This study aims to investigate the effects of omega-3 fatty acid supplementation on muscle performance, neuromuscular function, inflammation, and oxidative stress at rest and after eccentric exercise. The findings may provide insights into the role of omega-3 fatty acids in muscle function, physiology, and metabolism following muscle-damaging exercise. Additionally, the effects of omega-3 fatty acids on cardiovascular health will be investigated.
The study is a double-blind, randomized controlled trial. Two parallel groups of 16 healthy males and females each (sex counterbalanced, 20-45 years) will randomly take 3 g of fish oil \[1500 mg EPA (eicosapentaenoic acid) + 750 mg DHA (docosahexaenoic acid), Now Foods, USA\] or a placebo (3 g of olive oil) for 12 weeks. Before taking the supplement, participants will perform a unilateral isokinetic eccentric exercise session of maximal voluntary intensity (10 repetitions x 10 sets of knee extensions at a speed of 60 degrees per second). Dominant and non-dominant legs will be allocated equally and randomly across participants. Physiological assessments and blood samples will be collected immediately before exercise and 2 and 4 days after eccentric exercise (bout 1). Then, participants will start the supplementation for 12 weeks. At the end of the supplementation period, participants will perform the same eccentric exercise protocol (bout 2) with their opposite leg. The same assessments and blood sampling will be conducted at the same time points (i.e., immediately before and 2 and 4 days after eccentric exercise).
Study Type
INTERVENTIONAL
Allocation
RANDOMIZED
Purpose
SUPPORTIVE_CARE
Masking
QUADRUPLE
Enrollment
32
Participants will take three capsules per day (a total of 1,500 mg EPA and 750 mg DHA). They will take 1 g with each main meal (breakfast, lunch, and dinner).
Participants will take 3 g of olive oil. They will take 1 g with each main meal (breakfast, lunch, and dinner).
European University Cyprus
Nicosia, Engomi, Cyprus
Muscle Strength
Muscle strength of the knee extensors will be measured using an isokinetic dynamometer during maximal voluntary contractions. The unit of measure is Newton-meters (Nm).
Time frame: Changes in muscle strength after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Maximal Oxygen Uptake (VO₂ max)
VO₂ max will be assessed during a graded exercise test on a cycle ergometer. The measure unit is milliliters per kilogram of body mass per minute (ml/kg/min).
Time frame: Changes in VO₂ max after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Muscle Fatigue Index (MFI)
MFI will be calculated as the percentage decline (%) in peak torque across a series of repeated maximal contractions of the knee extensor muscles on an isokinetic dynamometer.
Time frame: Changes in MFI after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Muscle Oxygenation
Muscle oxygenation will be measured in the vastus lateralis using near-infrared spectroscopy (NIRS). Parameters include oxyhemoglobin (O₂Hb), deoxyhemoglobin (HHb), and total hemoglobin levels, which will be expressed in arbitrary units (AU).
Time frame: Changes in muscle oxygenation after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Creatine Kinase
Creatine kinase activity will be measured in International Units (IU) via the enzymatic colorimetric method in fasting plasma samples.
Time frame: Change in creatine kinase activity after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
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Range of Motion (ROM)
ROM of the knee joint will be measured in degrees (°) using an isokinetic dynamometer, with full extension defined as 0° and flexion recorded up to 120°.
Time frame: Change in ROM after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Muscle Thickness
Muscle thickness of the vastus lateralis will be measured using B-mode ultrasound with a linear probe, in millimeters (mm).
Time frame: Change in muscle thickness after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Echo Intensity
Echo intensity of the vastus lateralis will be assessed using B-mode ultrasound with a linear probe and quantified from grayscale image analysis, expressed in arbitrary units (0-255).
Time frame: Change in echo intensity after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Blood Lipids Profile
Fasting blood samples will be collected and analyzed to assess the participant's lipid profile, including: Total Cholesterol (TC). Expressed in mg/dL. Low-Density Lipoprotein Cholesterol (LDL-C). Expressed in mg/dL. High-Density Lipoprotein Cholesterol (HDL-C). Expressed in mg/dL. Triglycerides (TG). Expressed in mg/dL. Blood will be drawn after a minimum of 8-12 hours of fasting. Lipid concentrations will be determined using standard enzymatic colorimetric methods on an automated clinical chemistry analyzer.
Time frame: Change in blood lipids from baseline (immediately before supplementation) to 12 weeks after the supplementation period.
Omega-3 Index
The Omega-3 Index will be assessed as a biomarker of long-chain omega-3 fatty acid status. It is defined as the combined percentage (%) of eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) in red blood cell (RBC) membranes, expressed as a proportion of total identified fatty acids. Blood samples will be collected following an overnight fast, and red blood cells will be isolated for fatty acid analysis. EPA and DHA levels will be quantified via gas chromatography, and the Omega-3 Index will be reported as a percentage (%).
Time frame: Change in omega-3 index in red blood cells from baseline (immediately before supplementation) to 12 weeks after the supplementation period.
Rate of Force Development (RFD).
RFD will be calculated as the slope of the torque-time curve during the early phase of voluntary contraction. It will be measured in Newton-meters per second (Nm/s).
Time frame: Change in RFD after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Electromechanical Delay (EMD)
EMD will be defined as the time lag between the onset of electrical activity (via surface EMG) and the onset of force production, measured in milliseconds (ms).
Time frame: Change in EMD after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Force Matching Error (FME)
FME will be assessed during a submaximal isometric contraction using an isokinetic dynamometer and expressed as the percentage difference between the target and produced force (%).
Time frame: Change in FME after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Joint Position Sense (JPS)
JPS will be evaluated as the absolute angular difference between a target knee joint angle and the participant's reproduced angle, measured in degrees (°) using an isokinetic dynamometer.
Time frame: Change in JPS after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Delayed Onset Muscle Soreness (DOMS).
DOMS will be assessed using a 1-10 visual analog scale (VAS), where 1 indicates no pain and 10 indicates extreme pain.
Time frame: Change in DOMS after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Countermovement Jump (CMJ)
CMJ will be measured in centimeters (CM) using a force plate.
Time frame: Change in CMJ after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Contact Time
Duration of foot contact with the force plate during landing, expressed in milliseconds (ms).
Time frame: Change in contact time after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Stability Index
The stability index will be assessed from the center of pressure displacement using a force platform and expressed in millimeters (mm).
Time frame: Change in stability index after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
F2-Isoprostanes
F2-Isoprostanes will be quantified as a marker of lipid peroxidation using ELISA and expressed in picograms per milliliter (pg/mL).
Time frame: Change in F2-Isoprostanes after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Glutathione
Glutathione will be measured as an indicator of endogenous antioxidant capacity using a colorimetric assay and expressed in micromoles per gram of hemoglobin (μmol/g Hb).
Time frame: Change in glutathione after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Protein carbonyls
Protein carbonyls will be measured as a marker of oxidative protein damage using a spectrophotometric assay and expressed in nanomoles per gram of hemoglobin (nmol/g Hb).
Time frame: Change in protein carbonyls after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Interleukin-6 (IL-6)
IL-6 will be measured as a marker of inflammation using ELISA and expressed in picograms per milliliter (pg/mL).
Time frame: Change in IL-6 after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Tumor Necrosis Factor-alpha (TNF-α)
TNF-α will be measured as a marker of inflammation using ELISA and expressed in picograms per milliliter (pg/mL).
Time frame: Change in TNF-α after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Pennation Angle
The pennation angle of the vastus lateralis will be measured under resting conditions using longitudinal B-mode ultrasound imaging and expressed in degrees (°).
Time frame: Change in pennation angle after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Fascicle Length
Fascicle length of the vastus lateralis will be determined under resting condition from B-mode ultrasound images, either measured directly from visible fascicles or estimated using trigonometric calculations, and expressed in millimeters
Time frame: Change in fascile length after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Muscle Stiffness
Muscle stiffness, reflecting the resistance of the muscle to an external force, will be measured and expressed in newtons per meter (N/m) using the Myoton Pro device.
Time frame: Change in muscle stifness after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Muscle Tone
Muscle tone will be assessed using the Myoton Pro device, representing the natural oscillation frequency of the muscle at rest, and expressed in hertz (Hz).
Time frame: Change in muscle tone after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Mechanical Stress Relaxation Time
Mechanical stress relaxation time will be assessed using the Myoton Pro device, representing the time required for the muscle to return to its original shape after a mechanical impulse, and expressed in milliseconds (ms).
Time frame: Change in mechanical stress relaxation time after the first bout (week 0) and the second bout of exercise (week 12) from baseline (immediately before each bout) and at day 2 and day 4 after each exercise bout.
Central and Peripheral Blood Pressure
Central and peripheral systolic and diastolic blood pressure will be measured using the SphygmoCor system and expressed in millimeters of mercury (mmHg).
Time frame: Change in blood pressure from baseline (immediately before supplementation) to 12 weeks after the supplementation period.
Arterial stiffness
Arterial stiffness will be assessed using the SphygmoCor system by measuring carotid-femoral pulse wave velocity (PWV), expressed in meters per second (m/s).
Time frame: Change in arterial stiffness from baseline (immediately before supplementation) to 12 weeks after the supplementation period.