Peripheral arterial disease (PAD) affects over 200 million people worldwide and is caused by narrowing of the arteries. Intermittent claudication, characterized by pain when walking, is its main symptom. Despite the proven benefits of exercise, there is no optimal protocol for treatment. To analyze the effects of different exercise conditions on proteomic and physiological markers in patients with PAD and to determine their preferences regarding exercise conditions. A clinical cross-over study with four randomized experimental conditions (aerobic training, traditional strength training, circuit training and control) will be conducted. Patients with grade IIa-IIb PAD according to the Leriche-Le Fontaine scale will be recruited from the Department of Angiology and Vascular Surgery Department of the Hospital Clínico Universitario de Valladolid. In each experimental session, measurements of cardiorespiratory capacity, post-exercise oxygen debt, ankle-brachial index, blood flow, peripheral oxygen saturation, perceived exertion, heart rate variability and blood proteome changes will be performed.
Study Type
INTERVENTIONAL
Allocation
RANDOMIZED
Purpose
TREATMENT
Masking
SINGLE
Enrollment
16
Intermittent walking will be performed on a treadmill (F2W DUAL, BHFitness®, Spain). There will be 8 sets of 2 minutes in duration at an intensity equivalent to the level at which the patient experienced claudication symptoms. The recovery period between each set will be two minutes. All training sessions will conclude with a cool-down period of 5 minutes of walking at 50-60% of peak heart rate, followed by 3 minutes of joint mobility exercises. Throughout the session, heart rate will be continuously monitored using a heart rate monitor (Polar H10, Polar Electro Ltd., Kempele, Finland) and the Polar Team application (Polar Electro Ltd., Kempele, Finland).
Two sets will be performed with a moderate-to-high effort level (15-16 repetitions out of 20RM) consisting of 6 multi-joint exercises. The recovery period after each set will be two minutes of passive rest. Each participant will be asked to move the load as quickly as possible during each repetition. All training sessions will conclude with a cool-down period of 5 minutes of walking at 50-60% of peak heart rate, followed by 3 minutes of joint mobility exercises. Throughout the session, heart rate will be continuously monitored using a heart rate monitor (Polar H10, Polar Electro Ltd., Kempele, Finland) and the Polar Team application (Polar Electro Ltd., Kempele, Finland).
Two sets (rounds) will be performed with a moderate-to-high effort level (15-16 repetitions out of 20RM) in a circuit consisting of 6 multi-joint exercises. The recovery time between exercises will be the minimum required to change exercises, and the recovery between sets (rounds) will be two minutes of passive rest. Each participant will be asked to move the load as quickly as possible during each repetition. All training sessions will conclude with a cool-down period of 5 minutes of walking at 50-60% of peak heart rate, followed by 3 minutes of joint mobility exercises. Throughout the session, heart rate will be continuously monitored using a heart rate monitor (Polar H10, Polar Electro Ltd., Kempele, Finland) and the Polar Team application (Polar Electro Ltd., Kempele, Finland).
Blood Flow
An ultrasound machine will be used to measure blood flow in the femoral artery, quantifying changes in the velocity and diameter of the artery in the lower extremity with the lowest ABI. Blood flow measurements will be conducted with the participant in a supine position. During the first experimental visit, the measurement site will be determined, where all measurements will take place. This measurement site corresponds to the common femoral artery, just below the inguinal ligament, and approximately 2 cm above the bifurcation into the superficial and deep femoral arteries.
Time frame: Week 1 (baseline), Weeks 2-4 (15 minutes post-intervention)
Peripheral oxygen saturation
Changes in peripheral oxygenation levels in the gastrocnemius will be evaluated during physical exercise. For this purpose, near-infrared spectroscopy (NIRS) will be used, which utilizes a wavelength of 700-900 nm to assess the concentration of oxygenated and deoxygenated hemoglobin and the total blood volume. The NIRS device MOXY Monitor (Moxy, Fortiori Design LLC, Minneapolis, MN, USA) will be used.
Time frame: Week 1 (baseline), Weeks 2-4 (during each intervention session)
Heart rate variability
By monitoring the heart rate of participants, heart rate variability (HRV) will be assessed, which reflects the variation in the time intervals between beats (R-R intervals) and serves as a useful tool for monitoring patient health, as it estimates the balance between the sympathetic and parasympathetic branches of the autonomic nervous system. To evaluate HRV, a Polar H10 heart rate monitor (Polar Electro Oy, Kempele, Finland) and a mobile application, specifically Elite HRV, will be used. Heart rate variability will be monitored daily during the week leading up to the initial assessment visits. Participants will be instructed to take the measurement for the first 5 minutes after waking up and while fasting.
Time frame: Week 1 (baseline), Weeks 2-4 (during intervention and 24, 48, and 72 hours post-intervention)
Proteomics
A blood sample will be collected during the last 5 minutes of the pre-session and after completing the experimental condition, just before a 15-minute post-session rest. Venous blood will be drawn from the antecubital vein and stored at 4°C to prevent clotting and minimize protein degradation. Samples will be centrifuged at 1500 g for 10 minutes at 4°C, discarding the remaining sample. Ten microliters of protease inhibitor will be added to each 1.0 ml plasma aliquot, stored at -80°C for later analysis, with only one freeze-thaw cycle permitted. All samples will be prepared within 1 hour of collection and must show no hemolysis. Peptide identification will be performed using Proteome Discoverer (version 2.1.0.81) and SEQUEST-HT, analyzing data in the Uniprot database (January 25, 2024; 20,433 reviewed entries). Identified peptides with a false discovery rate (FDR) of 1% or less will quantify the relative abundance of each protein.
Time frame: Weeks 2-4 (5 minutes prior to exercise and 5 minutos post-intervention)
Post-exercise oxygen debt
Post-exercise oxygen debt (EPOC) will be calculated every minute for 12 minutes following the completion of exercise using the following formula: EPOC (L·min-¹) = post-exercise VO2 (L·min-¹) - resting VO2 (L·min-¹). Subsequently, EPOC will be converted into energy expenditure with the conversion of 1L of O2 = 4.64 kcal to exclude the rapid glycolytic resynthesis of adenosine triphosphate (ATP) as part of the conversion of O2 consumption into energy expenditure (EE).
Time frame: Weeks 2-4 (after every intervention session)
Perceived exertion
During the four experimental conditions, perceived exertion will be monitored using the Rating of Perceived Exertion (RPE) scale. The RPE will be recorded after each set or circuit round. Immediately after completing the last set of each condition, participants will be asked to indicate their perceived exertion for the entire session. Scales go from 0 (no exertion) to 10 (maximal exertion possible).
Time frame: Weeks 2-4 (right after every intervention session and 24, 48, and 72 hours post-intervention)
Satisfaction with the type of training
At the end of each session, satisfaction will be evaluated using a numerical scale from 0 to 10, where the highest score indicates maximum satisfaction with the type of training.
Time frame: Weeks 2-4 (right after every intervention session and 24 and 48 post-intervention)
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