Treatment of male infertility and results of first , S. Simi

ORIGINAL article
Journal of Andrological Sciences 2009;16:91-97
Treatment of male infertility and results of first
level assisted reproductive techniques
I. Natali, P. Turchi*, S. Simi*, S. Cipriani**, G. Mario Lentini, L. Carignani, I. Ingrassia, L. Niccoli
Sterility Centre, Gynecology and Obstetrics Unit, Azienda USL 3 Pistoia, Pescia, Italy; * Andrology Service, Azienda
USL 4 Prato, Prato, Italy; ** [email protected]@4 Group for Epidemiologic Research Milan, Italy
Key words
Male infertility • Couple infertility • Sterility
• Assisted reproductive techniques • IUI •
PTS • Capacitation test
Objective. To study the impact of a systematic clinical approach to male
infertility on seminal parameters and pregnancy rate both spontaneous and
obtained by artificial insemination.
Materials and methods. Two groups of male dispermic partners of infertile
couples were compared. Group I: 26 consecutive men submitted only to
seminal evaluation before assisted reproductive techniques; Group II: 41
consecutive men submitted to a complete clinical evaluation and treatment,
if indicated, before artificial insemination.
Results. Mean values of sperm concentration in Group II were 6.70 x 106/ml
before treatment and 11.90 x 106/ml thereafter. The sum of a + b motility
and only type a motility were (mean ± standard deviation) 58.1 ± 24.3 and
6.7 ± 7.7 before treatment and 82.3 ± 14.1 and 35.8 ± 25.9 thereafter, respectively. The statistical test showed a significant difference (p < 0.05) in all
the seminal parameters taken into account. The pregnancy rates obtained in
Group I, per cycle (3.2) and per couple (7.7) were lower compared to those
obtained in Group II, with male treatment, per cycle (12) and per couple
Conclusions. Treatment of pathological conditions, in males, leads to an
improvement in the sperm count and in the sum of a and b motility and
the a motility alone. Even if the scientific literature fails to validate guidelines
for the treatment of male infertility, the present data strongly support the
importance of meticulous screening of infertile males and the possibility to
achieve improvements in fertility by means of the commonly used treatment
Infertility, once considered to be mainly due to female factors, is now
known to be attributed equally to both the male and female components 1 2. The gradual increase in the role of male factors in determining
couple infertility, occurring over the last few years, has led to increased
attention being focused on the physio-pathological mechanisms of male
infertility and its treatment, including also the use of assisted reproductive techniques (ART) 3-5. Male infertility can result from gonadal or
Corresponding author:
Ilaria Natali, Sterility Centre, Gynecology and Obstetrics Unit, Azienda USL 3 Pistoia, Pescia, Italy – Tel. +39 0572 460342 – Fax +39 0572 460223 – E-mail: [email protected]
I. Natali, et al.
extra-gonadal diseases 6, but also from environmental pollutants 7 and increased age at which a man,
today, seeks paternity 8-10. The difficulty in defining the
mechanism of injury, intrinsic in this multi-factoriality,
results in medical treatment of male infertility having
uncertain indications and often not leading to the expected benefits. If we exclude treatment of infertility
due to hypogonadotropic hypogonadism, scientific
evidence of the benefits of drug therapy is, still today,
somewhat limited 11. Furthermore, studies on the
clinical applications of treatment for male infertility,
correctly performed from a methodological viewpoint,
are, unfortunately, a minority. Indeed, the few published trials often lack appropriate selection of patients, frequently classified generically as “infertile” or
as belonging to seminal categories (oligozoospermic,
asthenozoospermic, etc.) regardless of the pathogenesis. Management of male infertility has evidencebased indications only as far as concerns hormone
treatment of hypogonadotrophic hypogonadism and
surgical blockage of the seminal tract 12. Hormone
treatment, conversely, lacks a clear role in the empirical treatment 13 although it has been reported
that male treatment with follicle-stimulating hormone
(FSH), before intracytoplasmic sperm injection (ICSI)
improves the implantation rate, probably by improving
embryo quality 14. Infection of the accessory glands often contributes to the condition of male infertility. It has
been reported that the use of carnitines, after treatment with non-steroidal anti-inflammatory drugs or
natural anti-oxidant, produces encouraging results in
patients with prostate-vesciculo-epididymitis 15-16. The
administration of natural anti-oxidant in patients with
abacteric leukocytospermia, leads to an improvement
in sperm function 17. The effect of antimicrobial drugs
on accessory gland infections, on seminal parameters
and pregnancy rate (PR), is still under debate and the
conclusions of various studies are, sometimes, controversial. Although in chronic bacterial prostatitis, the
indication to antibiotic treatment is established by the
most authoritative guidelines, there is not sufficient evidence concerning the benefit in terms of PR. Similar
considerations can be made regarding the surgical approach, for which, excluding the treatment of seminal
tract obstructions, there are no clear indications. The
chance of correction of varicocele itself, in infertility,
is the object of debate. Despite this uncertainty in the
indications to pharmacological and surgical treatment,
clinical assessment of the male partner, in an infertile
couple, is considered a critical step in the diagnostic
and therapeutic iter of the infertile couple, both by the
scientific community and, in Italy, by the law that regulates the ART (Law 40/2004). The aim of the present
study was to establish, in the management of infertile
couples, whether treatment of the male partner would
improve seminal parameters, to increase pregnancies,
both spontaneous and obtained by artificial insemination, compared to couples in whom male treatment is
not carried out. To answer these points, a comparison
has been made of the results, namely of seminal values and PR, obtained in two homogeneous groups of
couples treated in a first level ART centre.
Materials and methods
Overall, 67 consecutive infertile couples, seen at
the Infertility Centre of the Hospital of Pescia (USL
3 Pistoia, Italy), and assessed for male factor,
were divided into two groups, uniform for men’s
and women’s age (< 40 years) and exclusion of
infertility related to endometriosis or tubal factors.
The first group (control group) comprised 26 consecutive couples, with the above-mentioned features,
seen at the Centre between 1996 and 2002. Males,
in this group, were considered infertile due to varying
degrees of dyspermia but had not undergone clinical
evaluation. The second group (study group) consisted of 41 consecutive couples also with the features
described, seen at the Centre between 2002 and
2006. Males, in this group, underwent andrological
examination and, where indicated, medical and/or
surgical treatment. This split is not randomly assigned, but chronologically, since the couples were
observed consecutively both before and after the
beginning of a systematic collaboration between the
Centre and the Hospital Andrology Service.
Evaluation and treatment of males and females
For basal assessment of males, 2 semen specimens
were required for examination, one month apart, one
from the other, and one capacitation test. Semen examinations were repeated, at each control following
medical or surgical treatment. All semen tests were
performed in the laboratory of the Centre and always
by the same operator.
The diagnostic protocol, applied after 2002, has
been standardized and included, not only a detailed clinical history and physical examination, but
also echography with colour Doppler evaluation
for spermatic funiculus, as well as endocrinological and bacteriological assessment. Trans-rectal
prostato-vesicular echography and a genetic analysis (karyotype and evaluation for Y chromosome
microdeletions) were performed only in cases of
azoospermia and severe asthenoteratozoospermia.
The therapeutic criteria are outlined in Table I. The 41
Treatment of male infertility and results of first level assisted reproductive techniques
cases, in Group II, belong to a group of men “treated”
(irrespective of the problem and the treatment performed) since the purpose of the present study was
not to evaluate the efficacy of one as opposed to
another, but to consider the overall results where the
better combined diagnostic and therapeutic criteria
were applied. Cases of genito-urinary phlogosis,
with or without positive cultures, were treated with
antibiotics and antioxidants; varicocele was treated
in accordance with the American Urological Association (AUA) and American Society for Reproductive
Medicine (ASRM ) guidelines 18; idiopathic infertility
was treated with anti-oestrogens or FSH (when FSH
values were < 6.9 IU/L) or were not treated (Table I).
The diagnostic protocol for females included: hormonal profile in the various phases of the cycle,
evaluation of tube patency, and examinations to
exclude infectious disease. Patients were assigned
to hormone treatment, defined on the basis of their
hormone profile and ovarian and endometrial echographic characteristics when inseminations were
scheduled, and, in some cases, even when targeted
intercourse without insemination was programmed.
Preparation of semen samples
Spermograms were performed according to WHO
guidelines 19. On the day of insemination, or spontaneous intercourse, males were required to present
with not more than 3 days abstinence and sperm was
collected by masturbation. Immediately after liquefaction, semen was centrifuged on a gradient density
[Purception, Sage IVF (Cooper Surgical Group), USA].
If the entire volume exceeded 2 ml, it was divided into
samples of 2 ml and layered onto a gradient comprising by the phases 40% and 80% in 15 ml conical
centrifuge tubes. The samples were centrifuged at
300xg for 20 minutes, and the pellet obtained was
then suspended in 2 ml of medium [Sperm Wash,
Sage IVF (Cooper Surgical Group), USA], and centrifuged at 300xg for 7 minutes. The pellet obtained was
again suspended in 4 ml of medium for insemination
[Sperm Assist, Sage IVF (Cooper Surgical Group),
USA] ready for insemination. Data on the volume,
physico-chemical properties, concentration and type
of sperm motility were recorded both before and after
preparation of the semen.
Insemination procedure
The insemination was performed using a dual way
catheter (Coppetta Cervix Adapter, Wisap, Germany)
for sperm fallopian sperm perfusion (FSP), after
thorough cleaning of the uterine portio. Capacitate
(4 ml) was introduced followed by 3 ml of air. The
system remained attached to the uterine portio for
~30 minutes after being appropriately clamped 20.
Patients received support treatment during the luteal
phase with progesterone until the pregnancy test. In
the case of a positive test, treatment was continued
Table I. Models of male factor treatment.
Male infertility factor
Idiopathic Oligo-AstenoTeratozoospermia (iOAT)
(only in cases of recent infertility, in couple
without evident gynaecological problems
and female age < 37 yrs)
Anti-oestrogens; gonadotropin; anti-oxidants
Tamoxifen 20 mg/day for 3-6 mos; rFSH or
hMG 75-150 IU 3 times/wk x 3-6 mos; L
carnitine 2 g/day; L acetyl-carnitine 1g/day
x 3-6 mos; Vitamin E 600 mg/day x 3-6
Prostatitis & vesiculitis
FANS & cortisones; antioxidants; antibiotics
(if positive cultures)
Celecoxib 10 mg/day x 2 mos); Nimesulide
(200 mg/day); L-acetyl carnitine (1 g/
day) + L carnitine (2 g/day) for 12 wks;
Fluorquinolones (E. coli, Proteus, Klebsiella,
Chlamydia); doxycyline or azithromycin
If > 2 degrees, with seminal alterations,
FSH < 10 IU/l, and female age < 37 yrs
Surgical repair or percutaneous
Hypogonadotropic hypogonadism
Urinary or recombinant gonadotropin
HCG (1000-2000 IU 2-3 times/wk) +
FSH (75-150 IU 3 times/wk)
Induction of an antetrograde ejaculation
Penile vibromassage; TESE
Retrograde ejaculation
Induction of a antetrograde ejaculation
Imipramine 25-75 mg, desipramine 50 mg
Obstructive azoospermia (OA)
Restoring (where indicated) patency
Non obstructive azoospermia (NOA)
Cryo-conservation from TESE or TESA
Sending to III level
I. Natali, et al.
until the 10th week of pregnancy. Cases in which the
gestational sac, vitelline sac and fetal heartbeat were
identified have been considered as pregnancies.
Description of sample and statistical analysis
All seminal parameters analyzed refer to capacitate
semen. The following aspects were then compared:
1) concentration of spermatozoa in Groups I and II;
2) concentration at baseline and after treatment in
Group II; 3) type a + b motility in Group I vs. II; motility type a + b before and after treatment in Group
II; type a motility in Group I vs. II; 4) type a motility
before and after treatment in Group II.
Parametric (Student t test) and non parametric tests
(Wilcoxon test and Testing signs) were applied. The
statistical analysis of data was performed on the findings shown in Tables II and III. Statistical differences
have been considered significant for p < 0.05 values.
Sperm concentrations in Group I were between 1.2
and 60 x 106/ml, with a 95% confidence interval (95%
CI) between 10.30 and 24.50 x 106/ml and a mean
value of 17.40 x 106/ml. Spermatozoa concentration
values, in Group II, before treatment, were between
0 and 26 x 106/ml, 95% CI 4.70-8.60 x 106/ml, mean
6.70 x 106/ml; finally, sperm concentration following
treatment ranged from 0.2 to 65 x 106/ml, 95% CI
8.20-15.60 x 106/ml, and a mean equivalent to 11.90
106/ml (Table II). In Group I, the sum of a + b motility was between 40 and 95% (mean 71.7, SD 17.6),
while motility type a ranged from 0 to 60% (mean
13.1, SD 13.1). In Group II, the sum of a + b motility,
before treatment was between 0 and 90% (mean
58.1, SD 24.3), and after treatment, between 40 and
95% (mean 82.3, SD 14, 1), only type a motility, before treatment, ranged from 0 to 30% (mean 6.7, SD
Table II. Description of samples (Groups I and II). Group I includes males not subject to andrological assessment. Group II includes males
subject to clinical evaluation and treatment. Concentration 1 is before and concentration 2 after treatment. The concentrations of sperm
are obtained after capacitation.
Upper limit
Lower limit
Concentration Group I
Concentration 1 Group II
Concentration 2 Group II
Table III. Description of sample (mean, range and SD) (all parameters are referred to capacitate) in the two groups. Group I - Line
1: Concentrations, Line 2: Sum of a and b motilities, according to WHO classification, Line 3: only type a motility. Group II - Line 1:
Concentrations before treatment, Line 2: Concentrations after treatment, Line 3: Differences in concentrations 1 and 2, Lines 4 and 5:
Sum of a and b motilities before (1) and after (2) treatment, Lines 6: Differences in the sum of a and b motilities before (1) and after
(2) treatment. Lines 7 and 8: Type a motility before (1) and after (2) treatment and Line 9: Differences between values before and after
treatment. Motility is expressed as %.
Group I
Mean (range)
95% CI
17.4 (1.2-60)
Motility type a + b
71.7 (40-95)
Motility type a
13.1 (0-60)
Mean (range)
95% CI
Group II
Concentration 1
6.7 (0-26)
Concentration 2
11.9 (0.2-65)
5.3 (-9-43)
Motility type a + b 1
58.1 (0-90)
Motility type a + b 2
82.3 (40-95)
Difference mot. a + b 1 and mot. a + b 2
Difference conc. 1 and conc. 2
24.2 (-36-66)
Motility type a 1
6.7 (0-30)
Motility type a 2
35.8 (0-90)
29.1 (-16-75)
Difference mot. a 1 and mot. a 2
Treatment of male infertility and results of first level assisted reproductive techniques
Table IV. Significance of tests: p values.
Parametric test
Non-parametric test
1) Concentration between the 2 Groups
2) Concentration in Group II
3) Motility type a + b between Groups
< 0.0001
< 0.0001**
4) Motility type a + b in Group II
5) Motility type a between Groups
6) Motility type a in Group II
< 0.0001
< 0.0001**
* Wilcoxon Test; ** Signs Test.
7.7) and after treatment values were between 0 and
90% (mean 35.8, SD 25.9) (Table III).
Application of the parametric test (Student T test),
showed a statistically significant difference (p < 0.05)
in all the parameters taken into account; given the
small number of samples, non-parametric analysis
was also performed which led, for each analysis, to
a complete overlap of the results (Table IV).
Finally, with regard to the pregnancies obtained,
the small number of samples did not allow statistical tests to be performed, however, also reported
were data on pregnancy rates either spontaneous or
evolutive obtained, without male treatment, in Group
I, per cycle (3.2) and per couple (7.7) and those obtained with male treatment, in Group II, per cycle (12)
and per couple (46.34) (Table V).
Treatment of male disorders leads to an improvement in the concentration of sperm (both in the
treated group and when comparing the two groups),
the sum of a and b motility compared to the control
group, and motility a and b of spermatozoa of the
same cases after treatment compared with the previous data. Tests applied became more significant
(p < 0.0001) if referred to the sum of motilities and
the only to type a motility, considering the same subjects before and after treatment (Group II).
Analysis of samples
The present study can be considered part retrospective, i.e., data related to the control group or Group I,
and part prospective, i.e., data on the study group or
Group II. Each parameter (concentration, a + b motility,
a motility) taken into account in this study refers to the
semen treated in the laboratory, namely capacitate,
and, therefore, no conclusions can be drawn on basal
seminal parameters. When studies are focused on
evaluations of the effect of a treatment, methodological approaches require that the recruited subjects be
randomly assigned to one or the other treatment arm,
in order to obtain two homogeneous groups. This is,
indeed, a limit of the present study, namely, the fact
that the two groups did not emerge from a randomization procedure and this is immediately obvious from
the level of concentration which is statistically different.
Albeit, the aim, in this study, was not to compare two
treatments arms, but rather to compare findings in a
group of couples in which the male had not been clinically evaluated with those obtained in another group
of couples in which the male has been both assessed
and treated. Moreover, in the latter group, a statistically significant difference has been recognized in the
levels of concentration after treatment compared with
basal findings, thus representing a statistically significant improvement in a factor that should produce an
improvement in the final objective. This statistically
significant result, even if on a limited number of cases,
offers useful information regarding the effectiveness of
treatment aimed at improving the levels of concentration, which is a favorable prognostic factor with regard
to the possible outcome in a pregnancy. These data
could support the proposal launched for a randomized
study to be carried out on two arms each undergoing
specific treatment to be carried out according to strati-
Table V. Pregnancy rate per cycle before and after male treatment.
No. couples
No. cycles
Without male treatment
With male treatment
* Number of couples without male treatment is equal in Tables 5 and 6 as couples who achieved pregnancy spontaneously were in temporary suspension of treatment.
** Group of couples (33 to 41) in which pregnancies were achieved after having undergone insemination. In remaining group of 12 pregnancies were achieved
I. Natali, et al.
fied randomization for some important factors including also the type of diagnosis.
sent useful predictive criteria of the results of assisted
fertilization. It cannot be excluded, however, that the
use of a single value, including various parameters (up
to 9: concentration, motility, circular speed, straight
speed, average speed, linear and lateral movements
of the head, morphology, according to the WHO
1999, and to the Tygerberg criteria) may represent an
estimate of pregnancies closer to reality 24. Bollendorf
et al., in 1996, published details of a study on 950 IVF
cycles and 1448 IUI cycles. In the IUI cycles, following preparation of the semen, couples without sperm
type a in the capacitate had a PR equal to 2.5% while
couples with sperm type a in the capacitate had a PR
of 10.2% 19. Thus, the presence of sperm with type
a motility in the capacitate significantly increases the
chances of pregnancy in IUI cycles. In in vitro fertilization, proof of the link of the spermatozoa to the pellucide zone (PZ), associated with the presence of type
a sperm, is a significant test in predicting failure of
fertilization of oocytes in cases of unexplained infertility and moderate male factor infertility 25.
Analysis of data
Motility type is an important parameter in the evaluation of effectiveness of spermatozoa for oocyte
fertilization 21 and our records confirm that improvement in this parameter correlates with an increase in
pregnancies. Unfortunately, due to the limited number of cases, it is not possible to perform a statistical analysis of the pregnancies observed, however,
it is quite clear that as a result of improvements in
seminal parameters, the PRs (both spontaneous and
obtained with insemination), are higher (Tables 5,
6). Few studies have correlated an improvement
in seminal parameters with an increase in PR. In
a study published in 1998, the only parameter
significantly related with the PR was sperm motility following semen preparation, regardless of the
concentration. Almost 50% of couples with good
sperm motility achieve pregnancy, but only 8.3% of
couples with low motility conceive 22. The same conclusions were reached in another retrospective study
on 504 couples, submitted to 1,636 cycles of intrauterine inseminations 23. Couples, in that study, were
divided into 5 groups, based on the total number of
mobile sperm after preparation (TMSC). However, it
was noted that it was not the parameter TMSC that
correlated with the PR, but the percentage of sperm
moving in the capacitate. The percentage of live
births increased significantly when the percentage of
mobile sperm reached 40% or more whereas below
this value, only one live birth was achieved. In our
study, moreover, we can see not only an increased
number of pregnancies per couple resulting from
insemination (from 7.7% in Group I to 46.34% in
Group II), but also an increase in spontaneous pregnancies (3.8% vs. 14.6%), as well as an increase in
evolutive pregnancies in the spontaneous (3.8% vs.
14.6%) and induced pregnancies (7.7% vs. 39%).
Examination of sperm motility in semen, after capacitation, may represent a significant parameter associated with the probability of success of insemination,
thereby supporting the relative reliability of basal seminal parameters. Both the sum of motility a + b, and
only type a motility of capacitate sperm could repre-
First level treatment (artificial insemination)
Couples suffering from idiopathic infertility or from
male subfertility, should be directed first towards
first level treatment, since success is comparable
to that obtained with in vitro fertilization techniques,
with the advantage of low costs 26. With the exception of cases in which the use of in vitro fertilization
(IVF or ICSI) is due strictly to a male or female factor, proposing a first-level treatment must be part
of a gradual approach to the techniques of artificial
insemination and is, indeed, an attitude justified by
factors not of secondary importance. In fact, the use
of artificial insemination has proved useful in cases
of idiopathic infertility, low stages of endometriosis,
as well as in mild-moderate male infertility. Artificial
insemination is a valid low cost method, minimally
invasive and easily acceptable from the female’s
hormone treatment point of view. Complications are
rare and of little clinical importance 27.
As already pointed out, the scientific literature has,
so far, failed to validate guidelines for the treatment
Table VI. Pregnancy rate per couple without and with male treatment.
No. couples
Without male treatment
With male treatment
Treatment of male infertility and results of first level assisted reproductive techniques
of the various forms of male infertility, albeit our data
strongly support the importance of adequate screening of the apparently infertile male and the need to
achieve improvements in fertility by means of the
male infertility treatment options currently available.
It is tempting to suggest that the limited size of the
sample studied may not provide sufficient data to
assess the significance with any certainty since the
more the observations, the more reliable the results.
Albeit, the interesting data emerging from the present
investigation suggest that further studies on a larger
number of subjects and with adequate randomization
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