Human Reproduction Vol. 15, (Suppl. 5) pp. 98-110, 2000
The future of male infertility management and assisted
reproduction technology
David Mortimer
Genesis Fertility Centre, #550, 555 West 12th Avenue, Vancouver, BC V5Z 3X7, Canada
E-mail: [email protected]
Intracytoplasmic sperm injection (ICSI) is
undoubtedly a powerful, and sometimes the
only effective, form of infertility treatment.
Nonetheless, it is a non-specific treatment that,
combined with increasingly heroic techniques
to recover male germinal cells, has led to perceptions of men as just providers of gametes in the
infertility equation. In response to this nihilist
attitude, where women are investigated extensively and scant attention is paid to men, there
is a re-emerging awareness of andrology—particularly in countries with limited healthcare
resources. Structured management strategies,
using diagnostic information to recognize causative factors amenable to simpler, even systemic,
therapies with reasonable chances of pregnancy
rather than resorting prematurely to assisted
reproduction technology, represent rational,
cost-effective approaches to infertility management. Furthermore, genetic testing (particularly
cystic fibrosis gene defects and Y-chromosome
microdeletions) is essential for couples to make
fully informed decisions on their options. Recognition that free radical-induced damage to the
sperm genome (e.g. from smoking or in-vitro
sperm manipulation) underlies deleterious
paternal effects on preimplantation development promotes further synergy between
andrology and embryology. Although societies
strike different balances between considerations
of affordability and cost-effectiveness of assisted
reproduction technology, ICSI represents a last
resort, to be used when less-invasive, lower-cost
98
treatments have been deemed inappropriate or
have failed. Consequently, rather than assisted
reproduction technology eliminating the need
for andrology, the future will see increasingly
tighter integration of multidisciplinary infertility care, embracing careful diagnosis and patient
education before obtaining truly informed consent and embarking upon cost-effective
treatment.
Key words: andrology/assisted reproduction/costeffective treatment/infertility management/male
infertility
Introduction
Infertility is a problem manifest by couples, not
individuals. A given man or woman might be
sterile, or have a form of reproductive dysfunction
that causes them to have reduced fertility potential
('be infertile'), but such attributes can only be
expressed when the individual is part of a reproductive unit, i.e. a couple attempting to achieve a
pregnancy. When considering the contribution of
the male partner of a couple to the aetiology of
their infertility, the impact of a given problem such
as a low sperm count will depend upon not only
the severity of the problem per se but also upon
the female partner's relative fertility potential.
Hence 'male infertility' is at best a misnomer,
although we might consider the existence of 'male
factor infertility'—but not to the exclusion of
possible (likely?) coexistent female factor infertil© European Society of Human Reproduction and Embryology
Male infertility treatment
ity in a given couple. These principles have been
discussed, and their application described, many
times (e.g. Comhaire et al., 1987; Cummins and
Jequier, 1994; Hargreave, 1994a; Rowe et al.,
1994; Irvine, 1998; Kamischke and Nieschlag,
1998, 1999a) and whereas in the past failures to
apply them were more often due to patriarchal
social beliefs, the development of assisted reproduction technology now constitutes a frequent
reason for their abandonment. This review considers recent developments in the relationship
between andrology and the 'modern' application
of clinical assisted reproduction technology.
It is certainly true that the development of our
understanding of the regulation of the male and
female reproductive processes advances asynchronously. While great progress has been made
in elucidating the neuroendocrine regulation of
gametogenesis in the female, spermatogenesis
remains even today much less well understood.
The existence of multiple spermatogenic waves
occurring simultaneously, even within a single
seminiferous tubule, represents a far more complex
system to unravel than the 28-30 day ovarian cycle.
Consequently, for the male partner of an infertile
couple we still have little more in our laboratory
armamentarium than the traditional descriptive
semen analysis and a range of poorly standardized,
and often laborious or technically complex, sperm
function tests. Whereas specific endocrine treatments exist for a variety of dysfunctions of the
ovarian cycle, there is no proven successful treatment for defective spermatogenesis beyond the
extreme case of hypogonadotrophic hypogonadism
(Wu, 1994). Sperm quality is rarely investigated
in any depth, being considered only in terms
of visual appraisal of sperm motility and sperm
morphology using light microscopy, and problems
of gamete approximation (insemination, sperm
transport through the female tract, sperm storage
and longevity in vivo) are almost entirely ignored.
Hence sperm transport, as well as various aspects
of sperm-oocyte interaction, anti-sperm antibodies
(ASAB) are often ignored because of confusion as
to their detection and the interpretation of test
results, despite extensive evidence that they can
affect sperm-cervical mucus interaction (Bronson,
1999; Hjort, 1999). Indeed, ASAB are an excellent
example of how a pathological process impacts
upon fertility: their effects are specific at the
cellular level, but their expression is variable at
the organism level. Therefore, rather than taking
exceptions as evidence to disprove a rule (e.g.
'I've had patient with ASAB who impregnated his
wife'), they should be recognized as the extreme
of a distribution of effects arising as a complex
product of the severity (and intrinsic variability)
of a pathological process in the man and the
relative fertility of his partner. Indeed, such understanding is central to how clinical medicine and
infertility care should be practised.
Infertility management and assisted
reproduction technology
The possible causes of, and approaches for the
treatment of, male factor infertility are summarized
in Table I. Clearly certain problems are amenable
to surgical intervention, and sexual dysfunction is
frequently remedied by medical or psychological
therapy (e.g. Hargreave, 1994b; Kamischke and
Nieschlag, 1999b; Shah, 1999). Exposure to reproductive toxins is finally being recognized as an
environmental, lifestyle or workplace hazard, the
impact of which can be alleviated by avoidance
(e.g. de Jager and Bornman, 1999). However, the
fundamental problems of making more, or better,
spermatozoa remain devoid of effective treatment
(e.g. Hargreave, 1994b; Kamischke and Nieschlag,
1998, 1999a). Consequently, the majority of therapeutic modalities for male factor infertility have
been founded upon the simple principle of
bypassing the problem(s):
• pericervical artificial insemination of homologous spermatozoa (AIH) for problems of insemination;
• various forms of epididymal sperm aspiration
to obtain spermatozoa in cases of agenesis or
inoperable blockage of the excurrent ducts;
• intrauterine insemination (IUI) to bypass problems of sperm transport at the cervical level;
• gamete intra-Fallopian transfer (GIFT) or IVF
to bypass more severe problems of sperm transport or less serious problems of poor sperm
quantity or quality;
• IVF to compensate for more serious problems
of poor sperm quantity or quality;
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CMortimer
Table I. Causes of male factor infertility and initial approaches for treatment
Cause of infertility
Category
Sterility
Absolute
Functional
Subfertility
Sperm production
Extrinsic factors
Sperm delivery
Treatment approach
Problem
Medical
ART
Spermatogenic arrest
Sterilizing defect
Tract agenesis
Tract dysgenesis
Tract blockage
Sperm dysfunction
Endocrine?
None
None
Surgical?
Surgical?
None
TESE?
ICSI?
MESA/TESE
MESA/TESE
(MESA/TESE)
ICSI
Too few spermatozoa
Abnormal spermatozoa
Antisperm antibodies
Free radicals (ROS)
Toxicological exposure
Partial obstruction
Ejaculatory disorders
Impotence
?
7
(Immunosuppression)
Antioxidants?
Reduce exposure
Surgical
Medical or surgical
Psychological and/or
pharmacological
IVF or ICSI
IVF? or ICSI
ICSI
IVF
ART = assisted reproduction technology; TESE = testicular sperm extraction; ICSI = intracytoplasmic sperm injection;
MESA = microepididymal sperm aspiration; ROS = reactive oxygen species.
• sub-zonal sperm insertion (SUZI) to bypass
sperm transport and penetration of the oocytecumulus complex (although an acrosome reaction was still required—but could be induced
artificially — and sperm-oocyte fusion had to
occur); and
• intracytoplasmic sperm injection (ICSI) as the
ultimate solution that bypassed all considerations of sperm physiology and function by
delivering the male partner's haploid genome
contribution directly into the oocyte.
Most recently (i.e. during the second half of the
1990s) we have gone even further and, in cases
where no mature spermatozoa were available even
from the epididymis, spermatozoa are sought from
within the testis. For such men, who are otherwise
medically sterile, various workers have reported
ICSI treatment using testicular spermatozoa
[extracted from the lumen of the seminiferous
tubule post-spermiation (TESE)] or more immature
spermiogenic stages such as elongated or elongating spermatids (ELSI) and even round spermatids
(ROSI) recovered or extracted from the Sertoli
cells, and ICSI has even been attempted using
isolated round spermatid nuclei (ROSNI). However, the competence of these progressively earlier
germinal cells as gametes is compromised both
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when assessed using in-vitro sperm function testing
(Aslam and Fishel, 1999) and when used clinically
(Silber and Johnson, 1998; Al-Hasani et al, 1999;
Aslam et al, 1999; Ghazzawi et al, 1999; Prapas
et al, 1999; Vanderzwalmen et al, 1999; also see
Table II), raising doubt as to the real value of such
heroic feats of sperm retrieval, and giving concern
that the man is now seen as little more than a
source of (any) gametes in the infertility equation.
In a traditional specialist infertility centre the
female and male partners of each couple receive
a careful and thorough investigation, including
comprehensive medical histories, physical examinations, and a work-up using a range of medical
and surgical diagnostic tests. The basic standards
for such infertility care were formalized by the
World Health Organization in its publication the
WHO Manual for the Standardized Investigation
and Diagnosis of the Infertile Couple (Rowe et al,
1994) which constitutes the de-facto minimum
standards worldwide. Because of the asynchronous
development of treatment options for male factor
infertility compared to female factors, a revised
edition considering advances for managing the
infertile male has been prepared recently (Rowe
et al, 2000). However, many infertile couples now
present (or are referred by their family doctor)
Male infertility treatment
Table II. Spermatids as gametes. Outcomes from intracytoplasmic sperm injection (ICSI) using either elongated spermatids
(ELSI), round spermatids (ROSI) or round spermatid nuclei (ROSNI) according to a recent literature survey by Aslam et al.
(1999)
Outcome measure
ELSI
ROSI
Treatment cycles (with embryo transfer)
Fertilization (2 pronuclei)
Embryos per transfer
Implantation rate per embryo transferred (%)
Clinical pregnancies
Live births
Deliveries per embryo (%)
55 (55)
57.5%
2.1
6.1
12
11
5.6
80 (75)
31.3%
3.6
6.5
10
9
5.8
ROSNI
9(9)
64.0%
<3.3
0
0
0
0.0
Note that the implantation rates per embryo transferred are substantially lower than those achieved by modern IVF/ICSI units
(-20-25%).
more-or-less directly to an 'IVF Clinic' where,
typically, the female partner might have already
undergone, or be expected to undergo, a WHOcompatible comprehensive work-up, including history and physical, extensive endocrine testing,
ultrasonographic examination of the ovaries, hysterosalpingograhy and/or laparoscopy (with tubal
patency test) and perhaps hysteroscopy. In contrast,
the male partner will typically receive little
attention beyond a perfunctory history and semen
analysis. Only rarely are the male partners of
assisted reproduction technology couples examined
physically or undergo such procedures as scrotal
thermography, ultrasound and vasography comparable to their partners' investigations. While this
situation might be explained as due to the limited
availability of specialist clinical andrologists, or
to poor perceptions of treatment availability for
problems that might be identified in the male
partner, the need for and value of extensive workup of the female partner should also be reconsidered
if treatment is pre-ordained to be IVF-based.
Another problem is the widespread consideration
of poor quality semen as a diagnosis. For the
male partner of an infertile couple a finding of
'oligoasthenoteratozoospermia' at semen analysis
is not a diagnosis but rather a symptom of some
unknown problem(s). Basing treatment upon such
a non-specific symptom can only be considered
arbitrary (even presuming, for the sake of argument,
that the semen analysis laboratory was run to an
adequate standard to minimize technical error and
was therefore able to provide accurate results, see
below). Of even greater concern is the trend in
some centres to use ICSI as the first-line treatment,
often regardless of the aetiology of a couple's
infertility.
Consequences of ICSI
In some centres, and indeed in some regions or
countries, there is a growing tendency for ICSI to
be used as the first choice of treatment for infertile
couples. In some circumstances this has arisen
in response to simplistic management by health
insurance providers, in others as a consequence of
misplaced concerns for avoiding treatment failure
by using a technique that is presumed to solve all
problems, and in others because ICSI is seen as
the most successful form of treatment. Overall, this
has been considered 'insidiously lazy' (Cummins,
1999) and a dangerous loss of control over the
clinical decision-making process (Cummins and
Jequier, 1994; Jequier and Cummins, 1997). However, consider those couples whose infertility is
amenable to treatment using, for example, interuterine insemination (IUI). A success rate of
-20% per treatment cycle can be achieved by
IUI combined with mild ovarian stimulation (e.g.
Branigan et al., 1999) giving, over three sequential
cycles, a cumulative pregnancy rate of -50%. This
is greater than that achieved in many centres using
IVF or ICSI, of which only a single cycle would
typically be performed in a given 3 month period.
Furthermore, the cost of three IUI cycles is less
than one of IVF or ICSI, and the risk of sideeffects lower (provided that appropriate care is
taken to control the stimulation—and limit the
number of IVF or ICSI embryos transferred—to
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D.Mortimer
minimize the risks of multiple pregnancy). A
similar comparison can be made for couples where
the man's spermatozoa are perfectly capable of
achieving fertilization in vitro and so for whom
IVF will have the same success rate as ICSI, but
typically at a lower cost per treatment cycle.
Further arguments can be made to undermine the
perceived greater success of IVF compared to IUI,
or ICSI compared to IVF (or IUI) when allowance
is made for the expected better prognosis of those
patients with less severe problems: IUI-suitable
patients treated using IVF will certainly show better
fertilization rates (and perhaps embryo quality, see
below) than patients who were unsuitable for IUI
due to decreased sperm quality, and their inclusion
in the IVF population will create an overall increase
in the outcome of the population treated using IVF.
This same argument was used to explain the higher
success rate of GIFT, and the often poorer IVF
success rates seen in centres that treated a significant proportion of their patients using GIFT.
Beyond these perceptions of improved success
rates using ICSI as the first choice option, there is
a range of more critical concerns (e.g. Cummins,
1999; Lamb, 1999), including reports that ICSI
in rhesus monkeys produces offspring with an
increased prevalence of chromosomal abnormalities (Hewitson et al, 1999a) and can facilitate
transmission of foreign DNA into embryos (Chan
et al, 2000). ICSI as the panacea for (male)
infertility leads to many couples having an incomplete, or no, diagnosis of the cause of their infertility
and significant factors can go unnoticed, for
example systemically induced sperm chromatin
damage caused by infections of the male genital
tract (e.g. Ochsendorf, 1999), by smoking (e.g.
Fraga et al, 1994), or in cases with oligoasthenozoospermia (e.g. Aitken et al, 1998). Furthermore,
the tendency to use therapeutic 'overkill', and
failing to use appropriate medical or surgical treatments, especially if more cost-effective, might
be considered poor practice. From the woman's
perspective, unnecessary ovarian stimulation has
potentially serious side-effects, not to mention
the risks associated with ovarian hyperstimulation
syndrome.
Preferential use of ICSI to avoid possible poor
or failed fertilization covers two failings: (i) failure
102
to identify specific causes, or even the risk, of
sperm dysfunction during diagnostic work-up by
the physician responsible for the couple, or (ii) a
tendency to tolerate lower technical standards
within the laboratory that themselves lead to poor/
no fertilization at IVF, but which are obviated by
using ICSI. Accepting either situation must be
considered unprofessional at best.
Finally, there are genetic sequelae of treating
many cases by ICSI, a topic that is the subject of
heated debate (e.g. Tournaye et al., 1997; Barratt
et al, 1999; Cummins, 1999; Lamb, 1999;
Schlegel, 1999). However, this does not imply that
ICSI per se causes genetic abnormalities of the
embryos and resulting offspring (Bonduelle et al.,
1999), nor does it concern the increased prevalence
of sex chromosome aneuploidy in men with severe
oligozoospermia (e.g. Pfeffer et al, 1999), but
rather recognizes a series of genetic, and hence
heritable, disorders:
• The high risk of Y chromosome microdeletions
in men with severe oligozoospermia or azoospermia (Patrizio, 1997; Vogt, 1999), which will be
transmitted to any sons created using assisted
reproduction technology and who will, in their
turn, be sterile (Kleiman et al, 1999; Page
et al, 1999).
• The risk of transmitting cystic fibrosis to the
offspring in couples where the man has congenital bilateral absence of the vasa deferentia
(CBAVD), which is due to mutations of the
cystic fibrosis transmembrane conductance regulator gene (Schlegel and Shin, 1999), and where
the female partner is a AF508 carrier.
• The growing number of single gene defects that
cause male infertility or sterility and can be
inherited by sons produced using assisted reproduction technology (e.g. Vogt, 1997, 1999;
Cummins, 1999).
In the most extreme cases, assisted reproduction
technology can now be considered to have made
male sterility a heritable condition (Tournaye et al,
1997; Barratt et al, 1999). While couples who
have been fully informed of their diagnosis and
likely outcome often chose still to proceed with
treatment, failing to inform them of all pertinent
information can be considered to negate their
ability to provide informed consent to treatment
(e.g. Cummins, 1999).
Male infertility treatment
Role of diagnostic laboratory andrology
The semen analysis remains the initial laboratory
investigation for all male partners of infertile
couples. After decades of effort to improve the
value and reliability of this fundamental test, we are
finally seeing significant improvements in semen
analysis standards in many countries as a result of
greater adherence to established methods (e.g.
Mortimer, 1994a,b; Mortimer and Mortimer, 1999;
World Health Organization, 1999), the development of coherent training courses by organizations
such as the British Andrology Society, the
European Society for Human Reproduction and
Embryology and the Nordic Association for
Andrology (Mortimer, 1994c; Bjorndahl and Kvist,
1998; Punjabi and Spiessens, 1998; Vreeburg and
Weber, 1998), recognition of the value of internal
quality control (e.g. Cooper et al., 1992; Clements
et al., 1995), and the appearance of external quality
assurance programmes (e.g. Neuwinger et al.,
1990; Matson, 1995).
Laboratory investigation of the male partners of
infertile couples can be viewed in two opposing
perspectives: (i) trying to predict fertility, and
(ii) trying to identify pathology that will cause
infertility or sterility. Although the former, prognostically motivated attitude is by far the more
common, in reality the second is more useful—
and readily applicable. There are far too many
confounding factors that prevent reliable prognosis,
short of stating that achieving a pregnancy (or
fertilization in vitro) is, for the majority of men,
the most likely outcome and can therefore be
expected to correlate with most measurements
made during diagnostic tests. The most useful
information is obtained when a test gives an
abnormal result that is predictive of failure—but
not simply failure of the overall process (i.e.
conception), rather failure of a specific component
or step in the process that can be used to identify
a treatment strategy. In this way the information
is not used to generate an overall prognosis for
the couple, but to direct the management of their
treatment. These concepts, which have been
described as 'structured management', are discussed in detail elsewhere (Mortimer, 1999) and have
been considered in the WHO's updated recommendations for management of male factor infertility (Rowe et al, 2000).
Functional Life of the Spermatozoon
Spermatogenesis
Spermiogenesis
I
S
Spermiation
CAPUT
CORPUS
-OYCAUDA
-M1S
Maturation: chromatin
plasmalemma
cytoplasmic droplet
biochemical
Storage
EJACULATION
VAGINA
CERVIX
UTERUS
0
tSTH-
Capacitation
Sperm reservoir
-MUS
L.
Capacitation & hyperactivation
Penetration of cumulus & corona
Binding to zona pellucida
Induction of acrosome reaction
Penetration of zona pellucida
Binding to oolemma
Incorporation into oocyte
Pronucleus formation
A
SYNGAMY
y
A
1
M
D
P
U
U
I.
C
T
Figure 1. Diagrammatic representation of the functional life
of the human spermatozoon showing the various processes
occurring within the reproductive tracts of the male and
female partners.
A 'modern' view of sperm testing in relation to
the functional life of the spermatozoon (Figure 1)
is shown in Table III. In these terms a semen
analysis is not simply descriptive, using a series
of counts to determine the adequacy of sperm
production in abstract terms, but rather it is performed in an attempt to identify the likelihood of
the man being able to achieve colonization of his
partner's cervical mucus by his spermatozoa and
hence have at least a chance of establishing a
pregnancy in vivo. It is possible that in-vitro
sperm-mucus interaction tests can help achieve
this determination more directly, but their performance is confounded by access to the partner's
cervical mucus — although other tests of sperm
function such as the hyaluronate migration test
(Mortimer et al., 1990; Neuwinger et al., 1991;
103
D.Mortimer
Table III. Laboratory assessment of the infertile male
Type of testing
Aspect being assessed
Tests available
Semen analysis
Sperm production, quantitative
Sperm production, qualitative
Sperm maturation
Autoimmunity to spermatozoa
Semen analysis: concentration, total count
Semen analysis: motility, vitality, morphology
Sperm morphology (creatine kinase?)
Antisperm antibody tests: Immunobead test,
SpermMAR test (Friberg, Isojima or Kibrick tests)
ROS analysis: differential between WBCS and
spermatozoa
SCSA, nick translation, chromomycin A3 staining,
Comet assay, TUNEL assay
'Trial wash' (i.e. trial sperm preparation) with
post-preparation assessment
Sperm-mucus interaction (e.g. Kremer test)
Hyaluronate migration test
CASA ± pharmacological sperm stimulation
ARIC test (uses A23187; rhZP3 in future?)
Hemizona assay using salt-stored zonae
(artificial zonae using rhZP3 in future?)
(Zona-free hamster egg penetration test)
First IVF treatment cycle?
Karyotype (high resolution G-banding)
(FISH for more specific analyses?)
PCR for CFTR gene defects
PCR for AZFa,b,c sequence-tagged sites
Molecular genetics (PCR, sequencing, etc.)
Free radicals
Sperm DNA damage
Suitability for ART
Sperm function tests
Migration
Hyperactivation
Acrosome reaction
Zona binding
Fertilizing ability
Genetic testing
Numerical or structural anomalies
CF testing in men with CBAVD
Y-chromosome microdeletions
Single gene defects
MAR = mixed agglutination reaction; ROS = reactive oxygen species; WBC = white blood cell; SCSA = sperm chromatin
structure assay; TUNEL = TDt (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling;
CASA = computer-aided sperm analysis; ARIC = acrosome reaction after ionophore challenge; FISH = fluorscent in-situ
hybridization; PCR = polymerase chain reaction; CFTR = cyctic fibrosis transmembrane regulator.
Aitken et al, 1992) could resolve this difficulty.
More specific evaluations of sperm fertilizing ability have been described in detail elsewhere
(reviews: Mortimer, 1994a; ESHRE Andrology
Special Interest Group, 1996; Oehninger et al,
1997, 1998; De Jonge, 1999; Huszar et al, 1999)
and the most commonly employed tests include:
• Sperm hyperactivation as a concomitant of capacitation and functional requirement for sperm
penetration of the cumulus complex and of the
zona pellucida (analysed using computer-aided
sperm analysis or 'CASA'; see Mortimer, 1997).
Analysis of human sperm hyperactivation has
been hampered by its multiphasic and temporally
variable expression, with substantial interindividual variability, although an assay
employing a single time point and stimulation
of hyperactivation expression by progesterone
and pentoxifylline has been reported (Mortimer
et al, 1997).
• The ability to undergo the acrosome reaction in
104
response to appropriate agonists such as the
ionophore A23187 as used in the 'acrosome reaction after ionophore challenge' or 'ARIC test
(Cummins et al, 1991; ESHRE Andrology
Special Interest Group, 1996; De Jonge, 1999)
(although use of a powerful ionophore is seen by
many to limit the value of the ARIC test because
the fertilizing spermatozoon, both in vivo and
in vitro, undergoes its acrosome reaction on the
surface of the zona pellucida in response to binding to the sperm receptor of the zona, ZP3.)
• Sperm binding to the zona pellucida (e.g. the
hemizona assay or 'HZA'; see ESHRE
Andrology Special Interest Group, 1996;
Oehninger et al, 1997, 1998). However, the
dependence of this test upon the availability of
human zonae pellucidae does limit its widespread
application.
It should be noted that the latter two tests will
become more amenable to routine clinical application with the availability of bioactive recombinant
Male infertility treatment
What the Human Sperm Contributes at Fertilization
NUCLEUS : paternal haploid genome (imprinted)
PROXIMAL
CENTRIOLE
paternal centrosome
CYTOPLASM :
oocyte activating factor(s)
Figure 2. Illustration of the components of the human spermatozoon contributed to the zygote at fertilization.
human ZP3 (rhZP3:
Chapman, 1998).
Van
Duin,
1994;
What does the spermatozoon contribute at
fertilization?
A further dimension to sperm assessment is
recognized when considering what the human spermatozoon contributes to the embryo at fertilization
(see Figure 2). While the chromatin contained in the
highly condensed sperm nucleus is unanimously
recognized as being the 'payload' of the spermatozoon, it does not influence embryonic development
until around day 3 when the embryo activates its
own genome; development prior to that stage being
controlled by mRNA stored in the oocyte. But the
spermatozoon contributes two more essential components at fertilization: (i) yet-to-be-defined cytoplasmic oocyte activation factor(s) (Dale et al,
1999; Perry etal., 1999); and (ii) the proximal centriole that becomes the centrosome of the zygote and
orchestrates its first cleavage division (e.g. Hewitson
etal., 1999b; Moomjy etal., 1999). It is now known
that sperm mitochondria are tagged with ubiquitin
(most probably during spermiogenesis) and thereby
marked for destruction in the zygote (Sutovsky et al,
1999) and, although sperm mitochondria (or their
DNA content) can be tracked during the first few
cleavage divisions using fluorescent probes or PCR
(e.g. Rinaudo et al, 1999; St. John et al, 1999), it
seems most probable that they are not destined to be
contributed to the next generation (Cummins, 1997,
1998; Cummins etal, 1997; Shoubridge, 1999).
Although a deleterious effect of free oxygen upon
human spermatozoa was originally described by
John MacLeod in the 1940s, it was not until the
1980s that the effects of free radicals (or reactive
oxygen species: ROS) upon sperm function, and
their role in the aetiology of male infertility, were
well established (Aitken and Clarkson, 1987).
Shortly thereafter, the detrimental influence of ROS
generation during in-vitro sperm preparation was
discovered (Aitken and Clarkson, 1988) and widespread evidence for how this phenomenon could
impact upon sperm function tests and IVF was
collated (Mortimer, 1991). ROS are generated by
both leukocytes present in semen and the spermatozoa themselves (ICrausz et al., 1992; Aitken,
1995; Whittington and Ford, 1999). However, only
those spermatozoa with excess retained spermatid
cytoplasm generate ROS (Aitken et al, 1994;
Aitken, 1995; Huszar et al, 1998, 1999), again
emphasizing an increased risk of dysfunction in
semen from men with defective sperm production.
It is now accepted that ROS not only affect the
sperm plasma membrane by causing phospholipid
peroxidation, and hence decrease membrane fluidity
and induce a loss of sperm function, but that they
also affect the sperm DNA, causing strand breaks
that can be revealed by various tests of sperm DNA
integrity such as nick translation (Sakkas et al,
1997) and the sperm chromatin structure assay
(SCSA: Evenson, 1999; Evenson etal, 1999).
Of particular importance in the practice of
assisted reproduction technology is a recent report
105
D.Mortimer
that sperm preparation using colloidal silica-based
density gradient separation (Percoll; Pharmacia
Biotech, Uppsala, Sweden; and PureSperm;
Nidacon International AB, Goteborg, Sweden) permits the recovery of selected populations of 'better'
spermatozoa with a lower incidence of DNA nicks
(Sakkas et al., 2000). Parallel swim-up migration
from semen did not achieve comparable selection
of the genetically less compromised cells, although
other studies using the SCSA (which assesses the
susceptibility of sperm DNA to in-vitro pH- or
temperature-induced damage) have shown that
swim-up (Spano et al., 1999) and glass wool
filtration (Larson et al., 1999) can also select
spermatozoa with better chromatin stability. The
relevance of these observations is that while many
assisted reproduction technology laboratories
employ density gradient preparation for their IVF
spermatozoa, many still use simple washing for
ICSI since the risk of ROS -induced damage to
sperm dysfunction arising from simple centrifugal
washing (Mortimer, 1991) was not considered
important because sperm function is believed to
be unimportant for ICSI. However, since those
men deemed as requiring treatment by ICSI would
be more likely to have increased proportions of
abnormal spermatozoa, they would be most susceptible to ROS-induced sperm DNA damage.
Since it is well established that ROS-induced
damage to spermatozoa involving substantial
degradation of their DNA does not necessarily
affect their fertilizing ability (Aitken et al.,
1998), colloidal silica density gradients must be
considered the most appropriate generally applicable technique of sperm preparation (Mortimer,
2000).
Paternal effect on preimplantation
development
During the last few years a paternal influence upon
mammalian preimplantation development has been
recognized and at least indirect evidence found for
its action in human embryos (review: Sakkas, 1999).
For example, Janny and Menezo (1994) found that
more embryos stopped development when zygotes
were produced by IVF from men with abnormal
semen quality. Also, when culturing supernumerary
embryos remaining after day 2 embryo transfer it
was found that many fewer ICSI-derived embryos
106
(i.e. embryos produced from men with poorer semen
quality) reached the blastocyst stage compared to
IVF-derived embryos (Shoukir et al, 1998). The
relative significance of pre-existing damage to the
sperm DNA (i.e. that occurring in the male tract,
perhaps arising from infections: Ochsendorf, 1999)
compared to what might be induced during in-vitro
manipulation for assisted reproduction technology
remains unknown.
Provision of infertility health care
The management of male infertility in future will
continue to be greatly affected by 'non-scientific'
issues which cannot be ignored. The availability
of, and access to, medical services are controlled
by complex interactions between political, economic and social pressures, and payment for
infertility treatment is a balance between government, insurance and personal funding sources. For
example, in Australia, a highly effective national
infertility support organization has ensured strong
government support for IVF so that all infertile
women receive substantial coverage towards the
medical and laboratory costs of IVF. Moreover,
the gonadotrophins are provided under the
Pharmaceutical Benefits Scheme. However, the
availability of subsidized gonadotrophins does not
apply to artificial insemination cycles, and so IUI
is not covered by Medicare — making IVF the
cheapest option for patients in Australia. Yet in
nearby New Zealand, there is no government
support for IVF and the provision of infertility
health care is strongly controlled by resource
allocation issues. In Canada, a country that prides
itself upon the universal provision of health care
by the state, only one province (Ontario) has
partial government support for IVF, and in some
provinces, all infertility treatment is confined to
private or academic practitioners.
With cost-effectiveness becoming more integral
to health care provision, funding decisions in future
will be based increasingly upon outcomes analysis
and affordable technology. In some places it might
now appear financially sensible to provide one
cycle of ICSI as the primary (and sometimes sole)
treatment for all infertile couples because it saves
extensive and costly diagnostic investigations, but
more careful testing, increased success rates (e.g.
resulting from improved embryo culture systems),
Male infertility treatment
and complete analysis of the true cost of treatment
(e.g. including the associated costs arising out of
a multiple pregnancy resulting from the transfer
of more than two embryos) will lead to changes
in funding patterns, and to changes in patient
expectations.
Conclusions
A continued increasing recognition of the clinical
importance of sperm dysfunction, and a growing
awareness of the importance of the costeffectiveness of infertility treatment, foreshadow
changes in infertility treatment during the coming
decade. There will be increasing focus on genetic
testing of men with significantly impaired fertility
and, as the general uselessness of pathology laboratory semen analyses becomes more widely recognized, there will be a continuing integration of
specialized andrology laboratories into multidisciplinary infertility programmes and assisted reproduction technology units. These laboratories will
provide standardized, quality-controlled semen
analysis and sperm function assessments, both
before and during assisted reproduction technology
treatment (Mortimer, 1994c,d; World Health
Organization, 1999, Rowe et al, 2000). With
improved biotechnology, in particular the production of bioactive rhZP3, a new generation of sperm
function tests is foreseen that will allow routine
investigation of defects of sperm function, both
within the diagnostic process and during progressive treatment of the infertile couple. Structured
management of couples will become more widespread, especially in countries where health care
resources for infertility care are more limited, and
although ICSI will undoubtedly continue as a
highly successful mode of assisted reproduction
technology treatment, it will be seen more as a
last resort for those who cannot, or did not, achieve
a pregnancy by less invasive forms of treatment.
Contrary to the perceptions of some, recent
advances in assisted reproduction technology have
not diminished the value and importance of
diagnostic andrology but rather have revealed it to
the perspicacious as having even greater significance than recognized previously, a perspective
reflected in the revised edition of the WHO clinical
manual for the infertile male (Rowe et al., 2000).
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