Dr. Sandro Esteves é palestrante internacional do 17o. Congresso da Sociedade Iraniana de Medicina Reprodutiva, ocorrido entre 2-4 março em Teerã.

Confira as aulas: "Role of Sperm Indexes in Embryo Quality" e "Sperm Retrieval Techniques".

Resumos:

Role of sperm indexes in embryo quality: what to do?

Sandro C. Esteves, MD, PhD

Spermatozoa are highly specializedcells with the purpose of not onlydelivering competent paternal DNA to the oocyte but also to provide a robust epigenetic contribution to embryogenesis. The identification of sperm fertility markers and the ability to selecthealthy spermatozoa for ART have a dual objective of choosing the best treatment strategy and optimizing ART outcomes. Currently, sperm indexes determination in the clinical setting is generally based on cell morphology and DNA content. Both sperm morphology and DNA integrity results, obtained from raw semen samples, have been shown to be of prognostic value for unassisted and assisted conception and useful in the selection of the best assisted conception modality.

These assays, however,provide an assessment of the distribution of cells in a given ejaculatethat may not be representative of the sperm population used in the ART treatment cycle. In fact, severe teratozoospermia,using Kruger’s strict criteria on pre-ART semen analysis, does notcorrelate to fertilization and embryo formation (including blastocyst development) in ICSI cycles. Nonetheless, if a more holistic approach to sperm morphology is taken, two prognostic groups can still be identified in cases of severeteratozoospermia (<4% normal) because certain morphology patterns and sperm abnormalities are known to affect ICSI outcomes. The first group includes mostly genetically determined sperm pattern defects, such asglobozoospermia, short tail syndrome and small-headed spermatozoa (in most cases combined with very small acrosomes). All of these types represent untreatable conditions that have been associated with abnormal sperm function andpoor ART outcomes. The second group includes unspecifiedor non-genetically determined sperm defects or patternscaused by environmental factors, medication, infection and related infertility conditions, including varicocele. Treatment of these conditions has been shown to optimize sperm morphology indexes with a positive impact on ART outcomes. Although the technician microscopically selects morphologically normal individual sperm during ICSI, form normalcy does not necessarily imply normal DNA content. As such, sperm DNA testing has been advocated to be an independent and reliable marker of fertility potential since sperm chromatin andDNA integrity is essential to ensure that the fertilizing sperm cansupport normal embryonic development of the zygote.At present, conflicting reports exist on the role of sperm DNA fragmentation index for embryo development, and it is apparent that DNA fragmentation does not significantly impair zygote and cleaving embryo morphology because major activation of the embryonic genome only beginafter the 4-cell stage. These observations do no underscore the importance of finding ways to increase sperm DNA integrity, since it has been suggested that DNA fragmentation is associated with late paternal effects that may lead to early miscarriages or diseases in the offspring. The etiology of sperm DNA damage is multi-factorial and may be due to primary (ageing, cryptorchidism, genetic defects, idiopathic) and or secondary (drugs, environmental, tobacco smoking, genital tract inflammation, infection,testicular hyperthermia and varicoceles) factors. Specific or non-specific treatments, including antioxidant supplements, are generally associated with reduced levels of sperm DNA damage and/or improved fertility potential.

Taken in conjunction, it is apparent that there is no unique sperm factor able to predict embryo development, but several candidate biomarkers are involved in this complex process.As a result, a wide variety of techniques have been proposed, including externalization of phosphotidylserine (magnetic-activated cell sorting),cell charge (zeta charge),maturity markers (hyaluronic acid binding) and detailed morphological analysis (intracytoplasmic morphologically selection sperm injection).Currently, these are several shortcomings for the routine application of these new methods to a busy IVF laboratory,both financially and logistically, and current data fail to indicate superiority of any of these methods over conventional ICSI.It is clear that better sperm fertility tools are urgently required. In this context, metabolomics and proteomic sperm profiling are under investigation and may be translated into clinical practice in the near future.

References

Menkveld R, Holleboom CAG, Rhemrev JPT. Measurement and significance of sperm morphology. Asian J Androl 2011; 13:59–68.

French DB, Sabanegh ES Jr, Goldfarb J, Desai N. Does severe teratozoospermia affect blastocyst formation, live birth rate, and other clinical outcome parameters in ICSI cycles? FertilSteril. 2010; 93:1097-103.

Barratt CL, Aitken RJ, Björndahl L, et al. Sperm DNA: organization, protection and vulnerability: from basic science to clinical applications--a position report. Hum Reprod 2010; 25:824-38.

Tesarik J, Greco E, Mendoza C. Late, but not early, paternal effect on human embryo development is related to sperm DNA fragmentation. Hum. Reprod. (2004) 19 (3): 611-615.

Esteves SC, Oliveira FV, Bertolla RP. Clinical outcome of intracytoplasmic

sperm injection in infertile men with treated and untreated clinical varicocele.

J Urol. 2010; 184:1442-6.

SPERM RETRIEVAL TECHNIQUES FOR THE AZOOSPERMIC MALE

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.

References

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 intracytoplasmic sperm injection (ICSI) in obstructive azoospermic (OA) men according to the cause of obstruction. FertilSteril. 2010; 94 (Suppl):S233.

Prudencio C, Seol B, Esteves SC. Reproductive potential of azoospermic men undergoing intracytoplasmic sperm injection is dependent on the type of azoospermia. FertilSteril. 2010;94 (Suppl):S232-3.