“Surgical sperm retrieval techniques“.

Welcome to chapter 8.
The following chapter is called “Surgical sperm retrieval techniques“.
The author is Professor Fábio Firmbach Paqualotto.
1
After this chapter, the student should be familiar with the main surgical sperm retrieval techniques, such as
percutaneous sperm aspiration (PESA), testicular sperm aspiration (TESA), microsurgical epididymal sperm
aspiration (MESA), testicular sperm extraction (TESE) and microsurgical testicular sperm extraction (m-TESE).
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Azoospermia occurs in roughly 5 % of all examined infertile couples. The condition is either classified as
“obstructive” or “non-obstructive”. For each individual case it is important to also consider the specific etiology.
Obstructive azoospermia is the result of obstruction in either the upper or lower male reproductive tract
(epididymis, vas deferens, seminal vesicles or ejaculatory ducts). Sperm production may be normal but the
obstruction prevents the sperm from being ejaculated.
Causes of obstructive azoospermia include scarring from past infections, inguinal hernia or hydrocele
operations, vasectomy, and congenital absence of vas deferens.
Non-obstructive azoospermia is the result of a testicular failure. Sperm production is either severely impaired or
non-existent, although in many cases sperm may be found and surgically extracted directly from the testicles.
Causes of non-obstructive azoospermia include, systemic disease (including cancer), drugs, radiation and
toxins, genetic and hormonal disorders, testicular maldescent and testicular torsion.
Some cases of obstructive azoospermia are treatable using microsurgical reconstruction of the seminal tract (for
example vasectomy reversal). Pregnancy rates following reconstruction vary from 27 to 56 % of cases and
results are determined by a number of factors such as the site and duration of the obstruction.
Unreconstructable obstructive azoospermia and non-obstructive azoospermia have historically been relatively
untreatable conditions that required the use of donor spermatozoa for fertilization. Intracytoplasmic sperm
injection has transformed treatment for this type of severe male factor infertility. Depending on the type of
azoospermia, sperm can be retrieved either from the epididymis or the testes.
3
ICSI involves injecting a single sperm into an oocyte making it an ideal treatment for male factor infertility that
would be otherwise untreatable using conventional in vitro fertilization techniques, which require large numbers
of sperm.
Initial successes with surgically recovered spermatozoa were followed by an extensive research in alternative
methods for recovering spermatozoa. Sperm can be retrieved for intracytoplasmic sperm injection in a variety of
ways depending on the type of azoospermia. In non-obstructive azoospermia sperm needs to be directly
obtained from the testis. Testicular sperm extraction (TESE), testicular biopsy and testicular sperm aspiration
(TESA) are techniques used on men with non-obstructive azoospermia. The procedures can require multiple
biopsies, sometimes in both testicles. Testicular fine needle aspiration (TEFNA) is a straight forward procedure
that is well tolerated by men.
In many cases of obstructive azoospermia, sperm can be retrieved from either the epididymis or the testis. Men
with obstructive azoospermia may undergo microsurgical epididymal sperm aspiration (MESA), percutaneous
epididymal sperm aspiration (PESA) or any of the testicular sperm extraction techniques. Of the two epididymal
techniques, PESA is less invasive and does not require microsurgical skills or equipment.
Successful retrieval of sperm for ICSI (the outcome measure of many of the non-randomized trials) is the first
step towards the achievement of the more clinically relevant outcomes - fertilization, embryo development,
implantation, a clinical pregnancy, a live birth and finally, the most relevant outcome to every couple, a healthy
baby without malformation.
4
For sperm retrieval there are basically five techniques, percutaneous sperm aspiration (PESA), testicular sperm
aspiration (TESA), microsurgical epididymal sperm aspiration (MESA), testicular sperm extraction (TESE) and
microsurgical testicular sperm extraction (m-TESE).
5
In 1997 Tournaye et al. first described the MESA technique.
The main drawback of MESA is that the procedure invasive and expensive. It requires a basic knowledge of
epididymal anatomy and of microsurgical techniques. The major benefit of this procedure is its diagnostic
power; a full scrotal exploration can be performed concomitantly.
Furthermore, the number of spermatozoa retrieved is high, which facilitates cryopreservation.
Intracytoplasmic sperm injection may be performed later, and even in another center, using the frozen-thawed
epididymal spermatozoa, without jeopardizing the intracytoplasmic sperm injection success rate.
6
A scrototomy is usually performed under general or loco-regional (cord block) anesthesia. The preferable
method of choice is an operating microscopy with X50–80 magnification although binocular loupes may also be
effective. When this procedure is performed by a skilled surgeon with the proper microsurgical approach, there
is a minimal risk of fibrosis or postoperative obstructive of the epididymal tubules occurring from the procedure.
7
Percutaneous epididymal sperm aspiration (PESA) was first described by Craft et al. in 1993 and is considered
a less invasive technique than the variant MESA. PESE is performed under local or loco-regional anesthesia.
Epididymal spermatozoa may be aspirated blindly from the epididymis by means of a percutaneous puncture
using a 19 gauge needle. It can be performed on an outpatient basis, and the technique is quick and simple
compared to MESA and it is certainly more cost-effective.
It has been argued that there may be fewer complications after PESA than after MESA.
Although there is no substantial data in literature stating that epidiymal spermatozoa may be cryoperseved after
PESA, it has been occasionally mentioned. It has also been stated that PESA may be repeated in the same
patient several times with good results. Because of the blind percutaneous punction, inadvertent damage may
be caused to the fine epididymal structures which may cause uncontrolled bleeding, leading to
postinterventional fibrosis. Another disadvantage is the impossibility of performing a proper diagnostic work-up
and concomitant reconstructive microsurgery.
8
On the day of oocyte retrieval, percutaneous epididymal sperm aspiration is performed under local anesthesia.
By injection 2 mL of 1 % lidocaine without adrenalin in the pampiniform plexus, cord block anesthesia is
administered. The surgeon then stabilizes the epididymis.
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In the surgical technique, a 27-gauge needle is introduced into the proximal part of the epididymis. A 1 mL
syringe filled with human tubal fluid medium, performs a delicate suction. The sample is then replaced in the
dish and is assessed under a light microscopy (200X magnification) to see if there is sperm presence. Then the
epididymis is gently massaged so that the fluid moves into the tubing. Postoperatively, but still in the operating
room, pressure should be held on the aspirated site for five minutes. Then a fluff compression dressing and
scrotal supporter are applied and left in place for 24 hours.
10
To obtain a histological or cytological diagnosis of spermatogenesis or to recover spermatozoa, many different
methods have been described for percutaneous needle aspiration of the testis, using a biopty gun or a 19- or
21-gauge needle.
When using the biopty gun or the 19-gauge needle for aspiration, local or loco-regional anesthesia is required,
but if a 21-gauges or finer needle is used (fine needle aspiration), aspiration may be performed without any
anesthesia. The aspiration technique is simple and quick and is non-invasive except for biopty gun systems or
analogues.
A sperm recovery rate of 96 % may be obtained by a 21-gauge fine needle aspiration in patients with normal
spermatogenesis. While a 21-gauge needle provides material only suitable for cytological assessment, larger
needles may provide tissue cylinders which allow an accurate histopathological examination.
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Because of its minimally invasive character, the aspiration procedure is assumed to be
more patient-friendly. Scores were shown to be higher in patients undergoing aspiration, in
a prospective stress and pain assessment using a visual-analogue scale method. When
this data was expressed as proportions of patients experiencing more less pain than
expected, then fine needle aspiration had more patients experiencing less pain than
expected compared with open biopsies. The higher average stress scores in fine needle
aspiration patients may be due to some patients being very anxious about needles.
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The surgeon can immobilize the testicle by grasping it with the epididymis and cord between his fingers while
pulling the scrotum skin taut. The surgeon can then control the depth of the needle excursion, protecting the
epididymis from injury. In the superior pole of the testis a 21-gauge butterfly needle is inserted and quickly
advanced and pulled back multiple times, toward several parts of parenchyma.
The butterfly tubing is clamped adjacent to the hub of the syringe when the plunger reached 20cc. This
clamping allows the assistant to replace the 20cc syringe filled with 0.5cc of human tubal fluid medium without
releasing the negative pressure.
The sample is placed in the dish and assessed for the presence of sperm under light microscopy (200x
magnification).
References:
Pasqualotto, F. F., Rossi, L. M., Guilherme,P., Ortiz, V., Iaconelli, Jr., A., Borges Jr., Etiology-specific outcomes
of intracytoplasmic sperm injection in azoospermic patients. Fertil Steril 2005;83:606-611.
Pasqualotto FF, Rossi-Ferragut LM, Rocha CC, Iaconelli A Jr., Borges E Jr. Outcome of in vitro fertilization and
intracytoplasmic injection of epididymal and testicular sperm obtained from patients with obstructive and
nonobstructive azoospermia. J Urol 2002; 167: 1753-56.
13
Retrieving testicular spermatozoa by an open excisional biopsy is equivalent to a diagnostic testicular biopsy
procedure. The sampled tissue is disrupted and minced or digested by enzymes in order to release the
spermatozoa from the seminiferous tubules and to recover testicular spermatozoa.
This technique of testicular sperm extraction (TESE) is even simpler than PESA and may be performed under
local anesthesia. TESE has a 100 % recovery rate in patients with normal spermatogenesis.
Recovery rates are approximately 50 % in patients with testicular failure showing different degrees of maturation
arrest germ-cell aplasia (Sertoli cell-only patterns). Similar recovery rates are found in patients with 47,XXY
Klinefelter’s syndrome. The outcome after intracytoplasmic sperm injection remains poor, although testicular
sperm recovery represents a major breakthrough for patients suffering from non-obstructive azoospermia.
14
As proposed by Schlegel in 1999, a microsurgical approach may be used to take small tissue samples to
minimize tissue damage when taking multiple excisional biopsies.
After opening the tunica albuginea, the seminiferous tubules are exposed at 40–80X magnification and the more
distended tubules may be selected for micro-excision. In patients with incomplete Sertoli cell-only syndrome,
where there is a substantial difference in diameter for empty and filled tubules this technique may be very
useful.
A general problem with the needle techniques is the difficulty in choosing the right site in the testis. There is
evidence of focal areas of normal and abnormal tissue within the testis of participants with non-obstructive
azoospermia and different methods of biopsy require randomized evaluation.
Thorough exploration with a microscope enabled the surgeons to choose the largest tubules in the testis which
are associated with more spermatozoa. Evaluating this newer technique, a previous study reported better
retrieval results with microsurgical sperm extraction compared to conventional open testicular retrieval.
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The open approach offers a major advantage with the possibility of freezing testicular tissue and of performing
intracytoplasmic sperm injection whenever the final histopathology shows spermatozoa, which prevents
pointless ovarian stimulation for the female partner.
Intracytoplasmic sperm injection is however not always proposed, because in some patients the final
histopathology may not show testicular spermatozoa or late spermatids.
Histopathology is the best indicator of whether spermatozoa for intracytoplasmic sperm injection are present or
not in patients with maturation arrest, the predictive power is insufficient to exclude a patient based on a
negative biopsy result. In any case, cryopreservation of testicular spermatozoa may prevent repeated surgeries
if pregnancy does not occur after intracytoplasmic sperm injection.
Overall, considering the low pregnancy rates after intracytoplasmic sperm injection with fresh testicular
spermatozoa from patients with testicular failure, the use of frozen-thawed testicular spermatozoa may be
preferable to repeat surgery in these patients.
16
The onus remains with those in support of more invasive techniques of surgical retrieval of sperm, that require
greater surgical expertise to demonstrate that these techniques can be justified by performing suitably
powered randomized clinical trials. Such trials need to have a particular focus upon:
1.
Clear definition of the population of men studied in terms of etiology of azoospermia;
2.
Use of clinically relevant outcomes: not only clinical pregnancy and live birth rates, but also the rate of birth
of a normal healthy baby, and certainly not simply the success of retrieval of sperm suitable for
intracytoplasmic sperm injection;
3.
Cost effectiveness (with inclusion of a cost-benefit analysis).
As the prevalence of azoospermia is low, it remains unlikely that a single unit will attain numbers to confer
sufficient power to such a trial. Large multicenter trials would increase the power of the results.
17
Since the introduction of intracytoplasmic sperm injection technique, both epididymal and testicular
spermatozoa have been used successfully to achieve pregnancies.
A large variety of methods for recovering epididymal or testicular sperm have been proposed and each has its
advantages and disadvantages. The question may therefore arise as to which method should be preferred in
clinical practice.
From published scientific data, it appears that percutaneous aspiration of the testis may be the method of
choice in patients with normal spermatogenesis, because the sperm recovery rate is high in these patients and
most patients experience less pain with this procedure than with an open biopsy.
For patients with obstructive azoospermia who have not had a thorough medical examination, MESA is the
preferred method because a full scrotal exploration can be performed with this procedure and, if indicated, a
vasoepididimostomy may be performed concomitantly.
In azoospermic patients showing different degrees of maturation arrest or germ-cell aplasia, recovery rates are
approximately 50 % after multiple open biopsies. Because the fertilization rates are significantly lower after
intracytoplasmic sperm injection and because fertilization failure rates are quite high, ongoing pregnancies are
obtained in only 10 % of intracytoplasmic sperm injection cycles.
The method of choice by which to prevent repeat surgery and pointless ovarian stimulation in the female partner
may be the surgical recovery of testicular spermatozoa by multiple open biopsies, cryopreservation of the tissue
and finally thawing, however, only if the microscopic examination of the tissue has demonstrated the presence
of spermatozoa.
18
There is not enough data from randomized trials to recommend any particular surgical sperm retrieval technique
for either obstructive or non-obstructive azoospermia. Non-obstructive azoospermia is a difficult area to analyze
as the physiology of the testis may vary between individuals. Techniques are modified rapidly and there is much
variation between different centers and surgeons.
It is logical to use the least invasive and simplest (and thereby most cost-effective) method for surgical retrieval
of sperm in the absence of evidence to support more invasive or more technically difficult methods. The more
invasive methods should currently be reserved for situations where sperm cannot be retrieved by a less
invasive technique (such as needle aspiration of the epididymis or testis) or for evaluation in the context of a
randomized trial.
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