Aging is a major risk factor for cardiovascular disease (CVD), contributing to progressive macrovascular and microvascular dysfunction. Macrovascular impairments, such as arterial stiffening and endothelial dysfunction, reduce nitric oxide (NO) bioavailability, leading to hypertension and impaired blood flow regulation. Microvascular dysfunction exacerbates cardiovascular decline by compromising capillary perfusion and tissue oxygenation, increasing susceptibility to ischemic events. Preserving vascular function is essential for maintaining cardiovascular health in aging populations.

Low-intensity aerobic exercise (LIAE) improves cardiovascular health and mobility, particularly in populations unable to perform high-intensity exercise. However, LIAE alone often fails to induce significant vascular adaptations, such as improved arterial compliance or endothelial function, highlighting the need for optimized interventions.

Blood Flow Restriction (BFR) enhances vascular adaptations during low-intensity exercise by inducing localized hypoxia, stimulating endothelial function and arterial remodeling. While BFR may transiently increase blood pressure and arterial stiffness, its vascular benefits resemble those of high-intensity exercise at lower workloads. However, research on BFR's acute vascular effects during aerobic exercise in older adults remains limited.

Current studies have focused on perceptual and hemodynamic responses to LIAE+BFR but lack direct assessments of macrovascular (e.g., flow-mediated slowing) and microvascular function. Older adults may exhibit distinct vascular responses, yet data are scarce. This study examines acute vascular responses to LIAE+BFR and high-intensity interval exercise (HIIE) in young and older adults.

Hypothesis:

1. BFR combined with LIAE will elicit greater acute microvascular and macrovascular responses in older adults than in younger adults.
2. These responses will be comparable to HIIE without BFR, suggesting BFR enhances LIAE's efficacy to high-intensity levels.
3. Any transient vascular dysfunction (e.g., increased arterial stiffness) will normalize within 30 minutes post-exercise, supporting safety in aging populations.

In this parallel-group crossover randomized trial, young and older adult participants will be randomly assigned to two experimental conditions: LIAE+BFR and HIIE. These two conditions were specifically chosen because they are known to induce similar haemodynamic responses. Each participant will visit the laboratory on three separate occasions, with at least one week between visits allowing recovery and minimizing potential carryover effects.

On Visit 1, participants will be familiarized with the exercise and testing protocols, undergo body composition measurements, and have baseline macrovascular and microvascular function evaluated twice, 30 minutes apart, in a resting condition (control). On Visits 2 and 3, participants will perform the experimental conditions in a simple randomized order, with allocation concealed until arrival at the laboratory. The simple randomization scheme sequence will be generated by Dr. XM using a web-based randomization tool (http://www.randomizer.org/) and placed in sequentially numbered, opaque, sealed envelopes to ensure allocation concealment and minimize bias. Researchers conducting field assessments will remain blinded to group allocation until the day of the first visit.

Each session will begin with 15 minutes of supine rest on a cushioned examination table, followed by baseline assessments of brachial (bSBP) and central blood pressure (cSBP), heart rate (HR), microvascular function using finger photoplethysmography (Vicorder, 80 beats medical, Berlin, Germany)., macrovascular function using FMS, and central PWV (cPWV).

After baseline assessments, participants will engage in one of the randomized aerobic exercise conditions, lasting approximately 30 minutes. The workload will be equalized considering intensity and stimulus time (intensity x duration) and the protocol is directly inspired by a previous study. In the LIAE+BFR condition, participants will walk on a treadmill (Technogym Excite Run, Technogym SpA, Cesena, Italy) for 20 minutes, using a 5 cm × 75 cm pneumatic cuff (Occlusion Cuff Pro®, Occlusion Cuff LLC, 2024) inflated to 1.3 times the individual's ankle SBP (Suga et al., 2012) The intensity will be set between 30-39% of heart rate reserve (HRR), a widely used method for prescribing aerobic exercise intensity (Liguori et al., 2021). In the HIIE condition, participants will perform a 20-minute treadmill-based high-intensity interval training, alternating between 60-seconds high-intensity intervals at 60-89% HRR and 60-second active recovery intervals at 40-50% HRR. Treadmill speed will be adjusted dynamically to maintain target intensities, while the incline remains at 1%. HR, the Modified Borg Rate of Perceived Exertion (RPE) scale, and the Rating of perceived discomfort will be continuously monitored throughout both exercise sessions to ensure adherence to the prescribed intensity levels.

Immediately after exercise, participants will return to the examination table for a 30-minute recovery period, during which brachial blood pressure, heart rate, and micro- and macrovascular function will be reassessed at 5- and 30-minutes post-exercise and compared to resting values. These post-exercise time points aim to characterize the biphasic response of macro- and microvascular function.

All participants will report to the laboratory between 8:00 AM and 12:00 PM, with room temperature controlled between 20-22°C. Participants will be instructed to avoid caffeine and alcohol for 24 hours, fully void before the session, and refrain from vigorous physical activity for 24 hours before each session, including the familiarization session.

All evaluations will be conducted by four trained physiotherapy students per participant (4:1), each with over 30 hours of training in the evaluation and exercise protocols. Throughout all sessions, a certified professional in basic life support (BLS) and automated external defibrillator (AED) use will be present at the clinic.

Inclusion Criteria:

• Aged between 18-30 years and those over 60 years.

Exclusion Criteria:

• Past or current history of coronary heart disease, stroke, oncological disease, or major cardiovascular events.
• Individuals who had undergone surgery within the last two months
• Body mass index (BMI) greater than 30 kg/m²
• Declared sleep apnea
• Active kidney or liver disease
• Active tobacco smokers
• Sensory impairments
• Neurological or orthopedic functional impairments, musculoskeletal pathologies affecting exercise capacity
• Physically active for more than six months with a weekly activity level of 1,000 MET/min.

Data obtained in this study may be provided to qualified researchers with academic interest in vascular responses to aerobic training with blood flow restriction. Data or samples shared will be coded, with no PHI included. Approval of the request and execution of all applicable agreements (i.e. a material transfer agreement) are prerequisites to the sharing of data with the requesting party.

Data requests can be submitted starting 9 months after article publication and the data will be made accessible for up to 24 months. Extensions will be considered on a case-by-case basis.

Access to trial IPD can be requested by qualified researchers engaging in independent scientific research and will be provided following review and approval of a research proposal and Statistical Analysis Plan (SAP) and execution of a Data Sharing Agreement (DSA). For more information or to submit a request, please contact xmelo@egasmoniz.edu.pt

Outcomes will be assessed by the same experts in every time point. Participants and evaluators will be blinded until arrival of the participant for the fist experimental condition.