Impact of Circadian Exercise on Metabolic Dysfunction-Associated Steatotic Liver Disease in Postmenopausal Women

Metabolic dysfunction-associated steatotic liver disease (MASLD) affects approximately one in three adults and is a major contributor to the growing burden of cardiometabolic disease. Exercise is one of the most effective interventions for improving cardiometabolic health, reducing hepatic fat, and enhancing metabolic flexibility. However, the timing of exercise -a modifiable behavioral factor- may play a crucial yet underexplored role in determining its physiological benefits.

Preclinical studies have shown that circadian rhythms regulate key metabolic processes, including lipid metabolism and glucose homeostasis. Moreover, recent findings suggest that exercise performed at different times of day may elicit different responses, influencing the regulation of hepatic fat and systemic inflammation. These effects may be mediated, in part, through gut microbiota, which modulate host metabolism via the gut-liver axis. However, the interaction between exercise timing and gut microbiota in human physiology -particularly in women- remains poorly understood.

This knowledge gap is particularly critical in postmenopausal women, a population that experiences profound metabolic changes due to hormonal decline, including increased hepatic and visceral fat, chronic inflammation, and reduced insulin sensitivity. These alterations significantly elevate the risk for MASLD and other cardiometabolic diseases. Yet, postmenopausal women are consistently underrepresented in clinical trials exploring exercise interventions.

Based on emerging scientific evidence, the investigators hypothesize that morning exercise may lead to greater reductions in hepatic fat and improvements in cardiometabolic health compared to evening exercise in postmenopausal women with MASLD. Furthermore, the investigators propose that these effects may be partially mediated by exercise-induced changes in gut microbiota composition and function.

Thus, the main objective of the project is to investigate whether the timing of exercise modulates hepatic fat reduction, cardiometabolic adaptation, and gut microbiota remodeling in postmenopausal women with MASLD. To achieve this, the project will implement a randomized controlled trial in which 63 sedentary postmenopausal women diagnosed with MASLD will be randomly allocated into one of three groups: (i) morning exercise (07:00h), (ii) evening exercise (19:00h), or (iii) a usual-care control group receiving lifestyle recommendations.

Participants in the intervention groups will complete a 12-week supervised training program, combining aerobic and resistance exercise (3 sessions/week, 60-90 minutes per session), in accordance with WHO recommendations. Before and after the intervention, biological samples, magnetic resonance imaging, and metabolic assessments will be collected. The study will employ advanced multi-omics analyses, including metabolomics and semi-targeted metagenomics, to explore how gut microbiota changes relate to improvements in metabolism.

Inclusion criteria

• Women aged 45-75 years, postmenopausal for at least two years (stage +1a)
• Body Mass Index (BMI) > 25 and < 40 kg/m²
• Diagnosed hepatic steatosis (via ultrasound hyperechogenicity, FibroScan CAP score >280, or histological confirmation)
• Sedentary lifestyle (no regular structured exercise)
• Willingness to be randomized and adhere to study procedures, including all assessments and visits
• Sufficient Spanish proficiency to understand and follow study instructions
• Consent to store biological samples for future research
• Participants should have a healthy circadian rhythm

Exclusion criteria

• Contraindications for MRI (e.g., claustrophobia, pacemaker, metal implants)
• History of major cardiovascular, endocrine, neurological, or kidney disease, or any clinical abnormalities (to be judged by the study physician)
• First-degree family history of sudden cardiac death
• Alcohol or substance abuse
• Psychiatric, psychotic, eating, or sleep disorders (to be judged by the study physician)
• Prior bariatric surgery, diagnosed HIV/AIDS, or inflammatory/autoimmune diseases
• Cancer or any medical condition where exercise is contraindicated (to be judged by the study physician)
• Recent or unstable metabolic conditions (e.g., diabetes, recent medication changes, or use of drugs affecting metabolism)
• Recent (<3 months) use of antibiotics, statins, glucocorticoids, hormonal therapies, amiodarone, or myelosuppressive agents
• Participation in weight-loss programs or special diets (e.g., ketogenic, high-carb)
• Shift workers or caregivers with frequent nocturnal disruptions