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Andrology Lab Corner* |
Centro de Infertilidad Masculina, La Coruña, Spain, and Harvard Medical School, Boston, Massachusetts
| Correspondence to: Dr. Juan G. Alvarez, Centro de Infertilidad Masculina, C/Fernando Macias, 8, 1C, 15004 La Coruña, Spain (e-mail: jalvarez{at}androgen.es). |
| Received for publication May 12, 2003; accepted for publication May 12, 2003. |
In this review, we describe two main strategies directed to optimize or "nurture" sperm production and quality: preproduction nurturing directed to optimize sperm quality before ejaculation and postproduction nurturing directed to preserve and optimize sperm quality and function after ejaculation.
This article does not attempt to provide a comprehensive review of all the interventions currently used to treat male infertility, but rather it is intended to provide some examples of interventions that can be taken to optimize sperm production in the fertile and infertile male (preproduction nurturing) as well as reproductive techniques used to help bypass defects in sperm function that would probably lead to infertility of the couple under unassisted conditions (postproduction nurturing).
Preproduction Nurturing
A number of environmental factors including high temperature, proinflammatory factors, social habits, drugs, radiation therapy, and xenobiotics have been shown to have a negative impact on testicular function. Perhaps the best example of preproduction nurturing would be simple intervention(s) directed to minimize the impact of exposure to these factors and that might also contribute to improve sperm production and quality. On the other hand, a number of nutraceuticals have been advocated as a way of potentiating sperm production and quality in the subfertile male. In this section, environmental factors and nutraceuticals shown to impact on sperm production and quality are described.
High Temperature
The importance of scrotal temperature in the regulation of sperm production
and quality has long been recognized. Variations in testicular temperature
have been proposed as a possible determinant of sperm quality
(Bedford, 1991;
Spira, 1991;
Mieusset and Bujan, 1995;
Setchell, 1998). Several
studies have found a higher scrotal temperature in infertile men, compared
with fertile controls, regardless of the cause of impaired fertility, eg,
varicocele (Mieusset et al,
1987; Zorgniotti and Sealfon,
1988; Mieusset,
1991). Likewise, in population-based studies, a negative
association between scrotal temperature and sperm concentration has been
demonstrated (Hjollund et al,
2000). In a retrospective survey of 402 fertile French couples, it
was found that it took longer to conceive a child if the man was exposed to
heat, eg, baker, welder, than if he was not
(Thonneau et al, 1996).
The reason for the variation in scrotal temperature under normal conditions is not well understood, although determinants could be of genetic nature. In a recent study, a correlation in median scrotal temperature has been found among monozygotic twins but not in dizygotic twins and a single-born brother (Hjollund et al, 2002). The results of this study suggest a genetic component to the variation in scrotal temperature. A hereditary element in male fecundity may be expressed through scrotal temperature, which constitutes a mechanism independent of those responsible for the development of the sperm-producing epithelium.
An increase in scrotal temperature can be brought about by an increase in environmental temperature, eg, occupational heat exposure in workers of glass and ceramic industry; social habits such as exposure to hot baths, Jacuzzis, saunas, etc or an increase in the endogenous temperature, eg, varicocele, fever. An increase in scrotal temperature not only can result in disruption of the process of spermatogenesis and spermiogenesis but also has been shown to induce sperm DNA fragmentation (Evenson, 2000). Therefore, one way of optimizing sperm quality would be to maintain scrotal temperature at physiological levels by avoiding occupational exposure to heat, treating episodes of fever with antipyretics, or performing varicocelectomy in cases of varicocele, eg, grades II or III, especially in adolescents in whom the presence of a varicocele could significantly compromise their long-term testicular function (Romeo et al, 2003).
Proinflammatory Factors
Chronic inflammatory disease has been shown to affect male reproductive
function and fertility. Relevant inflammatory diseases include general and
chronic infectious diseases as well as localized acute or chronic infections
of the male genitourinary tract. Male accessory gland infections account for
almost 15% of all cases of male infertility seen in infertility clinics, but
fertility usually is not a clinical objective among patients with acute
systemic infections such as Gram-negative sepsis
(Hales et al, 1999). Infections of the male accessory glands frequently are associated with
increased white blood cell count in semen and elevated levels of
proinflammatory cytokines in semen and the testis. There is an increasing body
of evidence that demonstrates the importance of cytokines and chemokines in
the regulation of testicular and glandular function during pathophysiological
states as well as under normal physiological conditions when cytokines act as
growth and differentiation factors. Cytokines have been shown to affect
Sertoli cell function and alterations in cytokine levels in the testis could
affect spermatogenesis (Cohen and Pollard,
1995). Montag et al
(1999) reported that use of
anti-inflammatory therapy in a male with nonobstructive azoospermia and
leukocytospermia resulted in resumption of sperm production and a significant
reduction in leukocyte concentration in the ejaculate. Therefore, prompt
diagnosis and treatment of these conditions could protect the testis against
the negative effect of these proinflammatory factors.
Social Habits
Alcohol consumption and cigarette smoking have been shown to affect sperm
production and quality. Cigarette smoking has been associated with reduced
sperm count and motility (Kunzle et al,
2003), increased oxidative stress
(Saleh et al, 2002), and
oxidative DNA damage (Fraga et al,
1991). Chronic ethanol exposure in peripubertal fathers decreases
fecundity, and this may be mediated by testicular oxidative injury, perhaps
leading to accelerated germ cell apoptosis
(Emanuele et al, 2001). In
addition, testes of rats fed an ethanol-containing liquid diet had more
testicular DNA fragmentation than mice fed an isocaloric control diet. Ethanol
increases the number of apoptotic spermatogonia as well as spermatocytes.
Direct intratesticular injections of ethanol enhanced testicular DNA
fragmentation, suggesting an increase in apoptosis. Moreover, Fas ligand
levels are increased within the testes of rats that were chronically fed
ethanol. In vitro, ethanol treatment of cultured Sertoli cells enhanced the
production of Fas ligand. In addition, testicular levels of p53 mRNA are
increased in rats chronically fed ethanol. All of these observations suggest
that ethanol enhances testicular germ cell apoptosis
(Zhu et al, 2000).
Therefore, males should be aware of the potentially damaging effects of alcohol consumption on testicular function. Investigation into the interplay of constitutive genes and disease modifier genes in males exposed to alcohol would help to reveal whether some of these males develop or not any testicular pathology.
Drugs
A number of pharmaceutical and recreational drugs have been shown to affect
testicular and sperm function (Nudell et
al, 2002). Some of these include alkylating agents, cocaine,
marijuana, calcium channel blockers, cimetidine, colchicine, cyclosporine,
erythromycin, gentamicin, neomycin, nitrofurantoin, spironolactone,
sulfasalazine, glucocorticoids, and tetracyclines. In a thorough fertility
evaluation of the male, the physician should determine what medication the
patient is taking as well as recreational drug use
(Nudell et al, 2002).
Discontinuing the offending agent(s) can reverse most of the induced adverse
effects. However, in some instances, such as chemotherapeutic regimens, the
medications cannot be discontinued, and pretherapy sperm cryopreservation
remains essential (Sweet et al,
1996). Treatment with cytotoxic chemotherapy is associated with
significant gonadal damage in men. The likelihood of gonadal failure following
cytotoxic chemotherapy is dependent on the type of drug and dose. At present,
sperm banking is the best method to ensure preservation of paternity, although
hormonal manipulation to enhance recovery of spermatogenesis and
cryopreservation of testicular germ cells are possibilities for the future.
Patients with testicular cancer should be informed of the side effects from
chemotherapy on testicular function and offered the option of sperm banking of
ejaculated sperm (nonazoospermic patients) or testicular sperm (azoospermic
patients) (Chan et al, 2001;
Schrader et al, 2003) prior to
the initiation of the therapy.
Radiation Therapy
Ionizing radiation has been shown to affect testicular function and sperm
production. Rowley et al
(1974) showed that direct
exposure to low-dose radiation (0.15 Gy) causes a significant reduction in
sperm count, with temporary azoospermia occurring after exposure to doses of
0.3 Gy.
Radioiodine treatment for thyroid cancer may result in transient impairment of gonadal function (Hyer et al, 2002). The radiation dose absorbed by the testis after a single ablative dose of radioiodine is well below that associated with permanent damage to germinal epithelium and the risk of infertility in these patients is minimal. Patients requiring multiple administrations for persistent or metastatic thyroid cancer may be at greater risk of gonadal damage (Hyer et al, 2002).
Therefore, patients exposed to radiation therapy should be informed of the potential effects of this therapy on testicular function and offered the option of semen cryopreservation prior to the initiation of the therapy, particularly in patients requiring multiple radiotherapy sessions.
Xenobiotics
Xenobiotics and, in particular, xenoestrogens have been implicated in the
decline of semen quality observed in the last decades. Some of these include
pesticides (Tielemans et al,
1999), insecticides (Hauser et
al, 2002), polyvinyl chloride plastics, polychlorinated biphenyls
(PCBs) (Rozati et al, 2002),
phthalate esters (PEs), and 1,2-dibromo-3-chloropropane
(Olsen et al 1990; Teitelbaum, 1999). In a recent
study, the highest average of PCB and PE were found in urban fish eaters,
followed by rural fish eaters, urban vegetarians, and rural vegetarians
(Rozati et al, 2002). The
total motile sperm counts in infertile men were inversely proportional to
their xenoestrogen concentrations and were significantly lower than those in
the respective controls. PCBs and PEs may be instrumental in the deterioration
of semen quality in infertile men
(Dallinga et al, 2002
Rozati et al, 2002).
Reproductive toxicants such as lead and cadmium have been associated with male infertility (Hess, 1998; Benoff et al 2000; Telisman et al, 2000). Occupational exposure to lead has recently been reported to induce sperm DNA fragmentation (Danadevi et al, 2003).
Benoff et al (2000) and subsequently Marmar (2001), have suggested that cadmium can act as a cofactor in the expression of varicocele-associated pathology in infertile males. They also suggested that other cofactors, such as gene modifiers, could also modulate the expression of this pathology. This may explain, at least in part, why studies involving the efficacy of varicocelectomy on sperm production and quality have not yet yielded definite conclusions concerning its potential beneficial effect.
Xenoestrogens have recently been identified as endocrine disruptors that not only may cause the so-called testicular dysgenesis syndrome (Skakkebaek et al, 2001) but also alter meiosis in germ cells at different stages of development. In fact, cumulative exposure to xenoestrogens has been proposed as a mechanism leading to meiotic alterations in human spermatozoa (Del Mazo et al, 1982).
Use of Nutraceuticals
Nutraceuticals have been advocated as a way of potentiating sperm
production and quality in the subfertile male. In a recent study, the
administration of folic acid and zinc sulfate to subfertile males was shown to
result in a significant improvement in sperm concentration compared to
placebo. Treatment lasted 25 weeks and the daily dose of folic acid and zinc
were 5 mg and 66 mg, respectively. Although the beneficial effect on fertility
remains to be established, this finding opens new avenues of future fertility
research and treatment (Wong et al,
2002).
Arginine (Pryor et al, 1978), vitamin B12 (Sinclair, 2000), methylcobalamin (Moriyama et al, 1987), and ginseng (Salvati et al, 1996) have been used in the treatment of male infertility. However, most of these compounds have marginal effects on sperm production and quality and have not been tested for safety and efficacy in randomized placebo-controlled studies. In fact, ginseng has been shown to have estrogenic activity (Duda et al, 1996) and produce adverse reactions (Hammond and Whitworth, 1981; Dega et al, 1996).
Because ROS overproduction has been associated with defective sperm function (Aitken et al, 1989), infertile patients have been treated with antioxidant compounds including, ascorbic acid (Fraga et al, 1991), vitamin E (Kessopoulou et al, 1995), selenium (Scott et al, 1998), gluthatione (Lenzi et al, 1998), vitamins, selenium, gluthatione, ubiquinol, and carnitine (Moncada et al, 1992; Lenzi et al, 1993; Costa et al, 1994; Sikka et al, 1995; Vitali et al, 1995; Hawkes and Turek, 2001; Vicari and Calogero, 2001; Lenzi et al, 2003). However, the effect of this treatment on sperm quality is still controversial (Ford and Whittington, 1998; Geva et al, 1998; Lenzi et al, 1998; Tarin et al, 1998; Comhaire et al, 1999). In a randomized, placebo-controlled, double-blind study, oral high doses of vitamins C and E to infertile males did not show any statistically significant improvement in semen parameters (Rolf et al, 1999). However, in this study, patient recruitment was solely based on having a sperm concentration below 50 million/ml.
One of the main reasons studies looking at the efficacy of antioxidant therapy in the treatment of male infertility have not yet been conclusive may be due to inadequate patient selection. Not all infertile males have an increase in oxidative stress in their testis and semen. Therefore, in principle, only those men who have a quantifiable increase in oxidative stress should benefit from antioxidant therapy. Perhaps the best marker to identify these males would be reactive oxygen species (ROS) levels in semen (Agarwal and Saleh, 2002). Another important aspect of antioxidant therapy is whether the antioxidant(s) and dose used in vivo are appropriate. Previous studies have indicated that the combination of vitamins E and C at high doses in vitro results in DNA fragmentation (Donnelly et al, 1999).
Postproduction Nurturing
In this section, the most common procedures used to preserve and optimize sperm quality and function after ejaculation as well as those reproductive techniques used to help bypass defects in sperm function are described.
Sperm Processing
Density Gradient Centrifugation—
Because seminal plasma contains factors that inhibit the fertilizing
ability of spermatozoa, it is essential that spermatozoa be separated from it
quickly and efficiently prior to any attempts at fertilization. In addition,
separation of mature sperm from ROS-producing immature sperm and leukocytes
during sperm processing is of paramount importance in preventing ROS-induced
damage to the sperm membranes and DNA of the mature sperm
(Lopes et al, 1998). It has
been previously reported that defective spermiogenesis is associated with the
release of high ROS-producing immature spermatozoa into the ejaculate
(Aitken and Clarkson, 1987). It
has recently been reported that sperm subsets in the ejaculate at different
stages of maturation and subsequently isolated by a 3-step density gradient
produce different levels of ROS
(Gil-Guzman et al, 2001). The
interface between 50% and 70% ISolate (Irvine Scientific, Santa Ana, Calif)
contains mostly immature sperm with proximal cytoplasmic retention. These
sperm produce the highest levels of ROS. Further, ROS levels in this fraction
were inversely correlated with percent motility in the sample
(Gil-Guzman et al, 2001) and
directly correlated with DNA damage, as measured by the sperm chromatin
structure assay test (Ollero et al,
2001). Recently, Sakkas et al
(2000) reported that sperm
isolated from semen by density gradient centrifugation showed a significant
improvement in nuclear integrity, as assessed by chromomycin A3
positivity and DNA strand breakage assays. When prepared using the swim-up
technique, the spermatozoa recovered showed no significant improvement in
nuclear integrity. Therefore, density gradient centrifugation techniques can
potentially enrich the sperm population by separating out those with nicked
DNA and poorly condensed chromatin.
Swim-Up— Unlike density gradient centrifugation techniques, which rely mainly on the separation of sperm by cell density (precluding, therefore, the migration to the gradient pellet of the low-density immature sperm), swim-up techniques rely solely on the ability of motile sperm to migrate to the upper culture media layer. Therefore, motile, low-density, ROS-producing immature sperm may be harvested from this fraction. In addition, one swim-up technique involves an initial semen washing by centrifugation step, which has since been shown to result in iatrogenic DNA fragmentation (Twigg et al, 1998). Because oxygen radicals have a short lifetime (nano-to microseconds), bringing immature, ROS-producing spermatozoa and/or leukocytes in close proximity to mature spermatozoa places these sperm at high risk of ROS-induced damage.
Centrifugation of sperm suspensions per se is not harmful to sperm unless mechanical damage is induced by high centrifugal forces. What might be really harmful is the cocentrifugation of mature and immature sperm, leukocytes, or both. Therefore, centrifugation of unprocessed semen should be discouraged and the use of swim-up techniques that bypass a semen centrifugation step recommended, such as direct swim-up (WHO, 1999). Although seminal plasma is known to contain antioxidant enzymes, centrifugation of semen has the dual effect of bringing mature and immature sperm in close proximity. In addition, it will significantly reduce sperm exposure to seminal plasma that, for the most part, will be in the supernatant, leaving a relatively dehydrated pellet devoid of antioxidant enzyme protection. However, these limitations may not apply to semen samples that are devoid of leukocytes or contain relatively low levels of ROS-producing immature sperm. Therefore, damage can be minimized by 1) using the World Health Organization-advocated direct swim-up; 2) using density gradient centrifugation (in which the pellet obtained is comprised, for the most part, of mature sperm); 3) diluting semen samples of high sperm concentration; and 4) use of testicular sperm obtained by testicular sperm extraction instead of epididymal sperm in assisted reproductive technology. Concerning the latter, it has been shown that DNA fragmentation is significantly higher in epididymal compared with testicular sperm (Steele et al, 1999).
In Vitro Treatment With Pentoxifylline and 2-Deoxyadenosine
Previous studies have shown that sperm treatment with pentoxifylline
increases the curvilinear velocity, path velocity, straight-line velocity,
lateral head displacement, beat cross-frequency, and sperm hyperactivation in
both normozoospermic and asthenozoospermic specimens
(Tesarik et al, 1992).
However, pentoxifylline does not modify the percentage of motile spermatozoa.
In a separate study, it was concluded that an unselective use of
pentoxifylline, 2-deoxyadenosine, or both compounds together may restore sperm
function in some patients and perhaps improve fertilization in vitro, but in
others it may produce no change or may even be detrimental to sperm function
(Tournaye et al, 1994).
Pentoxyfilline, in addition to stimulating sperm motility, has also been shown
to induce sperm capacitation and the acrosome reaction in fresh
(Ain et al, 1999) and
cryopreserved sperm (Esteves et al,
1998).
In Vitro Treatment With Platelet-Activating Factor
Previous studies have shown that human spermatozoa express
platelet-activating factor (PAF) receptor
(Reinhardt et al, 1999) and
that treatment of sperm with PAF results in a receptor-mediated improvement of
sperm function (Levine et al,
2002). Furthermore, in vitro treatment of spermatozoa with PAF has
been reported to significantly improve pregnancy rates in intrauterine
insemination (Wild and Roudebush,
2001). A beneficial effect for PAF on sperm function in vitro is
supported by the fact that sperm from a PAF receptor knockout mouse strain
display a significantly reduced rate of capacitation. When used for in vitro
fertilization, sperm from PAF receptor knockout mice gave a significantly
lower rate of fertilization (21.5%) than did wild-type sperm (66.7%)
(Wu et al, 2001).
Sertoli/Germ Cell Coculture
In vitro culture systems capable of supporting human early germ cell
differentiation have been developed for treatment of azoospermic patients or
patients with germ cell maturational defects. Sertoli cells, spermatogonia,
and spermatocytes, isolated from testicular biopsies of azoospermic patients,
have been cocultured using Vero cell-conditioned medium alone or supplemented
with recombinant (r) follicle-stimulation hormone (FSH) or rFSH plus
testosterone (Sousa et al,
2002). Optimal results are achieved using the hormones in
combination. In vitro matured spermatids microinjected into oocytes elicit
37.5% fertilization and 28.6% blastocyst rates. Abnormal elongating and
elongated spermatids resulted in 8.3% and 27.3% fertilization rates,
respectively, but none of the embryos reached the blastocyst stage. Normal
elongating and elongated spermatids result in 30.5% fertilization and 42.9% of
blastocyst rates (Sousa et al,
2002). Fluorescence in situ hybridization analysis showed sex
chromosome anomalies in all embryos except in the case of morulae from normal,
late spermatids. These results suggest that meiosis and spermiogenesis can be
resumed in vitro and with normal differentiated spermatids showing a low
fertilization potential but regular rates of blastocyst formation. However,
most of the embryos do not reach the morula stage and show major sex
chromosome abnormalities (Sousa et al,
2002).
Supplementation of Culture Media With Antioxidants
Supplementation of culture media with antioxidants has been used to
minimize sperm lipid peroxidation (Alvarez
and Storey, 1983). More recently, oxidative DNA damage induced by
30 Gy X-irradiation was reported to be prevented by ascorbic acid (600 µM),
alpha tocopherol (30 and 60 µM), and urate (400 µM) therapy. These
antioxidants provided protection from subsequent DNA damage by x-ray
irradiation. In contrast, acetyl cysteine or ascorbate and alpha tocopherol
together induced further DNA damage. In contrast, supplementation in vitro
with the antioxidants ascorbate, urate, and alpha tocopherol when used
separately had beneficial effects on sperm DNA integrity
(Hughes et al, 1998). These
results underscore the importance of careful antioxidant application and dose
selection in preventing oxidative stress.
Incubation of sperm from oligoasthenozoospermic samples at 5% O2 has been shown to result in a significant improvement in motility parameters, the percentage of hyperactivated motility, and induced-acrosome reaction, compared with those observed after incubation in an atmosphere of 20% O2. Exposure to 5% rather than 20% oxygen tension also induced a significant increase in the percentage of sperm penetrating zona-free hamster eggs after capacitation for 17 hours. After 24-hour incubation, a significantly higher survival rate is observed under 5%, compared with 20%, oxygen tension (Griveau et al, 1998), confirming previous reports (Alvarez and Storey, 1985; Alvarez et al, 1987). These results strongly suggest that use of low oxygen tension might improve sperm function by minimizing oxidative damage to sperm.
Use of Hyaluronic Acid
A relationship has been found between diminished cellular maturity of human
spermatozoa and low-level expression of the testis-specific chaperone protein,
HspA2 (formerly known as CK-MM). Because HspA2 is a component of the
synaptonemal complex in rodents and assuming that this is also the case in
men, it has been postulated that the frequency of chromosomal aneuploidies
would be higher in immature versus mature spermatozoa
(Kovanci et al, 2001). Hyaluronic acid (HA) has been shown to improve the proportion of high HspA2
and mature spermatozoa in a given sample
(Sbracia et al, 1997).
Therefore, the use of HA could be beneficial in assisted reproduction, eg,
intrauterine insemination and in vitro fertilization (IVF). Because HA is a
physiological component of the cumulus oophorus that surrounds the oocyte and
of the female and male reproductive tracts, clinical use of HA should not
cause ethical concerns. However, it should be emphasized that the release of
mature sperm, with a normal HspA2 profile, into the seminiferous tubules does
not ensure that these spermatozoa retain their fertilizing potential before
they leave the testis. Mature sperm will still be susceptible to DNA damage
during migration through the seminiferous tubules and the epididymis
(Steele et al, 1999). This
damage may not be compatible with the initiation and maintenance of a term
pregnancy, and yet these sperm retain a normal HspA2 profile.
Intracytoplasmic Sperm Injection
Intracytoplasmic sperm injection (ICSI) has been shown to be an effective
tool in the treatment of cases of severe oligozoospermia and
oligoasthenozoospermia (which before the introduction of ICSI would require
the use of donor sperm) or when sperm are unable to undergo capacitation, bind
to the zona pellucida, undergo the acrosome reaction, or fuse with the
oocyte's plasma membrane (Liu and Baker,
2000; Liu et al,
2003). Microinjection of spermatozoa directly into the oocyte's
cytoplasm could help bypass defects in these sperm functions.
Calcium Ionophore-Induced Egg Activation
Oocyte activation is a series of events triggered by the fertilizing
spermatozoon and necessary for the initial stages in embryonic development.
Calcium plays a pivotal role in this process
(Marangos et al, 2003). During
ICSI, there is an initial considerable but short (<2 minutes) increase in
[Ca2+]i that is detected immediately after the penetration of the
microinjection needle into the ooplasm, regardless of whether a spermatozoon
is simultaneously microinjected or not
(Tesarik et al, 1994).
Furthermore, this intracellular calcium rise, induced by the microinjection
pipette, does not itself provoke oocyte activation. After a lag period of 4-12
hours, oocytes undergo a second wave of [Ca2+]i changes. These
changes are sperm dependent and follow 1 of 2 alternative patterns, a
nonoscillatory one and an oscillatory one. The nonoscillatory pattern
resembles the changes described previously during parthenogenetic
(micropipette-initiated) activation of mammalian oocytes. The oscillatory
pattern is similar to the changes accompanying normal fertilization in
different mammalian species. Therefore, the initial [Ca2+]i rise
provoked by the ICSI procedure is not responsible for oocyte activation, and
the release of a sperm factor(s) is required to initiate this process
(Tesarik et al, 1994).
Oocytes can be activated in vitro following treatment with calcium ionophore A23187 and 6-dimethylaminopurine (6-DMAP) (Tesarik et al, 2000). In this procedure, oocytes are exposed for 10 minutes at 37°C to a solution of 10 µmol/L ionophore A23187 in Gamete-100 medium (prepared from a 2 mmol/L stock solution in dimethylsulphoxide), washed 3 times in fresh Gamete-100 medium, and incubated for 3 hours in IVF-50 medium supplemented with 2 mmol/L 6-DMAP (37°C, 5% CO2 in air). Oocytes can be activated by the combined treatment with ionophore A23187 and 6-DMAP. Therefore, in those cases of failed fertilization because of sperm failure to induce oocyte activation, calcium ionophore and 6-dimethylaminopurine could be used.
Recently a novel sperm-specific phospholipase C (PLC) has been identified as the sperm factor that triggers Ca2+ oscillations in mouse eggs indistinguishable from those observed at fertilization. PLC removal from sperm extracts abolishes Ca2+ release in eggs. Moreover, the PLC content of a single sperm is sufficient to produce Ca2+ oscillations as well as normal embryo development to blastocyst (Saunders et al, 2002). This PLC has the potential to be used to induce egg activation in in vitro fertilization.
Selection of Nonapoptotic Sperm
Apoptotic sperm expressing phosphatidylserine (PS) on the outer leaflet of
the membrane can be separated by magnetic-activated cell sorting (MACS) after
binding to superparamagnetic annexin V-conjugated microbeads (ANMBs)
(Paasch et al, 2003).
Spermatozoa from donors show lower levels of bound annexin V and activated
caspases than spermatozoa from infertile patients. MACS results in a
significant decrease of spermatozoa with activated caspases in both donors and
infertile patients. Separation effects of the MACS technique have been
confirmed with flow cytometry using antiannexin V antibodies and electron
microscopy. Therefore, ANMBMACS removes spermatozoa with PS-bound annexin V
and produces a higher-quality sperm fraction
(Paasch et al, 2003).
Summary
In spite of the significant progress made in human reproduction in the last decades, there are some conditions in the male that still lack a specific treatment for infertility, eg, microdeletions of the Y chromosome (AZFa, AZFb), meiotic defects, sperm maturational arrest, aneuploidies, defective centromeres, defects in oocyte activation, etc.
Nevertheless, the formidable advances made in the field of assisted reproduction in the last decade have given us the opportunity and means to help bypass defects in sperm function that would otherwise leave the couple as infertile. As indicated above, the introduction of ICSI has afforded the treatment of many forms of previously untreatable male factor infertility, eg, oligozoospermia, oligoasthenoteratozoospermia, azoospermia, sperm function abnormalities.
Concerning preproduction nurturing, although preventing or minimizing exposure to some of the factors described herein is known to favorably impact on testicular function and sperm quality, there are still some questions that remain to be answered. For example, can sperm production be resumed once exposure to a toxicant (eg, chemotherapy, xenobiotics) has been discontinued? Hopefully, future studies will provide answers to these questions.
Footnotes
* Andrology Lab Corner welcomes the submission of unsolicited
manuscripts, requested reviews, and articles in a debate format. Manuscripts
will be reviewed and edited by the Section Editor. Papers appearing in this
section are not considered primary research reports and are thus not subjected
to peer review. All submissions should be sent to the Journal of
Andrology Editorial Office. Letters to the editor in response to
articles as well as suggested topics for future issues are encouraged. 
References
Agarwal A, Saleh RA. Role of oxidants in male infertility: rationale, significance, and treatment. Urol Clin North Am. 2002;29:817-827.[Medline]
Ain R, Uma Devi K, Shivaji S, Seshagiri PB.
Pentoxifylline-stimulated capacitation and acrosome reaction in hamster
spermatozoa: involvement of intracellular signalling molecules. Mol
Hum Reprod. 1999;5:618-626.
Aitken RJ, Clarkson JS. Cellular basis of defective sperm function and its association with the genesis of reactive oxygen species by human spermatozoa. J Reprod Fertil. 1987; 81:459-469.
Aitken RJ, Clarkson JS, Fishel S. Generation of reactive oxygen species, lipid peroxidation and human sperm function. Biol Reprod. 1989;41:183-197.[Abstract]
Alvarez JG, Storey BT. Spontaneous lipid peroxidation in rabbit and mouse epididymal spermatozoa. Dependence of rate on temperature and oxygen concentration. Biol Reprod. 1985; 32:342-351.[Abstract]
Alvarez JG, Storey BT. Taurine, hypotaurine, epinephrine and albumin inhibit lipid peroxidation in rabbit spermatozoa and protect against loss of motility. Biol Reprod. 1983; 29:548-555.[Abstract]
Alvarez JG, Touchstone JC, Blasco L, Storey BT. Spontaneous lipid
peroxidation and production of hydrogen peroxide and superoxide in human
spermatozoa. Superoxide dismutase as major enzyme protectant against oxygen
toxicity. J Androl. 1987; 8:338-348.
Bedford JM. Effects of elevated temperature on the epididymis and testis: experimental studies. Adv Exp Med Biol. 1991; 286:19-32.[Medline]
Benoff S, Jacob A, Hurley IR. Male infertility and environmental
exposure to lead and cadmium. Hum Reprod Update. 2000; 6:107-121.
Chan PT, Palermo GD, Veeck LL, Rosenwaks Z, Schlegel PN. Testicular sperm extraction combined with intracytoplasmic sperm injection in the treatment of men with persistent azoospermia postchemotherapy. Cancer. 2001;92:1632-1637.[Medline]
Cohen PE, Pollard JW. Cytokines and growth factors in reproduction. In: Bronson R et al, eds. Reproductive Immunology. Cambridge, Mass: Blackwell Science; 1995.
Comhaire FH, Mahmoud AM, Depuydt CE, et al. Mechanisms and effects
of male genital tract infection on semen quality and fertilizing potential:
the andrologist's viewpoint. Hum Reprod Update. 1999; 5:393-398.
Costa M, Canale D, Filicori M, et al. L-carnitine in idiopathic asthenozoospermia: a multicenter study. Andrologia. 1994; 26:155-159.[Medline]
Dallinga JW, Moonen EJ, Dumoulin JC, Evers JL, Geraedts JP,
Kleinjans JC. Decreased human semen quality and organochlorine compounds in
blood. Hum Reprod. 2002; 17:1973-1979.
Danadevi K, Rozati R, Saleha Banu B, Hanumanth Rao P, Grover P. DNA damage in workers exposed to lead using comet assay. Toxicology. 2003; 187:183-193.[Medline]
Dega H, Laporte JL, Frances C, et al. Ginseng as a cause for Stevens-Johnson syndrome? Lancet. 1996; 347:1344.
Del Mazo J, Perez-Castillo A, Abrisqueta JA. Trisomy 21: origin of nondisjunction. Hum Genet. 1982; 62:316-320.[Medline]
Donnelly ET, McClure N, Lewis SE. The effect of ascorbate and
alpha-tocopherol supplementation in vitro on DNA integrity and hydrogen
peroxide-induced DNA damage in human spermatozoa.
Mutagenesis. 1999; 14:505-512.
Duda RB, Taback B, Kessel B, Dooley DD, Yang H, Marchiori J, Slomovic BM, Alvarez JG. pS2 expression induced by American ginseng in MCF-7 breast cancer cells. Ann Surg Oncol. 1996; 3:515-520.[Abstract]
Emanuele NV, LaPagli N, Steiner J, Colantoni A, Van Thiel DH, Emanuele MA. Peripubertal paternal EtOH exposure. Endocrine. 2001; 14:213-219.[Medline]
Esteves SC, Sharma RK, Thomas AJ Jr, Agarwal A. Cryopreservation of
human spermatozoa with pentoxifylline improves the post-thaw agonist-induced
acrosome reaction rate. Hum Reprod. 1998; 13:3384-3389.
Evenson DP, Jost LK, Corzett M, Balhorn R. Characteristics of human sperm chromatin structure following an episode of influenza and high fever: a case study. J Androl. 2000; 21:739-746.[Abstract]
Ford WCL, Whittington K. Antioxidant treatment for male
subfertility: a promise that remains unfulfilled. Hum
Reprod. 1998;13:1416-1419.
Fraga CG, Motchnik PA, Shigenaga MK, Helbock HJ, Jacob RA, Ames BN.
Ascorbic acid protects against endogenous oxidative DNA damage in human sperm.
Proc Natl Acad Sci USA. 1991; 88:11003-11006.
Geva E, Lessing JB, Lerner-Geva L, et al. Free radicals,
antioxidants and human spermatozoa: clinical implications. Hum
Reprod. 1998;13:1415-1424.
Gil-Guzman E, Ollero M, Lopez MC, Sharma RK, Agarwal A, Alvarez JG.
Differential production of reactive oxygen species by subsets of human
spermatozoa at different stages of maturation. Hum
Reprod. 2001;16:1922-1930.
Griveau JF, Grizard G, Boucher D, Le Lannou D. Influence of oxygen
tension on function of isolated spermatozoa from ejaculates of oligozoospermic
patients and normozoospermic fertile donors. Hum
Reprod. 1998;13:3108-3113.
Hales DB, Diemer T, Hales KH. Role of cytokines in testicular function. Endocrine. 1999; 10:201-217.[Medline]
Hammond TG, Whitworth JA. Adverse reactions to ginseng. Med J Aust. 1981; 1:492.
Hauser R, Altshul L, Chen Z, Ryan L, Overstreet J, Schiff I, Christiani DC. Environmental organochlorines and semen quality: results of a pilot study. Environ Health Perspect. 2002; 110:229-233.[Medline]
Hawkes WC, Turek PJ. Effects of dietary selenium on sperm motility in healthy men. J Androl. 2001; 22:764-772.[Abstract]
Hess RA. Effects of environmental toxicants on the efferent ducts, epididymis and fertility. J Reprod Fertil Suppl. 1998; 53:247-259.[Medline]
Hjollund NH, Bonde JP, Jensen TK, Olsen J. Diurnal scrotal skin temperature and semen quality. Int J Androl. 2000; 23:309-318.[Medline]
Hjollund NH, Storgaard L, Ernst E, Bonde JP, Christensen K, Olsen
J. Correlation of scrotal temperature in twins. Hum
Reprod. 2002;17:1837-1838.
Hughes CM, Lewis SE, McKelvey-Martin VJ, Thompson W. The effects of
antioxidant supplementation during Percoll preparation on human sperm DNA
integrity. Hum Reprod. 1998; 13:1240-1247.
Hyer S, Vini L, O'Connell M, Pratt B, Harmer C. Testicular dose and fertility in men following I(131) therapy for thyroid cancer. Clin Endocrinol. 2002;56:755-758.[Medline]
Kessopoulou E, Powers HJ, Sharma KK, Pearson MJ, Russell JM, Cooke ID, Barratt CL. A double-blind randomized placebo cross-over controlled trial using the antioxidant vitamin E to treat reactive oxygen species associated with male infertility. Fertil Steril. 1995; 64:825-831.[Medline]
Kovanci E, Kovacs T, Moretti E, Vigue L, Bray-Ward P, Ward DC,
Huszar G. FISH assessment of aneuploidy frequencies in mature and immature
human spermatozoa classified by the absence or presence of cytoplasmic
retention. Hum Reprod. 2001; 16:1209-1217.
Kunzle R, Mueller MD, Hanggi W, Birkhauser MH, Drescher H, Bersinger NA. Semen quality of male smokers and nonsmokers in infertile couples. Fertil Steril. 2003; 79:287-291.[Medline]
Lenzi A, Culasso F, Gandini L, et al. Placebo-controlled,
double-blind, cross-over trial of glutathione therapy in male infertility.
Hum Reprod. 1993; 8:1657-1662.
Lenzi A, Gandini L, Picardo M. A rationale for glutathione therapy.
Hum Reprod. 1998; 13:1419-1421.
Lenzi A, Lombardo F, Sgro P, Salacone P, Caponecchia L, Dondero F, Gandini L. Use of carnitine therapy in selected cases of male factor infertility: a double-blind crossover trial. Fertil Steril. 2003;79:292-300.[Medline]
Levine AS, Kort HI, Toledo AA, Roudebush WE. A review of the effect
of platelet-activating factor on male reproduction and sperm function.
J Androl. 2002;23:471-476.
Liu DY, Baker HWG. Defective sperm-zona pellucida interaction: a
major cause of failure of fertilization in clinical in vitro fertilization.
Hum Reprod. 2000; 15:702-708.
Liu D, Clarke GN, Martic M, Garrett C, Baker HWG. Frequency of
defective sperm-zona pellucida interaction in severely teratozoospermic
infertile men. Hum Reprod. 2003; 18:802-807.
Lopes S, Jurisicova A, Sun JG, Casper RF. Reactive oxygen species:
potential cause for DNA fragmentation in human spermatozoa. Hum
Reprod. 1998;13:896-900.
Marangos P, FitzHarris G, Carroll J. Ca2+ oscillations at
fertilization in mammals are regulated by the formation of pronuclei.
Development. 2003; 130:1461-1472.
Marmar JL. The pathophysiology of varicocales in the light of
current molecular and genetic information. Hum Reprod
Update. 2001;7:461-472.
Mathevon M, Buhr MM, Dekkers JC. Environmental, management, and genetic factors affecting semen production in Holstein bulls. J Dairy Sci. 1998;81:3321-3330.[Abstract]
Mieusset R. Scrotal hyperthermia; etiologic factors: facts and hypotheses. Adv Exp Med Biol. 1991; 286:211-216.[Medline]
Mieusset R, Bujan L. Testicular heating and its possible contributions to male infertility: a review. Int J Androl. 1995;18:169-184.[Medline]
Mieusset R, Bujan L, Mondinat C, Mansat A, Pontonnier F, Grandjean H. Association of scrotal hyperthermia with impaired spermatogenesis in infertile men. Fertil Steril. 1987; 48:1006-1011.[Medline]
Moncada ML, Vicari E, Cimino C, et al. Effect of acetylcarnitine treatment in oligoasthenospermic patients. Acta Eur Fertil. 1992;23:221-224.[Medline]
Montag M, van der Ven H, Haidl G. Recovery of ejaculated spermatozoa for intracytoplasmic sperm injection after anti-inflammatory treatment of an azoospermic patient with genital tract infection: a case report. Andrologia. 1999; 31:179-181.[Medline]
Moriyama H, Nakamura K, Sanda N, et al. Studies on the usefulness of a long-term, high-dose treatment of methylcobalamin in patients with oligozoospermia. Hinyokika Kiyo. 1987; 33:151-156.[Medline]
Nudell DM, Monoski MM, Lipshultz LI. Common medications and drugs: how they affect male fertility. Urol Clin North Am. 2002; 29:965-973.[Medline]
Ollero M, Guzman-Gil E, Lopez MC, Sharma RK, Agarwal A, Larson K,
Evenson DK, Thomas A Jr, Alvarez JG. Characterization of subsets of human
spermatozoa at different stages of maturation: implications in the diagnosis
and treatment of male infertility. Hum Reprod. 2001; 16:1912-1921.
Olsen GW, Lanham JM, Bodner KM, Hylton DB, Bond GG. Determinants of spermatogenesis recovery among workers exposed to 1,2-dibromo-3-chloropropane. J Occup Med. 1990; 32:979-984.[Medline]
Paasch U, Grunewald S, Fitzl G, Glander HJ. Deterioration of plasma
membrane is associated with activated caspases in human spermatozoa.
J Androl. 2003;24:246-252.
Pryor JP, Blandy JP, Evans P, et al. Controlled clinical trial of arginine for infertile men with oligozoospermia. Br J Urol. 1978;50:47-50.[Medline]
Reinhardt JC, Cui X, Roudebush WE. Immunofluorescent evidence of the platelet-activating factor receptor on human spermatozoa. Fertil Steril. 1999; 71:941-942.[Medline]
Rolf C, Cooper TG, Yeung CH, Nieschlag E. Antioxidant treatment of
patients with asthenozoospermia or moderate oligoasthenozoospermia with
high-dose vitamin C and vitamin E: a randomized, placebo-controlled,
double-blind study. Hum Reprod. 1999; 14:1028-1033.
Romeo C, Ientile R, Impellizzeri P, Turiaco N, Teletta M,
Antonuccio P, Basile M, Gentile C. Preliminary report on nitric oxide-mediated
oxidative damage in adolescent varicocele. Hum Reprod. 2003; 18:26-29.
Rowley MJ, Leach D, Warner G, Heller CG. Effects of graded doses of ionizing radiation on the human testis. Radiat Res. 1974; 59:665-678.[Medline]
Rozati R, Reddy PP, Reddanna P, Mujtaba R Role of environmental estrogens in the deterioration of male factor fertility. Fertil Steril. 2002;78:1187-1194.[Medline]
Sakkas D, Manicardi GC, Tomlinson M, et al. The use of two density
gradient centrifugation techniques and the swim-up method to separate
spermatozoa with chromatin and nuclear DNA anomalies. Hum
Reprod. 2000;15:1112-1116.
Saleh RA, Agarwal A, Sharma RK, Nelson DR, Thomas AJ Jr. Effect of cigarette smoking on levels of seminal oxidative stress in infertile men: a prospective study. Fertil Steril. 2002; 78:491-499.[Medline]
Salvati G, Genovesi G, Marcellini L, et al. Effects of Panax ginseng C.A. Meyer saponins on male fertility. Panmineva Med. 1996;38:249-254.
Sandler B, Faragher B. Treatment of oligospermia with vitamin B12. Infertility. 1984; 7:133-138.
Saunders CM, Larman MG, Parrington J, et al. PLC: a sperm-specific
trigger of Ca2+ oscillations in eggs and embryo development.
Development. 2002; 129:3533-3544.
Sbracia M, Grasso J, Sayme N, Stronk J, Huszar G. Hyaluronic acid
substantially increases the retention of motility in cryopreserved/thawed
human spermatozoa. Hum Reprod. 1997; 12:1949-1954.
Schachter A, Goldman JA, Zukerman Z. Treatment of oligospermia with the amino acid arginine. J Urol. 1973; 110:311-313.[Medline]
Schrader M, Mller M, Sofikitis N, Straub B, Krause H, Miller K. "Oncotese": testicular sperm extraction in azoospermic cancer patients before chemotherapy-new guidelines? Urology. 2003; 61:421-425.[Medline]
Scott R, MacPherson A, Yates RW, Hussain B, Dixon J. The effect of oral selenium supplementation on human sperm motility. Br J Urol. 1998;82:76-80.[Medline]
Setchell BP. The Parkes Lecture. Heat and the testis. J Reprod Fertil. 1998;114:179-194.
Sikka SC, Rajasekaran M, Hellstrom WJG. Role of oxidative stress
and antioxidants in male fertility. J Androl. 1995; 16:464-468.
Sinclair S. Male infertility: nutritional and environmental considerations. Altern Med Rev. 2000; 5:28-38.[Medline]
Skakkebaek NE, Rajpert-De Meyts E, Main KM. Testicular dysgenesis
syndrome: an increasingly common developmental disorder with environmental
aspects. Hum Reprod. 2001; 16:972-978.
Sousa M, Cremades N, Alves C, Silva J, Barros A. Developmental
potential of human spermatogenic cells co-cultured with Sertoli cells.
Hum Reprod. 2002; 17:161-172.
Spira A. Epidemiologic aspects of the relationship between temperature and male reproduction. Adv Exp Med Biol. 1991; 286:49-56.[Medline]
Steele EK, McClure N, Maxwell RJ, Lewis SEM. A comparison of DNA damage in testicular and proximal epididymal spermatozoa in obstructive azoospermia Mol Hum Reprod. 1999; 15:831-835.
Sweet V, Servy EJ, Karow AM. Reproductive issues for men with cancer: technology and nursing management. Oncol Nurs Forum. 1996;23:51-58.[Medline]
Tarin JJ, Brines J, Cano A. Antioxidants may protect against infertility. Hum Reprod. 1998; 13:1415-1416.
Teitelbaum DT. The toxicology of 1,2-dibromo-3-chloropropane (DBCP): a brief review. Int J Occup Environ Health. 1999; 5:122-126.[Medline]
Telisman S, Cvitkovic P, Jurasovic J, Pizent A, Gavella M, Rocic B. Semen quality and reproductive endocrine function in relation to biomarkers of lead, cadmium, zinc, and copper men. Environ Health Perspect. 2000;108:45-53.[Medline]
Tesarik J, Nagy ZP, Mendoza C, Greco E. Chemically and mechanically
induced membrane fusion: non-activating methods for nuclear transfer in mature
human oocytes. Hum Reprod. 2000; 15:1149-1154.
Tesarik J, Sousa M, Testart J. Human oocyte activation after
intracytoplasmic sperm injection. Hum Reprod. 1994; 9:511-518.
Tesarik J, Thebault A, Testart J. Effect of pentoxifylline on sperm
movement characteristics in normozoospermic and asthenozoospermic specimens.
Hum Reprod. 1992; 7:1257-1263.
Thonneau P, Ducot B, Bujan L, et al. Heat exposure as a hazard to male fertility. Lancet. 1996; 347:204-205.[Medline]
Tielemans E, Burdorf A, te Velde ER, Weber RF, van Kooij RJ, Veulemans H, Heederik DJ. Occupationally related exposures and reduced semen quality: a case-control study. Fertil Steril. 1999; 71:690-696.[Medline]
Tournaye H, Wieme P, Janssens R, Verheyen G, Devroey P, Van
Steirteghem A. Incubation of spermatozoa from asthenozoospermic semen samples
with pentoxifylline and 2-deoxyadenosine: variability in hyperactivation and
acrosome reaction rates. Hum Reprod. 1994; 9:2038-2043.
Twigg J, Fulton N, Gomez E, Irvine DS, Aitken RJ. Analysis of the
impact of intracellular reactive oxygen species generation on the structural
and functional integrity of human spermatozoa: lipid peroxidation, DNA
fragmentation and effectiveness of antioxidants. Hum
Reprod. 1998;13:1429-1436.
Vicari E, Calogero AE. Effects of treatment with carnitines in
infertile patients with prostato-vesiculo-epididymitis. Hum
Reprod. 2001;16:2338-2342.
Vitali G, Parente R, Melotti C. Carnitine supplementation in human idiopathic asthenospermia: clinical results. Drugs Exp Clin Res. 1995;21:157-159.[Medline]
Wild MD, Roudebush WE. Platelet-activating factor improves intrauterine insemination outcome. Am J Obstet Gynecol. 2001;184:1064-1065.[Medline]
Wong WY, Merkus HM, Thomas CM, Menkveld R, Zielhuis GA, Steegers-Theunissen RP. Effects of folic acid and zinc sulfate on male factor subfertility: a double-blind, randomized, placebo-controlled trial. Fertil Steril. 2002; 77:491-498.[Medline]
World Health Organization. WHO Laboratory Manual for the Examination of Human Semen and Sperm-Cervical Mucus Interaction. 4th ed. Cambridge, United Kingdom: Cambridge University Press; 1999 .
Wu C, Stojanov T, Chami O, Ishii S, Shimizu T, Li A, O'Neill C,
Shimuzu T. Evidence for the autocrine induction of capacitation of mammalian
spermatozoa. J Biol Chem. 2001; 276:26962-26968.
Zhu Q, Meisinger J, Emanuele NV, Emanuele MA, LaPaglia N, Van Thiel DH. Ethanol exposure enhances apoptosis within the testes. Alcohol Clin Exp Res. 2000;24:1550-1556.[Medline]
Zorgniotti AW, Sealfon AI. Measurement of intrascrotal temperature in normal and subfertile men. J Reprod Fertil. 1988; 82:563-566.
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