Physical Inactivity
Conditions
Keywords
High-Intensity Interval Training, Histone Lactylation, H3K18la, Monocytes, Trained Immunity, Innate Immunity, Blood Lactate, Epigenetics
Brief summary
This randomized, work-matched training-detraining-rechallenge study evaluated whether repeated high-intensity interval training produces persistent functional and epigenetic adaptations in human monocytes. Eighty healthy physically inactive adults aged 18-30 years were randomly assigned in a 2:2:1 ratio to high-intensity interval training (HIIT), moderate-intensity continuous training (MCT), or a non-training control group. HIIT and MCT completed 6 weeks of supervised cycling training matched for external mechanical work, followed by 4 weeks without prescribed structured training. All groups subsequently completed a standardized 30-minute cycling rechallenge. The primary outcome was the between-group difference in change in ex-vivo LPS-stimulated TNF-alpha release from baseline to the post-detraining assessment. Secondary outcomes included IL-6 release, H3K18 lactylation, monocyte subset distribution, blood lactate exposure, chromatin accessibility, H3K18la CUT&Tag occupancy, and RNA-seq transcriptional profiling.
Detailed description
Participants completed baseline cardiopulmonary exercise testing to determine peak oxygen uptake (VO2peak) and maximal aerobic power (MAP). Eligible participants were randomized to HIIT, MCT, or Control. The HIIT intervention consisted of 4 × 4-minute cycling intervals at 85% MAP separated by active recovery at 40% MAP, with standardized warm-up and cool-down periods. The MCT intervention consisted of continuous cycling at 55% MAP for a duration selected to match the external mechanical work of the HIIT sessions. Both exercise groups trained three times per week for 6 weeks. Blood and monocyte assessments were performed at baseline (T0), 72 hours after the final training session (T1), after 4 weeks without prescribed structured training (T2\_PRE), and 120 minutes after a standardized 30-minute cycling rechallenge at 60% of baseline MAP (T3\_POST). Classical monocytes were isolated and stimulated ex vivo with ultrapure Escherichia coli O111:B4 lipopolysaccharide (10 ng/mL) for 24 hours. TNF-alpha was the prespecified primary functional outcome, with IL-6 as a key secondary outcome. Epigenetic and molecular analyses included global H3K18 lactylation, ATAC-seq chromatin accessibility profiling, H3K18la CUT&Tag, and RNA-seq. Blood lactate kinetics and cumulative lactate exposure across the training intervention were also assessed. The primary comparison was HIIT versus MCT. The Control group served as a supportive non-training reference. Longitudinal outcomes were analyzed using mixed models for repeated measures under the intention-to-treat principle.
Interventions
BEHAVIORALHigh-Intensity Interval Training
Supervised cycling HIIT performed three sessions per week for 6 weeks. Each session included a 5-minute warm-up at 30% MAP, four 4-minute intervals at 85% MAP separated by three 3-minute active recovery periods at 40% MAP, and a 5-minute cool-down at 30% MAP.
BEHAVIORALModerate-Intensity Continuous Training
Supervised moderate-intensity continuous cycling performed three sessions per week for 6 weeks. Each session included a 5-minute warm-up at 30% MAP, 31 minutes and 16 seconds of continuous cycling at 55% MAP, and a 5-minute cool-down at 30% MAP. The protocol was designed to match the external mechanical work performed by the HIIT group.
Sponsors
Saif Rasheed Ghanim
Study design
Allocation
RANDOMIZED
Intervention model
PARALLEL
Primary purpose
BASIC_SCIENCE
Masking
SINGLE (Outcomes Assessor)
Masking description
Laboratory personnel performing blood processing, monocyte isolation, ex-vivo cytokine assays, flow cytometry, epigenetic assays, sequencing, and bioinformatic analyses were blinded to group allocation. Participants and exercise physiologists supervising the interventions were not blinded because of the nature of the exercise protocols.
Intervention model description
Participants were randomized in a 2:2:1 ratio to HIIT, MCT, or a non-training Control group and were followed in parallel through the training, detraining, and standardized exercise-rechallenge phases.
Eligibility
Sex/Gender
ALL
Age
18 Years to 30 Years
Healthy volunteers
Yes
Inclusion criteria
Inclusion Men and women aged 18-30 years. Healthy adults. Physically inactive, defined as participating in less than 150 minutes per week of structured moderate-to-vigorous exercise during the preceding 6 months.
Exclusion criteria
* Major cardiovascular, metabolic, respiratory, renal, endocrine, inflammatory, autoimmune, or hematological disease. Current tobacco use. Current use of relevant immunomodulatory, anti-inflammatory, corticosteroid, or cardiovascular medications. Acute infection within 4 weeks before baseline assessment. Musculoskeletal contraindications to maximal cycling exercise. Pregnancy or lactation.
Design outcomes
Primary
| Measure | Time frame | Description |
|---|---|---|
| Change in LPS-Stimulated TNF-Alpha Release From Classical Monocytes | Baseline (T0) and 4 weeks after completion of the 6-week training intervention (T2_PRE). | TNF-alpha concentration (pg/mL) was measured in culture supernatants after 24-hour ex-vivo stimulation of purified classical CD14++CD16- monocytes with 10 ng/mL ultrapure Escherichia coli O111:B4 lipopolysaccharide. The primary outcome was the change from baseline (T0) to the post-detraining assessment (T2\_PRE), with the primary comparison between the HIIT and MCT groups. |
Secondary
| Measure | Time frame | Description |
|---|---|---|
| Change in LPS-Stimulated IL-6 Release From Classical Monocytes | Baseline (T0) and 4 weeks after completion of the 6-week training intervention (T2_PRE). | IL-6 concentration (pg/mL) was measured in culture supernatants after 24-hour ex-vivo stimulation of purified classical CD14++CD16- monocytes with 10 ng/mL ultrapure Escherichia coli O111:B4 lipopolysaccharide. The outcome was the change from baseline (T0) to the post-detraining assessment (T2\_PRE), with comparison between the HIIT and MCT groups. |
| Exercise-Rechallenge Change in LPS-Stimulated TNF-Alpha Release From Classical Monocytes | Immediately before the standardized exercise rechallenge (T2_PRE) and 120 minutes after the rechallenge (T3_POST). | TNF-alpha concentration (pg/mL) was measured after 24-hour ex-vivo stimulation of purified classical CD14++CD16- monocytes with 10 ng/mL ultrapure Escherichia coli O111:B4 lipopolysaccharide. The outcome was the within-participant change from the post-detraining pre-rechallenge assessment (T2\_PRE) to 120 minutes after the standardized exercise rechallenge (T3\_POST), with the principal comparison between the HIIT and MCT groups. |
| Exercise-Rechallenge Change in LPS-Stimulated IL-6 Release From Classical Monocytes | Immediately before the standardized exercise rechallenge (T2_PRE) and 120 minutes after the rechallenge (T3_POST). | IL-6 concentration (pg/mL) was measured after 24-hour ex-vivo stimulation of purified classical CD14++CD16- monocytes with 10 ng/mL ultrapure Escherichia coli O111:B4 lipopolysaccharide. The outcome was the within-participant change from the post-detraining pre-rechallenge assessment (T2\_PRE) to 120 minutes after the standardized exercise rechallenge (T3\_POST), with the principal comparison between the HIIT and MCT groups. |
| Change in Monocyte H3K18 Lactylation Across Study Time Points | Baseline (T0), 72 hours after the final training session (T1), 4 weeks after training cessation before rechallenge (T2_PRE), and 120 minutes after the standardized exercise rechallenge (T3_POST). | Histone H3 lysine 18 lactylation (H3K18la) was quantified in purified classical CD14++CD16- monocytes and normalized to total histone H3 abundance. H3K18la was assessed to determine training-induced elevation, persistence after 4 weeks without prescribed structured training, and re-amplification following the standardized exercise rechallenge. |
| Genome-Wide Chromatin Accessibility in Classical Monocytes | Four weeks after completion of the 6-week training intervention, before the standardized exercise rechallenge (T2_PRE). | Genome-wide chromatin accessibility was assessed by ATAC-seq in purified classical CD14++CD16- monocytes. Differentially accessible regions between the HIIT and MCT groups were identified using a false discovery rate (FDR) \<0.05 and an absolute log2 fold-change \>1.0. |
| Genome-Wide H3K18la Occupancy in Classical Monocytes | Four weeks after completion of the 6-week training intervention, before the standardized exercise rechallenge (T2_PRE). | Genome-wide histone H3 lysine 18 lactylation (H3K18la) occupancy was profiled by CUT\&Tag in purified classical CD14++CD16- monocytes. H3K18la-enriched genomic regions were identified and their overlap with differentially accessible ATAC-seq regions was evaluated between the HIIT and MCT groups. |
| Genome-Wide Differential Gene Expression in Classical Monocytes | Four weeks after completion of the 6-week training intervention, before the standardized exercise rechallenge (T2_PRE). | Genome-wide transcript abundance was assessed by RNA sequencing (RNA-seq) in purified classical CD14++CD16- monocytes. Differential gene expression between the HIIT and MCT groups was evaluated using DESeq2, with differentially expressed genes defined using a false discovery rate (FDR) \<0.05 and an absolute log2 fold-change \>1.0. |
Countries
Iraq
Contacts
PRINCIPAL_INVESTIGATORSaif Rasheed Ghanim, PhD
University of Samarra, College of Physical Education and Sports Sciences
Outcome results
None listed