metabolic assessment before the fasted and fed exercise conditions. | Resting fat oxidation rate, expressed in grams per minute (g/min), will be calculated from oxygen consumption (VO2) and carbon dioxide production (VCO2) measured by indirect calorimetry during the resting metabolic assessment. Resting fat oxidation will be compared between the high and low aerobic fitness groups under fasted conditions and between the fasted and fed conditions within the high aerobic fitness group. |
| Resting Carbohydrate Oxidation Rate | During the resting metabolic assessment before the fasted and fed exercise conditions. | Resting carbohydrate oxidation rate, expressed in grams per minute (g/min), will be calculated from oxygen consumption (VO2) and carbon dioxide production (VCO2) measured by indirect calorimetry during the resting metabolic assessment. Resting carbohydrate oxidation will be compared between the high and low aerobic fitness groups under fasted conditions and between the fasted and fed conditions within the high aerobic fitness group. |
| PBMC Glycolytic Proton Efflux Rate (glycoPER) | Immediately before and immediately after the fasted 30-minute cycling test at 65% of VO2max. | Glycolytic proton efflux rate (glycoPER) will be calculated from the total proton efflux rate after correction for the mitochondrial contribution to extracellular acidification. Measurements will be obtained from PBMCs isolated from venous blood samples collected immediately before and immediately after the fasted submaximal exercise test using a Seahorse XFe96 extracellular flux analyzer. |
| Resting Metabolic Rate | Before fasted exercise in both groups and 1 hour after the meal before fed exercise in the high-fitness group. | Resting metabolic rate, expressed in kilocalories per day (kcal/day), will be measured using a Q-NRG indirect calorimetry system in canopy mode. Measurements will be performed after an overnight fast in both aerobic fitness groups. In the high aerobic fitness group, resting metabolic rate will also be reassessed one hour after the ad libitum meal and before the fed exercise test. |
Countries
Turkey (Türkiye)
Contacts
CONTACTBurcu ÖZALP, MSc student
CONTACTSüleyman BULUT, PhD
PRINCIPAL_INVESTIGATORSüleyman BULUT, PhD
Hacettepe University, Faculty of Sport Sciences
PRINCIPAL_INVESTIGATORMuhammed M ATAKAN, PhD
Hacettepe University, Faculty of Sport Sciences
Outcome results
None listed