Sperm retrieval techniques


Published on

Iran march 2011



Sandro C. Esteves, MD, PhD

Spermatozoa can be retrieved from either the epididymis or the testis, depending on the type of azoospermia, using different surgical methods such as PESA, MESA, TESA, TESE and micro-TESE.
In obstructive azoospermia (OA), sperm production is normal and gametes can be easily retrieved from the epididymis or the testicle in most cases, irrespective of the technique. PESA or TESA are simple and efficient methods for retrieving epididymal or testicular spermatozoa in men with OA. According to our data on OA, the etiology of the obstruction and the use of fresh or frozen-thawed epididymal/testicular sperm do not seem to affect ICSI outcomes in terms of fertilization, pregnancy, or miscarriage rates.
In cases of nonobstructive azoospermia (NOA), the efficiency of TESA for retrieving spermatozoa is lower than TESE, except in the favorable cases of men with previous successful TESA or testicular histopathology showing hypospermatogenesis. The use of microsurgery during TESE may improve the efficacy of sperm extraction with significantly less tissue removed, which ultimately facilitates sperm processing. Testicular histology results, if available, may be useful to predict the chances to retrieve sperm in men with NOA. Our data demonstrate that micro-TESE performs better than conventional TESE or TESA in cases of maturation arrest and Sertoli cell-only histological patterns, where tubules containing active focus of spermatogenesis can be positively identified using microsurgery. Testicular spermatozoa can be obtained even in the worst case scenario except in the cases of Y chromosome infertility with complete AZFa and/or AZFbmicrodeletions.
In both OA and NOA, sperm retrieval technique itself seems to have no impact on ICSI success rates. The main goal of PESA/TESA/TESE sperm processing is the recovery of a clean sample containing motile sperm. Such specimens are more fragile, and often compromised in motility, as compared to the ones obtained from ejaculates. Laboratory techniques should be carried out with great caution not to jeopardize the sperm fertilizing potential. Surgically-retrieved spermatozoa can be intentionally cryopreserved for future use. Spare left-over specimens that would be discharged after ICSI can also be cryostored. Different strategies can be developed according to each group’s results. If freezing of surgically-retrieved specimens provides results similar to those with the use of fresh sperm, then the use of freezing specimens would be preferable. If not, fresh specimens are preferable.
The reproductive potential of infertile men undergoing ART is related to the type of azoospermia. According to our data, the chances of retrieving spermatozoa (odds ratio [OR] = 43.0; 95% confidence interval [CI]: 10.3-179.5) and of achieving a live birth by ICSI (OR=1.86; 95% CI:l 1.03-2.89) were significantly increased in couples whose male partner had obstructive rather than non-obstructive azoospermia. Children conceived using sperm retrieved from men with OA and NOA should be followed-up because it is still unclear if there is an increased risk of birth defects when ICSI is carried out with non-ejaculated sperm.
Esteves SC, Glina S. Recovery of spermatogenesis after microsurgical subinguinal varicocele repair in azoospermic men based on testicular histology. IntBraz J Urol. 2005; 31:541-8.
Verza S Jr, Esteves SC. Sperm defect severity rather than sperm source is associated with lower fertilization rates after intracytoplasmic sperm injection. IntBraz J Urol. 2008,34:49-56.
Esteves SC, Verza S, Prudencio C, Seol B. Sperm retrieval rates (SRR) in nonobstructive azoospermia (NOA) are related to testicular histopathology results but not to the etiology of azoospermia. FertilSteril. 2010; 94(Suppl.):S132.
Esteves SC, Verza S, Prudencio C, Seol B. Success of percutaneous sperm retrieval and i

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  • My final topic of today is Azoospermia, defined as the absence of sperm in the ejaculate. In cases of azoospermia, two totally different clinical situations exist. In obstructive azoospermia, spermatogenesis is normal but a mechanical blockage exists in the genital tract, somewhere between the epididymis and the ejaculatory duct. Common causes of OA include vasectomy, post-infectious diseases, congenital conditions. In nonobstructive azoospermia, sperm production is extremely deficient or absent inside the testicles . Common causes of NOA include cryptorchidism, orquitis, Radio/Chemotherapy, use of gonadotoxic medication and sterioids, and genetic origin.
  • Certain cases of OA are potentially reversible by using microsurgical or endoscopic methods. Another option for this group of patients is ART. Sperm can be easily obtained from the epididymis or testicles by percutaneous or microsurgical methods.
  • This video will demonstrate what I just said. Observe this dilated seminiferous tubule surrounded by non-dilated ones.
  • NOA is a complete different story. It is an untreatable condition and men with NOA have been the most difficult to treat. The goal in NOA is to find testicular sperm for ICSI, but the problem is that sperm production is focal or absent in such cases, and geographic distribution of sperm production is not predictable. Percutaneous and open testicular biopsy are used to retrieve sperm; however, the open biopsy using microsurgery is the one that yields better retrieval rates. The principle is to open the testicle and look for enlarged seminiferous tubules using the operating microscope. Enlarged tubules are removed because they are likely to contain active spermatogenesis .
  • TESA is one of the most widely used techniques for sperm retrieval in NOA men, but controlled studies demonstrated that they are not reliable as compared to open biopsy. Even with open biopsy, successful retrieval rates are only fair, with the risk of removing excessive testicular parenchyma that may compromise testicular function.
  • Currently, several male infertility specialists consider that the microsurgically guided open biopsy is the technique of choice for sperm retrieval in NOA men. The first description of microsurgical dissection of testicular tubules to identify sperm containing regions was made by Peter Schlegel in 1999. After that, several studies have reported overall higher sperm retrieval rates by microdissection than by conventional biopsy or needle aspiration. I am gathering experience with this technique over the last 7 years. Here you can see how the seminiferous tubules are seen under the operating microscope, and a dilated tubule, which may harbor mature sperm, can be easily identified.
  • In this study, we reported our initial experience with micro-tese, focusing on the importance of Testicular Histology in determining the success of this technique in patients with NOA.
  • Half of our patient population have been previously submitted to testicular needle aspiration, and you can see that micro-TESE at least doubled the chance of finding sperm in the cases of MA and SCO as compared to TESA. From our data , Micro-TESE is not necessary for NOA patients with testicular histology showing HYPO, since the success rates of the less invasive needle aspiration are as good as the ones obtained with microdissection.
  • Now, I will present some of our own data on the reproductive potential of infertile men using assisted conception by ICSI.
  • In OA cases, on the other side, the absence of sperm in the ejaculate is exclusively due to an obstruction at some point of the ductal system, but spermatogenesis is normal. ICSI results in OA are very satisfactory and they are independent of the sperm source: sperm retrieved from the epididymis or testicles perform similarly and yields adequate fertilization, cleavage and pregnancy rates.
  • In this study, we reported our assisted conception experience using ICSI in a group of infertile men with different degrees of sperm abnormalities. Patients with sperm in the ejaculate were classified according to the severity of sperm defect: single defect means that only one of the seminal analysis parameters (count, motility or morphology) was below the normal values. We classified patients as having double or triple defect when a combination of 2 and 3 abnormal parameters were seen, respectively. Azoospermic patients were classified based on the type of azoospermia. Testicular sperm retrieved from NOA men have a significant reduction in the fertility potential when used for ICSI, as seen by decreased embryo quality and CPR, as compared to other subgroups. Please observe that Fertilization rates were related to the degree of sperm abnormality. Significantly lower normal fertilization rates were obtained when ejaculated sperm with triple sperm defect or testicular sperm from patients with non-obstructive azoospermia were used for ICSI in comparison with other groups. These findings may be related to the tendency of severely abnormal sperm to carry deficiencies which ultimately affects the capability of the male gamete to activate the egg and trigger the formation and development of a normal zygote and a viable embryo.
  • The chances of finding sperm, and the chances of achieving a live birth by ICSI is completely different depending on the type of azoospermia. Successful sperm retrieval are 43 times higher in man with OA compared to NOA. Although men with NOA should not be considered sterile, their chances of having their own offspring is significantly lower compared to the counterpart with OA.
  • Sperm retrieval techniques

    1. 1. Sandro Esteves, MD, PhD Director, ANDROFERT Center for Male Reproduction and Infertility Campinas, BRAZIL Esteves,
    2. 2. <ul><li>Azoospermia </li></ul><ul><ul><li>It is not a synonymous of sterility </li></ul></ul>
    3. 3. Obstructive Azoospermia <ul><ul><li>Epididymis </li></ul></ul><ul><ul><li>Testis </li></ul></ul><ul><ul><li>Simple and effective </li></ul></ul>Sperm retrieval for ART
    4. 4. Percutaneous Epididymal Sperm Aspiration (PESA) Please visit http://www.vimeo.com/21033482 to watch the video.
    5. 5. Esteves SC, Verza S, Prudencio C, Seol B. Success of percutaneous sperm retrieval and intracytoplasmic sperm injection (ICSI) in obstructive azoospermic (OA) men according to the cause of obstruction. Fertil Steril. 2010;94 (Suppl):S233.
    6. 6. Non-obstructive Azoospermia <ul><ul><li>Sperm production reduced or absent </li></ul></ul><ul><ul><li>Geographic location unpredictable </li></ul></ul>Sperm Retrieval for ART Untreatable condition TESA TESE
    7. 7. Azoospermia & ART Testicular Sperm Aspiration: TESA
    8. 8. NOA Testicular microdissection: micro-TESE <ul><li>Method to identify site(s) of production </li></ul><ul><ul><li>Based on the diameter of seminiferous tubules </li></ul></ul><ul><li>Microsurgical approach </li></ul><ul><ul><li>Identify site of production </li></ul></ul><ul><ul><li>Preserve vasculature of testis </li></ul></ul><ul><ul><li>Small quantity of tissue excised </li></ul></ul>Schlegel, Hum Reprod 1999; 14
    9. 9. Micro-TESE Please visit http://www.vimeo.com/21031980 to watch the video.
    10. 10. <ul><li>176 NOA men (mean age 36.9 years) </li></ul><ul><li>Microdissection TESE or TESA </li></ul><ul><li>Classified according to the Etiology of NOA </li></ul><ul><li>Biopsy for histology concomitant or prior to SR </li></ul>Sperm Retrieval Rates in NOA are RelATED TO Testicular Histopathology but not to the Etiology of Azoospermia Esteves SC, Verza Jr S, Prudencio C, Seol B; Fertil Steril 2010 Hypospermatogenesis (HYPO) Maturation Arrest (MA) Sertoli Cell Only Syndrome (SCO) Esteves, Androfert
    11. 11. Sperm Retrieval in NOA is not related to Etiology Chi-square; NS Esteves SC, Verza S, Prudencio C, Seol B. Sperm retrieval rates (SRR) in nonobstructive azoospermia (NOA) are related to testicular histopathology results but not to the etiology of azoospermia. Fertil Steril. 2010;94( Suppl.):S132.   Presence of Testicular Spermatozoa; N (%) By Method   TESA (N=61); overall SRR, N (%) 34/61 (55.7%) Hypo 26/26 (100.0%) MA 2/6 (33.3%) SCO 6/29 (20.7%) MA + SCO 8/35 (22.8%)* Micro-TESE (N=70); overall SRR, N (%) 39/70 (55.7%) Hypo 19/19 (100.0%) MA 7/12 (58.3%) SCO 13/39 (33.3%) MA + SCO 20/51 (39.2%)* *p value 0.03 By Cause of NOA   Varicocele (N=66) 45/66 (68.2%) Genetic (N=12) 6/12 (50.0%) Cryptorchidism (N=19) 12/19 (63.1%) Idiopathic (N=63) 33/63 (52.4%) Radio/chemotherapy (N=6) 3/6 (50.0%) Orchitis/Gonadotoxin/Endocrine (N=10) 10/10 (100.0%) Overall SRR; N (%) 109/176 (61.9%) p value NS
    12. 12. Results (2): Micro-TESE X TESA Sperm Retrieval in NOA is related to Testicular Histopathology Esteves SC et al Fertil Steril 2010; 94:S132 *TESA vs micro-TESE (MA + SCO): P=.03 Histology Sperm + TESA Sperm + Micro-TESE HYPO 26/26 (100.0%) 19/19 (100.0%) MA 2/6 (33.3%) 7/12 (60.0%)* SCO 6/29 (20.7%) 13/39 (33.3%)* Total 34/61 (55.7%) 39/70 (55.7%) Esteves, Androfert
    13. 13. TESA/TESE N=131; *hypospermatogenesis excluded Source: Esteves et al.; Fertil Steril 2010; 94:S132 P=.03 Micro-TESE 39%
    14. 14. Reproductive Potential of Infertile Men in ART Esteves, Androfert
    15. 15. Sperm Defect Severity Rather Than Sperm Source Is Associated With Lower Fertilization Rates After Intracytoplasmic Sperm Injection Verza Jr S & Esteves SC. ANDROFERT – Brazil Int Braz J Urol 2008; 34 NS Obstructive Azoospermia Epididymal (n=31) Testicular (n=8) Female Age (years) 31.5 ± 7.7 36.3 ± 5.1 Mature oocytes (n) 9.4 ± 5.8 9.4 ± 4.9 Embryo Transfer (n) 3.3 ± 1.3 3.7 ± 1.5 2PN Fertilization (%) 74.7 ± 21.2 69.1 ± 19.6 TQE on Day 3 (%) 44.6 ± 30.5 52.7 ± 29.6 Clinical Pregnancy (%) 51.6 50.0 Miscarriage (%) 18.8 25.0 Esteves, Androfert
    16. 16. * P<0.05 Sperm Defect Severity Rather Than Sperm Source Is Associated With Lower Fertilization Rates After Intracytoplasmic Sperm Injection Verza Jr S & Esteves SC. ANDROFERT – Brazil Int Braz J Urol 2008; 34 ICSI Ejaculated Sperm (n=220) Sperm Defect Testicular/ Epididymal Sperm (n=93) Normal Single Double Triple OA NOA 2PN Fertilization (%) 71.3 73.2 72.1 63.4* 73.6 52.2* TQE on Day 3 (%) 48.4 50.5 46.9 48.3 46.3 35.7* Clinical Pregnancy (%) 40.9 36.6 44.4 51.0 51.3 25.9* Miscarriage (%) 14.9 9.1 12.5 12.0 20.0 14.3 Esteves, Androfert
    17. 17. Prudencio C, Seoul B, Esteves SC. Reproductive potential of azoospermic men undergoing intracytoplasmic sperm injection is dependent on the type of azoospermia. Fertil Steril 2010; 94 (4): Suppl. S232-233 . Sperm Retrieval Rates and Reproductive Potential of Azoospermic Men in ICSI Odds ratio 43.0 1.86 95% CI 10.3 – 179.5 1.03 – 2.89 P-value <0.01 0.03