Clinical Trial Details
— Status: Completed
Administrative data
| NCT number |
NCT05753111 |
| Other study ID # |
S67315 |
| Secondary ID |
|
| Status |
Completed |
| Phase |
N/A
|
| First received |
|
| Last updated |
|
| Start date |
February 16, 2023 |
| Est. completion date |
May 26, 2023 |
Study information
| Verified date |
February 2024 |
| Source |
KU Leuven |
| Contact |
n/a |
| Is FDA regulated |
No |
| Health authority |
|
| Study type |
Interventional
|
Clinical Trial Summary
This study evaluates the effects of short term palmitoylethanolamide (PEA) supplementation on
functional responses (i.e. muscle soreness and performance parameters) to strenuous eccentric
exercise, and investigateq the systemic and muscle molecular mechanisms through which PEA
impacts on these functional responses.
Description:
Study design The protocol entails a double-blind, placebo-controlled, crossover trial design
in which non-elite male athletes (training load >5h/wk, n=14) are enrolled. Participants
undergo 2 experimental sessions in which they receive PEA or placebo in a randomized order.
The two sessions are separated by a 3-week wash out period to allow a full recovery from the
eccentric exercise bout. The scientific rationale to use a within-subjects design is based on
the assumption of reduced data variability and increased statistical power compared to a
between-subjects design. Additionally, a double-blind placebo controlled trial is used to
prevent treatment bias and placebo effects.
Participants are randomly allocated to receive PEA or appearance-matched placebo supplements
twice a day for a total of 6 days. Three days before the start of supplementation (D-3),
baseline measurements are collected during which participants report to the laboratory in a
fasted and rested state and mark their perceived muscle soreness on a visual analogue scale
(VAS). Next, blood samples and muscle biopsies are collected, after which maximal isometric,
eccentric and concentric voluntary contractions and jump height (Counter Movement Jump and
Squat Jump) are assessed. Furthermore, participants complete the Pittsburgh Sleep Quality
Index (PSQI) to evaluate sleep quality and quantity of the previous month. Three days after
the collection of baseline measurements (D0), participants return to the laboratory in a
rested and fasted state. After collection of a fasted blood sample, they receive their first
supplement with a light standardized breakfast (~500 kcal, 60% carbohydrates, 20% protein,
20% fat). Afterwards, they rate their perceived muscle soreness and fill in the St. Mary's
Hospital questionnaire (to evaluate their sleep quality and quantity from the previous
night). One hour after supplement intake, participants perform a unilateral eccentric
exercise (EE) bout consisting of 24 sets of 10 eccentric contractions of the knee extensors.
Measurements of maximal voluntary contractions and jump height are repeated 24, 48, 72 and
120h following EE. Additionally, blood samples are collected 24, 48, 72, 96 and 120h
following EE, and muscle biopsies are collected 48h following EE. Furthermore, participants
rate their perceived muscle soreness, complete a food diary and fill in the St. Mary's
Hospital questionnaire (daily), for the entire duration of the experimental protocol.
Screening Volunteers are screened to assess their eligibility to participate in the study
based on in- and exclusion criteria . During the screening, body mass is measured using a
digital balance with an accuracy of 0.1 kg and body height is measured with an accuracy of 1
cm. Furthermore, a medical questionnaire is obtained to assess participants' general health.
When volunteers are deemed eligible according to the screening results and the in- and
exclusion criteria, they are included in the study.
Familiarization To prevent any learning effect during the intervention trial, all
participants are familiarized with the test equipment at least one week before the start of
the trial. Participants are invited to perform 3 submaximal isometric, eccentric and
concentric voluntary contractions and 3 counter movement jumps and squat jumps as described
in detail below (see paragraph 76.3 on page 12). Participants are instructed about the proper
technique of the movements and are supervised by a staff member at all times. During the
familiarization, individual settings on the isokinetic dynamometer are determined and will be
used in all subsequent measurements. Additionally, participants are instructed on how to
complete their food diaries.
Randomization: Participants are randomly allocated to one of the supplemental groups (PEA
orplacebo) during the first session. After a wash out period of 3 weeks participants are
crossed over to the other supplement group. Randomization is performed by a researcher within
our lab who is not involved in any of the experimental procedures. As such, the participants
as well as the researchers performing the measurements are unaware of the order in which
participants receive the different supplements. The randomization sequence will be generated
by a member of the research group that is not involved in the current study using an online
tool (RANDOM.ORG - Integer Set Generator). Identity concealment codes are used in the
randomization list which is in the safe custody of the same member of the research group that
is not involved in the current study.
Blinding: To avoid bias, treatment arms will be blinded to Investigators/Trial
staff/participants, as follows: the supplement will be manufactured, packaged and labelled in
such a way that the visual appearance, nor the smell and/or touch of the supplement can be
distinguished from the placebo. Supplements are delivered to the participants by a member of
the research group that is not involved in any of the experimental procedures or analyses.
Furthermore, to maintain the blind, the supplement and placebo will follow the same
administration route and process.
Unblinding: After all participants have completed the Trial and all wet-lab analyses are
completed, the database will be locked and the collected Trial data will be unblinded to
allow statistical analysis of the Trial data. The PI will provide the designated researchers
with the unblinded treatment information for all participants after completing the data
analysis.
In case of an emergency in a blinded Trial, the Investigator has the sole responsibility for
determining if premature unblinding of a participant's treatment assignment is warranted.
Participant safety must always be the first consideration in making such a determination. The
date and reason for unblinding must be recorded, as applicable.
Muscle damaging eccentric exercise (EE) protocol Participants perform a 5-minute warm up on
the cycle ergometer at 100W. Afterwards, they take place on the isokinetic dynamometer where
their upper body is strapped to the chair to stabilize their trunk and their right leg is
strapped to the lever arm. The EE protocol consists of 24 sets of 10 isokinetic unilateral
eccentric contractions of the knee extensors at an angular velocity of -30⁰/s and a range of
motion from 180⁰ to 60⁰ (where 180⁰ refers to full extension). In between sets, 60 seconds of
rest is provided. Participants are verbally encouraged to maximally resist the downward
rotation of the lever arm during the eccentric phase of the movement. No resistance is
applied during the concentric phase of the movement. This eccentric exercise protocol showed
to effectively induce muscle damage as indicated in previous studies.
Supplements Participants receive PEA and placebo supplements on two separate occasions in a
randomized order. The PEA supplements contain 350 mg of Levagen+ (315 mg of PEA and 35 mg of
excipients) and the placebo supplements contain 350 mg of maltodextrine. Levagen+ consists
for 90% of palmitoylethanolamide and for 10% of a mixture of coconut oil (fractionated),
polyglycerol polyricinoleate, citrus oil, olive oil, lecithin, dl-alpha tocopheryl acetate,
and silicon dioxide to improve the bioavailability of PEA. Both PEA and placebo supplements
are packaged in non-see-through hard capsules to assure that there are no differences in
appearance or taste. Participants are instructed to ingest their supplement twice a day (in
the morning and one hour before bedtime) for 6 consecutive days starting after collection of
baseline measures.
Physical activity and Dietary control The participants are asked to maintain their habitual
activity patterns (e.g. biking to class). However, they are instructed not to perform any
moderate to strenuous exercise during and 72h before the start of the experimental session.
Furthermore, no exercise of any kind is allowed 24h before the collection of muscle biopsies.
Participants receive a standardized meal (1200 kcal; 180 g CHO, 33 g fat, 45 g protein) on
the evening before EE as well as on the evening before muscle biopsy collection. On testing
days, when participants have to report to the lab in a fasted state, a breakfast (500 kcal)
is provided upon completion of all measurements. The same breakfast is provided 1 hour before
the start of EE. Furthermore, participants complete a food diary daily, starting after
baseline measurements, in order to check for possible dietary variation between conditions.
Dietary analyses are performed using a web-platform for dietary recording and analysis of
macronutrient and micronutrient composition (https://mijn.voedingscentrum.nl/nl/eetmeter/).
Participants are not allowed to consume alcoholic or caffeinated beverages during the
experimental sessions.
Blood samples Blood (2x10 mL) is sampled from an arm vein (Venoject system), and plasma or
serum is immediately separated by centrifuging. Plasma and serum samples are distributed in
different 1 mL aliquots and stored at -20°C until usage for biochemical assays at a later
date. Blood samples are collected at baseline, immediately before EE and 24, 48, 72, 96 and
120h following EE in a fasted state.
Plasma samples are analysed by appropriate specific Enzyme-Linked Immunosorbent Assay (ELISA)
for CK (marker for muscle damage), IL-6 and TNFalpha (both markers of systemic inflammation).
PEA concentrations are assessed by high-performance liquid chromatography with tandem mass
spectrometry. Additional assays on blood samples may be performed if judged relevant after
the primary analyses have been performed. After all analyses have been performed, any
residual sample is stored for an additional 3 years period before being permanently
destroyed.
Muscle biopsies Muscle biopsies are collected at baseline and 48h following EE from the
dominant leg in a fasted state. Hence, a total of 4 biopsies per subject are taken throughout
the study. Biopsies are taken by an experienced physician (Dr. Ruud Van Thienen) from the m.
vastus lateralis of the trained leg using a Bergström needle with a 5 mm diameter as
previously described (29). In short, a small incision (~5mm) is made through the skin and
underlying fascia under local anaesthesia (Lidocaïne, 1ml) to facilitate smooth penetration
of the needle into the muscle belly. Once the needle is placed within the muscle belly, a
tube and syringe are attached to the upper extremity of the needle which is used to create a
vacuum by which the muscle is drawn into the needle. At this point, a smooth twisting
downward movement of the inner part of the needle cuts through the tissue to obtain the
muscle sample. After cutting of the tissue (~150-300 mg of muscle) the samples are freed from
any visual blood and connective tissue before being frozen in liquid nitrogen. The incision
that was made through the skin will be closed using adhesive strips (steristrips) and
bandaged with a waterproof plaster. Participants will wear the plaster for 5 days before they
are allowed to take it off. During these 5 days participants are allowed to take a shower but
they will be instructed not to immerse or plunge the plaster (i.e. taking a bad or swim).
Participants will be warned about the fact that the muscle biopsy procedure leaves a small
scar of ~5mm that most likely will be invisible after one or two years. Furthermore, the
local anaesthesia assures that there is no pain sensation during the procedure, but
mechanical sensors in the muscles remain sensitive. As such, participants may experience the
feeling of pressure but not pain. After the procedure the muscle may feel stiff for a day or
two.
Muscle biopsies are biochemically analysed within our lab (Exercise Physiology research lab,
KU Leuven) for parameters of muscle inflammation, stress, regeneration and anabolism.
Maximal voluntary contraction (MVC) of the knee extensors The isometric, eccentric and
concentric knee extensor torque of the trained leg are measured using an isokinetic
dynamometer (cfr. (28)) at baseline and 24, 48, 72 and 120h following EE. In short,
participants are seated with their upper body strapped to the chair to stabilize the trunk
and their trained leg strapped to the lever arm. For maximal isometric voluntary contraction
peak, the knee joint is placed at an angle of 90⁰. Participants are instructed to maximally
contract their knee extensors and hold it for 3 seconds. Maximal eccentric and concentric
voluntary contraction are measured at an angular velocity of 30⁰/s and a range of motion
between 180-60⁰ (where 180⁰ refers to full extension). Each movement is performed three times
with 60 seconds of rest between attempts. The maximal peak torque value obtained between
attempts is used for analysis.
Counter movement jump Participants perform a counter movement jump (CMJ) on a force platform
(SMARTJUMP, Fusion sport, Nottingham, UK) at baseline and 24, 48, 72 and 120h following EE.
Participants are asked to take an upright position (knee angle of 180⁰) with their feet on
hip width before they initiate a downward movement to squat position (knee angle of 90⁰)
after which they jump vertically for maximum height. Participants are instructed to place
their hands on their hips and to keep their body erect throughout the jump. CMJ is performed
three times with 60 seconds of rest between attempts. The highest jump recorded is used for
analysis.
Squat jump Participants perform a squat jump (SJ) on a force platform at baseline and 24, 48,
72 and 120h following EE. Participants are asked to start at a squat position (knee angle of
90⁰) and hold this for 3 seconds before they jump vertically for maximum height. Participants
are instructed to place their hands on their hips and to keep their body erect throughout the
jump. SJ is performed three times with 60 seconds rest between attempts. The highest jump
recorded is used for analysis.
Muscle soreness Muscle soreness of the trained leg is assessed at baseline, immediately
before EE and 24, 48, 72, 96 and 120h following EE using a visual analogue scale (VAS).
Participants are asked to hold a squat position for 3 seconds with their hands on their hips
and to mark their perceived muscle soreness on a scale that is represented by a 100mm line of
which the far-left represents 'no pain' while the far-right represents 'extreme pain'.
Sleep quality and quantity At baseline, participants' habitual sleep hygiene is evaluated
using the Pittsburgh Sleep Quality Index . In addition, participants complete the St. Mary's
Hospital questionnaire daily to evaluate their sleep quality and quantity from the previous
night, starting on the night before baseline measurements. The St. Mary's Hospital
questionnaire showed to be a reliable tool for assessing the previous night's sleep of a
subject.
