Clinical Trial Details
— Status: Not yet recruiting
Administrative data
| NCT number |
NCT00479960 |
| Other study ID # |
022006 |
| Secondary ID |
|
| Status |
Not yet recruiting |
| Phase |
Phase 0
|
| First received |
May 28, 2007 |
| Last updated |
May 29, 2007 |
| Start date |
June 2007 |
| Est. completion date |
June 2007 |
Study information
| Verified date |
November 2006 |
| Source |
Hadassah Medical Organization |
| Contact |
Abraham Benchetrit, MD |
| Phone |
97250-8962566 |
| Email |
benchet[@]012.net.il |
| Is FDA regulated |
No |
| Health authority |
Israel: Ethics Commission |
| Study type |
Interventional
|
Clinical Trial Summary
The phospholipids of mammalian spermatozoa possess a distinctive fatty acid composition with
high proportion of long chain polyunsaturated fatty acids. The lipid composition is a major
determinant of the membrane flexibility and sperm motility required for proper
fertilization. It also influences the sperm plasma membrane's fluidity, chilling sensitivity
and thermotropic lipid phase transition (LPT) and these parameters determine our ability to
cryopreserve these cells. Our hypotheses is that by providing dietary supplementation of
omega-3 polyunsaturated fatty acids an improvement of sperm parameters (number, motility,
viability) can be achieved. We also expect alteration spermatozoal plasma membrane fatty
acid composition, making it more chilling resistant.
Experimental methodology: 1) Characterize fatty acid composition of the spermatozoa of
normal and abnormal spermatozoa by gas chromatography. 2) Characterize sperm plasma membrane
LPT by FTIR spectrometer. 3) Run a randomized double-blind, placebo controlled, crossed-over
dietary fatty acid supplementation pilot trial in human sub-fertile patients. Large scale
trial will follow, if justified. In both trials sperm characterization of each participant
will be conducted before, during and following the trial.
Subfertile males will benefit greatly if their sperm parameters can be improved and
cryopreserved while ensuring enhanced post-thawing survival. We believe that changing the
fatty acid composition of sperm plasma membrane by simple dietary means will open the way to
improve the fertility of those that needs it the most.
Description:
The phospholipids of mammalian spermatozoa possess a distinctive and highly unusual fatty
acid composition, the most unique feature of which is a very high proportion of long chain
(C20-22) highly polyunsaturated fatty acyl groups. In most mammals, docosahexaenoic acid
(DHA; 22:6,n-3) is the dominant polyunsaturated fatty acid (PUFA) although, in several
species, docosapentaenoic acid (22:5,n-6) is also a major component 2. The lipid composition
is a major determinant of the membrane flexibility required for the characteristic flagella
movement of spermatozoa and for the fusogenic properties of the membranes associated with
the acrosome reaction and fertilization. In addition, it has been demonstrated that
lipid-derived mediators are involved in various signal transduction mechanisms that regulate
spermatozoa functions.
Sperm motility was therefore closely correlated with fertility, yet factors controlling
sperm motility in humans are not fully understood 12. The amount of PUFA, mainly DHA, and to
a lesser extent eicosapentaenoic acid (EPA), was showed to be positively associated with
sperm motility and essential for optimal fertility. The reduction in the amount of DHA in
sperm lipids have been correlated with reduced sperm concentration, sperm progressive
motility and the proportion of cells with normal morphology. In asthenozoospermic (poor
sperm forward motility), the levels of DHA (and total PUFA) was found to be lower than in
normozoospermic individuals despite similar serum fatty acid composition 18. Broiler
spermatozoa rich in EPA or DHA or both showed significantly increased fertility following
artificial insemination (AI).
Chilling injury to mammalian gametes has been described as the irreversible damage that
occurs upon cooling to low, but non-freezing, temperatures. Several studies suggested that
chilling injury is the major limiting factor for successful gamete cryopreservation. Other
studies suggested that membranes are the primary sites for structural and functional
chilling injury in sperm and oocytes. Additionally, membrane chilling injury is dependent on
the biochemical and biophysical properties of membranes. The plasma membrane thermotropic
lipid phase transition (Tm) from the fluid liquid crystalline phase to the more rigid gel
phase is associated with chilling injury. When membrane lipids undergo lipid phase
transition (LPT), fluidity decreases and the structure and functioning of the membrane
change. As a result, the membrane permeability increases and the cells are damaged. The
temperature at which Tm occurs is greatly affected by the fatty acid composition of the
membrane. Membranes consisted of lipids with multiple unsaturations or short carbon chains
are more fluid at low temperatures. The ability of the spermatozoal plasma membrane to
resist structural damage during cryopreservation may therefore be related to its fatty acids
composition and the strength of the bonds between membrane components 38. It was shown, for
example, that spermatozoa of Asian elephants, which are more chilling sensitive than African
elephant sperm, contain lower concentration of PUFA in their membranes and are especially
poor in DHA.
There are several ways to change the chilling sensitivity and Tm in mammalian gametes.
Spermatozoa can spontaneously interact with liposomes as was described in several studies.
Previous work suggested that phospholipids adsorbed on to the sperm cells, which were then
cooled to 4oC, increased the chilling resistance of the sperm cells from bovine, ram and
elephant.
The relationship between dietary components and reproduction is well established. Follicular
maturation, ovulation and oestrus in the female, and libido and fertility in the male, might
all be adversely affected by undernutrition. Men with infertile semen were found to consume
less omega-3 fatty acids than fertile men, and a significant correlation was established
between the consumption of alpha-linolenic acid (18:3n-3) on the one hand and sperm
concentration and progressive motility on the other hand. In boar, the output of the
accessory glands, but not the output of spermatozoa by the testes, was affected by the
reduced feed intake and is likely to reflect a suppression of androgen secretion. Various
reports showed that enrichment of diet with PUFA can alter the biophysical properties or
reproductive performance of mammalian and avian sperm and oocytes.
Objectives and expected significance of the research Male patients arriving at the male
fertility clinic frequently suffer from one or more of the following: asthenozoospermia (low
progressive motility), teratozoopermia (abnormal morphology) and/or oligozoospermia (low
sperm count), collectively known as O.T.A It is assumed that by supplementing the diet of
such patients with omega-3 PUFA, an improvement of these parameters can be achieved. Since
the number of normal cells in all such patients is limited, omega-3 PUFA supplementation may
confer the needed change to the sperm plasma membrane, making these cells more resistant to
chilling injury during preservation. This will provide us with a mean to ascertain that more
of the normal cells will survive the freezing and thawing processes involved in
cryopreservation. By achieving that we will be able to make sure that enough normal cells
will be available for fertilization.
If the hypothesis of this study is proved correct, it will mean that by a simple dietary
mean we will be able to improve reproduction in those that needs it the most.
Comprehensive description of the methodology and plan of operation
1. Experimental design: The experiment will be a randomized double-blind, placebo
controlled, crossed-over trial. The experiment will start with a pilot which will be
conducted on 20 participants, all of which are out-patients at the male fertility
clinic of the Shaare Zedek Medical Center, Jerusalem, Israel. The pilot is expected to
be followed by a large scale trial in a similar design (without the "cross-over" part).
Both pilot and large scale trial will seek and obtain the approval of the Helsinki
committee of the Shaare Zedek Medical Center prior to their initiation. All
participants will be asked to sign an informed consent.
2. Basic spermatozoa database: For general information on human spermatozoa, fresh and
frozen samples of normal and subfertile males will be analyzed for total lipids fatty
acids composition by gas chromatography. All samples will be evaluated for basic sperm
quality characteristics according to the World Health Organization (WHO) guidelines 66.
Samples will then be separated from the seminal plasma and stored till fatty acid
composition analysis. Determination of the lipid phase transition temperatures by the
FTIR spectrometer on fresh samples will also be conducted.
3. Dietary supplementation: At the beginning of the trial each participant will be
provided with 120 capsules of either placebo or commercially available omega-3 fatty
acids. The capsules will be consumed in the following manner: 2 capsules daily during
the first week, 3 capsules per day during the second week and starting on the third
week, 4 capsules per day till all are consumed. At this point there will be a break of
4 weeks before switching between the groups and resuming the same protocol once again.
During the entire period each participant will also be provided with vitamin E pills as
anti-oxidant.
4. During and post-experimental sperm characterization: Each of the participating patients
will provide 5 samples during the dietary supplementation pilot experiment: Once at the
beginning of the trial, once in the middle of each of the two segments (see dietary
supplementation paragraph for details), once at the cross-over point and once at the
end of the trial. All samples will be analyzed in the same way as described above.
5. Lipid phase transition determination: Approximately 10 of the participants in the pilot
trial will be requested to provide additional samples at the cross over point and at
the end of the trial for the determination of the lipid phase transition temperatures
by the FTIR spectrometer. These participants will be selected based on their sperm
fatty acid composition at the beginning of the trial so as to cover as wide range of
compositions as possible.
6. Large scale trial: After analyzing the results of the pilot trial, a decision will be
made if there is indication to proceed with a large scale trial. Assuming the results
will indicate such a need, a large scale trial will be conducted. Experimental design
will be randomized double-blind, placebo-controlled trial. The number of participating
patients will be determined at the time the decision to proceed with such a trial will
be made. At that time a decision will also be made what other male fertility clinics
from additional medical centers to invite to participate.
