NCT Number: NCT07481578
The goal of this study is to learn how different amounts of supervised indoor cycling training change gut health (gut bacteria, the substances gut bacteria make, and the gut barrier integrity), and how these changes relate to changes in fitness, muscle health, and signs of doing too much training (a state called 'overreaching'). The study includes healthy, recreationally active men aged 18 to 45 years.
The primary questions, for which the study is powered (sufficient participants included), are:
1. Does moderate load training change blood and faecal levels of butyrate (a short-chain fatty acid made by gut bacteria) after eight weeks compared with a control group? 2. Compared to moderate load training, do higher training loads lead to different responses in blood and faecal levels of butyrate?
Researchers will compare:
* A control group that does not complete structured training; * A moderate-load training group that completes eight weeks of supervised cycling (4x/week); * A high-load training group that completes four weeks of moderate-load training followed by four weeks of higher training load (twice the number of training sessions).
Participants will:
* Be randomly assigned to one of the three groups; * Complete 8 weeks of supervised indoor cycling sessions if assigned to a training group; * Complete four study assessment periods (baseline, after week four, after week eight, and after a short taper (rest period); * Provide blood, stool, skeletal muscle, urine, saliva, and breath samples during the assessment periods; * Complete fitness and performance tests and questionnaires during the assessment periods.
Interested in participating?
Request Info18 year–45 year
Male
Interventional
Not applicable
Leuven, Vlaams Brabant, 3001, Belgium
Location status: Recruiting
Sofie Engelborghs
CONTACT
Stefan De Smet
CONTACT
Stefan De Smet
PRINCIPAL_INVESTIGATOR
ICARUS is a monocentric, randomized, controlled, three-arm, parallel-group trial designed to determine whether exercise training load (moderate versus high) modulates the human gut ecosystem, with a primary focus on gut microbial short-chain fatty acid (SCFA) production, and to test whether interindividual gut responses relate to physiological adaptations and potential maladaptations (i.e., overreaching).
STUDY DESIGN: 45 healthy, recreationally active men will be randomized in a 1:1:1 allocation ratio to: (i) a non-training control group, (ii) a moderate-load cycling training group, or (iii) a high-load cycling training group. The intervention lasts 8 weeks and is followed by a 10-day taper period. Outcomes are assessed at four time points: baseline (T1), mid-intervention after 4 weeks (T2), post-intervention after 8 weeks (T3), and post-taper 10 days later (T4).
RANDOMIZATION: Randomization will be stratified by baseline habitual training volume (>3.5 hours versus ≤3.5 hours of exercise per week) using permuted blocks of varying size and a computer-generated allocation sequence. Allocation will be implemented using a secure REDCap web application to support allocation concealment from personnel involved in data collection and analysis.
SETTING AND RECRUITMENT: The trial is conducted at the Exercise and Muscle Physiology Research Group, Department of Movement Sciences, KU Leuven (Leuven, Belgium). Recruitment uses multiple channels, including email invitations, classroom announcements, posters/flyers, and social media.
FAMILIARIZATION AND STANDARDIZATION PROCEDURES: Approximately one week before the intervention starts, participants attend one familiarization session to minimize learning effects and standardize procedures across subsequent assessments. Familiarization includes key performance and physiological testing procedures and instruction on study devices and diaries. A personalized, standardized dietary plan is developed for the three days preceding each assessment period and the assessment days to improve within-participant dietary consistency across time points and to align with specific test requirements (e.g., fiber intake, carbohydrate loading, lactose/mannitol restriction where applicable).
INTERVENTION ARMS - CONTROL GROUP: Participants do not perform a structured training program. To control for expectancy effects, they receive a supervised weekly placebo capsule (1 g granulated sugar) presented as a 'novel plant extract' purported to support gut health and performance. Participants are asked to maintain their habitual exercise patterns and to monitor relevant behaviors as instructed.
INTERVENTION ARMS - MODERATE-LOAD TRAINING GROUP: Participants complete 8 weeks of supervised indoor cycling followed by a 10-day taper. Training is performed on an electromagnetically braked cycle trainer, supervised at the study center. The weekly structure comprises four sessions per week with a fixed sequence of session types (moderate-intensity continuous training, high-intensity interval training, moderate-intensity continuous training, and sprint interval training), and progressive load increases according to a standardized plan.
INTERVENTION ARMS - HIGH-LOAD TRAINING GROUP: Participants complete 4 weeks identical to moderate-load training group, followed by 4 weeks of high-load training in which training volume is doubled relative to MOD by increasing session frequency while maintaining the same intensity distribution.
EXERCISE INTENSITY PRESCRIPTION: Training intensity is individualized using physiological thresholds derived from an incremental cardiopulmonary exercise test with lactate measurements, including power at the first lactate threshold, second lactate threshold, and maximal power output. Training is prescribed based on cycling power output.
ASSESSMENT PERIODS: Each of the four assessment periods spans three consecutive days of home-based and study center-based procedures under standardized conditions and at the same time of day for each participant to limit circadian variability. Assessments integrate biospecimen collection and multi-system phenotyping, including gut-related measures (e.g., SCFAs, gut barrier integrity testing), exercise performance tests, skeletal muscle phenotyping, cardiometabolic testing, autonomic measures, and self-report instruments relevant to recovery and psychological well-being. Stool samples are collected at home and stored frozen prior to transfer to the study center for biobanked storage.
BIOSPECIMEN HANDLING AND STORAGE: Biological samples including blood, stool, skeletal muscle, urine, and saliva are labeled using a study-specific identifier and stored at -20°C of -80°C until analysis and/or future use in accordance with institutional biobank requirements and applicable regulations.
BLINDING AND BIAS MITIGATION: Because of the behavioral nature of the intervention, participants cannot be blinded to group assignment. To reduce expectancy effects, the control group receives placebo supplementation presented as performance-relevant. Where feasible, outcome assessors and laboratory personnel remain blinded to group allocation during data collection and analyses, although some partial unblinding may occur due to staffing constraints.
SAMPLE SIZE AND STATISTICAL PRINCIPLES: The planned sample size is 45 participants (15 per arm), accounting for attrition, and is based on power considerations derived from prior evidence on fecal butyrate responsiveness to endurance training, given the absence of robust human data for the primary circulating SCFA endpoint at the time of planning. Analyses will follow intention-to-treat principles. Group differences over time will be estimated using constrained longitudinal data analysis models that jointly model baseline and follow-up measures while constraining baseline means to be equal across groups, thereby providing ANCOVA-equivalent estimands and accommodating missing data under a missing-at-random assumption via maximum likelihood. A hierarchical testing strategy is planned for the primary and key secondary endpoints to control the type I error rate. Sensitivity analyses may include per-protocol analyses defined prior to database lock.
Healthy volunteers accepted: Yes
Only the study team can determine whether someone qualifies for participation.
Inclusion criteria
Exclusion criteria
The nature of the intervention does not allow blinding of study participants. However, to induce an 'expectancy-effect' in the control group, participants allocated to the control group will receive dietary placebo supplementation. Participants will be told (deceived) that this novel 'plant extract' is believed to have the potential to mimic training effects on gut health and exercise performance. The dietary placebo capsule will consist of 1 g sugar. Intake of the dietary placebo capsule will take place after a fasting state of six hours or more, at a fixed timepoint once a week, and under direct supervision of a research investigator (Sofie Engelborghs). The control group does not participate in any study-related training sessions, but will be asked to continue and monitor (Polar Pacer heart rate monitor) their normal physical activities and sports.
Other names: CON
The moderate load training group will enroll in an exercise intervention of 8 weeks. This intervention consists of cycling sessions on an electromagnetically braked ergometer under direct supervision of the investigators at the Exercise and Muscle Physiology Group. Training intensities are prescribed based on the power output associated with the first and second lactate threshold and maximal power output obtained during a maximal incremental step test with capillary lactate measurements. Three types of training sessions will be prescribed: moderate-intensity continuous training (2x/week), high-intensity interval training (1x/week), and sprint interval training (1x/week). The training load will gradually increase over time, by increasing the session duration and/or intervals.
Other names: MOD
Participants allocated to the high load training group will conduct a similar intervention as the moderate load training group, but with double the amount of training sessions (8x/week) throughout the last four weeks of the 8-week training program. The four weeks of high load training aims to induce a state of overreaching in some though not all participants allocated to the high load training group.
Other names: HIGH
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Diet and in particular fiber intake will be standardized for three days prior to and during the assessment days during which venous blood sample collection will take place. To promote microbial SCFAs production, participants will consume 15 g of oligofructose five hours prior to venous blood sample collection. Serum butyrate concentration will be assessed using gas chromatography-mass spectrometry.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Diet and in particular fiber intake will be standardized for three days prior to and during the assessment days during which fecal collection will take place. Participants will be asked to collect a stool sample at the first experiment assessment day. Fecal butyrate concentration will be assessed using gas chromatography with flame ionization detection.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Short-chain fatty acid concentration (i.e. acetate, propionate, and butyrate concentrations) will be assessed in serum (gas chromatography-mass spectrometry) and feces (gas chromatography with flame ionization detection).
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
The gut microbiome will be evaluated using taxonomic composition and functional genetic capacity (shotgun metagenomics, feces).
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Indices of gut barrier integrity will be assessed by lactulose mannitol ratio test (HPLC-ELSD) and plasma lipopolysaccharide binding protein (ELISA).
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
A gastrointestinal questionnaire will be used to report stool consistency, bowel movement frequency, and gastrointestinal distress.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Aerobic exercise performance will be measured on a bicycle ergometer through a 30-min simulated time-trial test.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Anaerobic exercise performance will be measured on a bicycle ergometer through a 6-s isokinetic sprint test, and a 30-s isokinetic sprint test.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Cardiorespiratory fitness will be evaluated through an incremental maximal cardiopulmonary exercise test (step test), throughout which peak oxygen uptake and the first and second lactate thresholds will be determined
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Knee extensor strength will be assessed using a custom-build dynamometer.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Muscle fiber type composition will be visualized and quantified using fluorescent immunohistochemical staining.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Markers of mitochondrial content, biogenesis, and remodelling will be quantified using a hierarchical analytical workflow combining western blotting, enzyme histochemistry, and spectrophotometric assays.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
If the oral glucose tolerance test indicates between-group differences in glucose tolerance or insulin sensitivity, skeletal muscle samples will undergo western blot analysis to quantify key insulin-signalling proteins.
Time frame: Each training session, once every weekend, and at day 2 and day 3 of each experimental assessment period.
Heart rate will be measured using a Polar Pacer monitor connected to a Polar H10 chest strap. Resting heart rate will be assessed once weekly, while submaximal and maximal heart rate will be continuously recorded during all training and testing sessions.
Time frame: Each training session, once every weekend, and at day 2 and day 3 of each experimental assessment period.
Heart rate variability (HRV) will be measured using a Polar Pacer monitor connected to a Polar H10 chest strap. RR intervals for HRV will be collected once weekly and during each experimental assessment timepoint, with measurements performed immediately upon waking in a supine position. HRV will be quantified using Kubios software, including time-domain, frequency-domain, and non-linear indices.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Changes in serum hsCRP will be evaluated as marker of systemic low-grade inflammation. Gut-local inflammation will be assessed via fecal calprotectin.
In case differential inflammatory or maladaptive responses are observed between study arms, a predefined subset of stored biosamples will be subjected to more detailed inflammatory and gut-barrier profiling within available resources. If moderate vs high load training induces distinct changes in intestinal or systemic inflammatory status, gene and protein expression levels of muscle pro- and anti-inflammatory cytokines will be analyzed using commercially available ELISAs
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Concentrations of resting and exercise-induced hormones related to the hypothalamic-pituitary-adrenal (HPA) axis, stress, energy regulation, and cortisol will be analyzed in venous blood and saliva samples using electrochemiluminescence immunoassay (ECLIA) or enzyme-linked immunosorbent assay (ELISA).
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Participants will complete the validated Recovery-Stress Questionnaire for Athletes (RESTQ-Sport) to assess perceived recovery-stress balance. The RESTQ-Sport comprises 77 items across 19 scales. Each item is rated on a 7-point Likert scale (0 = "never" to 6 = "always"). The recovery-stress balance will be calculated by subtracting the total stress score (sum of 10 stress subscales) from the total recovery score (sum of 9 recovery subscales), with higher values indicating more favourable recovery states.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Mood disturbance will be assessed using the validated Profile of Mood States (POMS) questionnaire (McNair et al., 1971). The POMS is a 65-item instrument that evaluates transient, fluctuating mood states. It comprises six subscales: five negative affect dimensions (Fatigue, Depression, Tension, Anger, and Confusion) and one positive affect dimension (Vigour). Participants rate how they have felt during the past week on a 5-point Likert scale (0 = "not at all" to 4 = "extremely"). The total mood disturbance (TMD) score will be calculated by summing the five negative subscales and subtracting the positive (Vigour) subscale, with higher TMD values indicating greater overall mood disturbance.
An energy index will be calculated as the difference between the POMS Vigour and Fatigue subscale scores, providing an indicator of perceived energy and readiness to perform.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Subjective workload will be assessed using the validated NASA Task Load Index (NASA-TLX) questionnaire, a reliable indicator of both physical and mental workload (Sandra & Lowell, 1988). The NASA-TLX comprises six dimensions: Mental Demand, Physical Demand, Temporal Demand, Frustration, Effort, and Performance. Participants will rate each dimension on a 20-point scale ranging from "very low" to "very high." The overall workload score will be calculated as the mean of the six dimension ratings, with higher scores indicating greater perceived workload.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Subjective well-being will be assessed using the Wellness Questionnaire (Bellinger, 2020), which comprises seven items evaluating physical and mental well-being. Participants will rate each item on a 1-10 Likert scale, where higher scores indicate better perceived well-being. The questionnaire will be completed at the end of each intervention week and at each experimental assessment period. An overall well-being score will be calculated as the sum of all item scores.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
The Wisconsin Upper Respiratory Symptom Survey (WURSS-21) will be used to monitor the incidence and severity of upper respiratory tract infections (Barrett et al., 2009). Participants will rate one global question, ten symptom-related questions, and nine functional-impairment questions (e.g., ability to think, sleep, breathe, walk) on a 0-7 Likert scale (0 = no symptoms, 1 = very mild, 3 = mild, 5 = moderate, 7 = severe), based on average symptom intensity over the previous seven days. The total WURSS-21 score will be calculated as the sum of all items, with a score ≥ 21 indicating the incidence of an upper respiratory tract infection (URTI).
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Peripheral microvascular function will be assessed using a vascular occlusion test combined with near-infrared spectroscopy on the gastrocnemius muscle. The protocol will consist of a 2-min baseline recording, 8 min of arterial occlusion, and 5 min of reperfusion. Muscle oxygen extraction will be quantified from the StO₂ decline during occlusion, and microvascular reperfusion capacity will be determined from the rate of StO₂ recovery after cuff release.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
A venous blood sample will be collected after an overnight fast to analyze fasting glucose and insulin concentrations. Fasting glucose levels and the homeostasis model assessment of insulin resistance (HOMA-IR) will serve as indicators of liver insulin resistance.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
The Matsuda index, derived from an oral glucose tolerance test, will be used as a marker for muscle and whole-body insulin resistance. In this test, 75 g of glucose will be administered following a 10-12 hour overnight fast, and venous glucose and insulin levels will be measured at 0, 30, 60, 90, and 120 minutes post-administration.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
Body composition will be measured by a Dual Energy X-ray Absorptiometry (DXA) scan.
Time frame: During three days each week during the study and three days before and during the experimental assessment periods.
Diet and nutritional intake will be monitored using a food diary (MijnEetmeter) during three days each week. Additionally, participants will follow a standardized dietary protocol for three days before and during the experimental assessment periods.
Time frame: At baseline (T1), mid-intervention after four intervention weeks (T2), post-intervention after eight intervention weeks (T3), and 10 days following intervention completion (T4).
If DXA reveals between-group changes in whole-body or regional fat-free mass, western blotting will be used to assess regulators of muscle protein synthesis and breakdown.
Time frame: From baseline (T1) until 10 days following intervention completion (T4).
Adherence and fidelity to the training intervention will be assessed by recording the number of completed sessions, and the sessions' power output, heart rate, and session duration. Adherence to the dietary supplementation in the control group will be ensured via directly observed administration.
Continuation of participants' habitual sport activities outside the intervention will be monitored using the Polar Pacer smartwatch.
Time frame: Post-intervention (T3, week 8)
Participants in the high-load training group will be classified at T3 as overreached when both of the following are present relative to T2: (i) a reduction in exercise performance, defined as a lower mean power output during either the 30-second sprint test or the 30-minute time trial, and (ii) a reduction in the energy index calculated from the POMS questionnaire as Vigour minus Fatigue. To account for day-to-day variability, the reduction in exercise performance must exceed the coefficient of variation derived from the control group, and the reduction in energy index must exceed the standard error of measurement of the change score derived from the control group. Participants showing a reduction in energy index without a decrement in exercise performance will be classified as acutely fatigued.
Contact information is provided by the study sponsor or research team.
Sofie Engelborghs
CONTACT
Stefan De Smet
CONTACT
Stefan De Smet
Other
ICARUS: Impact of Training Load on the Gut Microbiome and Its Relation to Exercise Performance, Muscle Phenotype, and Markers of Overreaching in Healthy Men: Phase A - Human Trial
Acronym: ICARUS
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