Sperm Retrieval Techniques - Looking for a Needle in the Haystack
2012 Summer Internship ProgramCleveland Clinic Reproductive Research Center Sandro Esteves, MD, PhD Director, ANDROFERTCenter for Male Reproduction and Infertility Campinas, BRAZIL
Learning Objectives Understand azoospermia and the differences between obstructive and nonobstructive subtypes Learn the available methods for sperm acquisition in azoospermia and their indications Learn the success rates of sperm retrieval in different azoospermia scenarios Understand the reproductive potential of azoospermic men undergoing assisted conceptionEsteves, 2
Review this lecture at:http://bit.ly/ccfsummerinterns2012 Pdf slides Videos Reference papers
SpermatogenesisWhere do we stand compared to our relatives? Chimpanzee Human Gorilla 100 lbs 180 lbs 600 lbs 64 cc 20 cc 14 cc >1 64 5 billion/mL million/mL million/mL
Sperm Count in Humans General Population of Unscreened Men Centiles 2.5% 50% 97.5%Sperm countper mL (x106) 4 64 237 Azoospermia • Complete absence of sperm in the ejaculate • 1-3% male population • ~10% male infertility population Cooper et l. Hum Reprod Update 2009; Esteves et al, CLINICS 2011
Semen Analysis and Azoospermia Centrifugation at 3,000g for 15 minutes The supernatant is discharged and the pellet is examinedEsteves, 6
Types of Azoospermia Non- Obstructive obstructive• Normal sperm production • Absent or minimal sperm• Mechanical blockage anywhere production within the along the reproductive tract testicles • Epididymis • Testicular failure • Vas Deferens • Ejaculatory Duct
Management of Azoospermia OA NOA non- treatable treatable ductal sperm reconstruc- retrieval tion and ICSI Esteves et al. An update on the initial assessment of the infertile male. CLINICS 2011;66:1-10.
1. Azoospermia is a descriptive term of ejaculates that lack spermatozoa without implying a specific underlying cause.2. Azoospermia is not synonymous of sterility. Treatment options are microsurgical ductal reconstruction (selected cases of OA) and sperm retrieval coupled with in vitro fertilization (ICSI).
Sperm Retrieval Goals Immediate use for Obtain ICSI spermfor ICSI Cryopreservation Future retrievalsMinimizedamage Testicular function
Sperm Retrieval Techniques Technique Acronym Indications Percutaneous Epididymal PESA OA cases only Sperm Aspiration Microsurgical Epididymal MESA OA cases only Sperm Aspiration Testicular Sperm Aspiration TESA; Failed PESA in OA TEFNA1 Epididymal agenesis in CAVD Favorable histopathology in NOA Previous SR success in NOA Testicular Sperm Extraction TESE Failed PESA or TESA in OA (single or multiple biopsies) NOA cases Microsurgical Testicular Micro-TESE NOA cases only sperm Extraction Esteves et al. Sperm Retrieval Techniques for Assisted Reproduction.Esteves, 12 Int Braz J Urol 2011; 37(5):570-83
Sperm Retrieval in Obstructive Azoospermia • Epididymis • Testicle • Simple and Effective Esteves SC & Agarwal A. Sperm Retrieval Techniques; In: Gardner D et al (Eds.), HumanEsteves, 13 Assisted Reproductive Technology. Cambridge University Press, pp. 41-53, 2011.
Percutaneous Sperm Retrieval in Obstructive AzoospermiaPlease visit http://androfert.com.br/videos to watch this video
PESA alone PESA + rescue TESA 97.3 % OBSTRUCTIVE AZOOSPERMIA 100% 96.6% 96.3% 78.1 % Successful Retrievals CBAVD Vasectomy Post-infection Esteves et al. Reproductive potential of men with OA undergoing percutaneous sperm retrieval and ICSI according to the cause of obstruction. J UrolEsteves, 15 2012, submitted.
Epididymal/Testicular sperm Ejaculated sperm 70 73.6 P>0.05 48.5 46.3 51.3 43.2 20 12.1 %2PN %Top quality % Pregnancy % Miscarriage Fertilization embryos Verza Jr S & Esteves SC. Sperm defect severity rather than sperm source is associated with lower fertilization rates after intracytoplasmic sperm injection. Int Braz J Urol 2008; 34:49-56.Esteves, 16
CBAVD Post-vasectomy Post-infection P>0.05 265 277 250 Maformation rate: 1.5% Perinatal mortality: 1.5% 34.4 32.2 36.4 35.8 37.0 35.5 % Live birth Gestational age (wks) Birth weight (gramsx10) Esteves et al. Reproductive potential of men with OA undergoing percutaneous sperm retrieval and ICSI according to the cause of obstruction. J UrolEsteves, 17 2012, submitted.
Non-obstructive Untreatable Azoospermia condition Small testes/elevated FSH/”sterile” Absent or minimal production for sperm to appear in ejaculate Heterogeneity of sperm production: 600-800 seminiferous tubules/testis; Single focus of production adequate to retrieve spermatozoa for ICSI Goal: To identify and retrieve sperm for ICSI, but… Geographic location unpredictableEsteves, 19
Can We Predict Sperm Retrieval Success in NOA? Important because: 1. Can minimize emotional and financial cost of IVF cycles. 2. Can minimize trauma/damage to testis during sperm harvesting.Esteves, 20
Predictive Value of Noninvasive Tests for Sperm Retrieval in NOA FSH Testosterone Testicular Volume Verza Jr. & Esteves. Fertil Steril 2011; 96: S53Esteves, 21
Predictive Value of Noninvasive Tests for Sperm Retrieval in NOA Y Chromosome Microdeletion Screening Prevalence of Yq microdeletions: 1:2.000-3.000 newborns Azoospermic men: 5-12%Esteves, 22 Esteves, Miyaoka & Agarwal. An update on the initial assessment of the infertile male. CLINICS 2011; 66:1-10.
Predictive Value of Noninvasive Tests for Sperm Retrieval in NOA Y Chromosome Microdeletion Screening AZFb deletion Absence of retrievable sperm Esteves SC & Agarwal A. Novel concepts in male infertility.Esteves, 23 Int Braz J Urol 2011; 37:5-15.
Predictive Value of Invasive Tests for Sperm Retrieval in NOA Testicular Histopathology Sensitivity Specificity (95% Accuracy (95% CI) CI) (%) HYPO 93 (66-100) 70 (54-82) MA 64 (31-89) 59 (44-73) 81.9 SCO 20 (08-37) 20 (07-41) Esteves, Miyaoka & Agarwal. Surgical Treatment of Male Infertility in the ICSI Era.Esteves, 24 CLINICS 2011; 66:1463-77.
Predictive Value of Testingfor Sperm Retrieval in NOA
Sperm Retrieval inNonobstructive Azoospermia OPEN BIOPSY
Nonobstructive Azoospermia TESA vs. TESEControlled studies Needle Open Biopsyfor NOA men AspirationFriedler et al., 4/37 (11%) 16/37 (43%)Human Reprod 12:1488, 1997Ezeh et al. 5/35 (14%) 22/35 (63%)Human Reprod 13:3075, 1998
Conventional TESE (open biopsy) in NOA Number of patients 25 20 15 10 5 0 1 2 3 4 7 8 9 10 14 Number of testicular fragments excised Ostad et al., Urology 52:692, 1998.Esteves, 28
Nonobstructive AzoospermiaTesticular microdissection - micro-TESE• Method to identify site(s) of production – Based on the diameter of seminiferous tubules• Microsurgical approach – Identify site of production – Preserve vasculature of testis – Small quantity of tissue excised Schlegel PN. Testicular sperm extraction: microdissection improves sperm yield with minimal tissue excision. Hum Reprod. 1999;14:131-135.
Please visit http://androfert.com.br/videos to watch this video
Schlegel 1999Amer et al. 2000 Micro-Okada et al. 2002 TESE 43%-53%Okubu et al. 2002Tsujimura et al. 2002 TESE 25%-41%Ramon et al. 2003Esteves et al. 2011
1. Sperm retrieval techniques are surgical methods to collect spermatozoa from the epididimys or the testis of azoospermic men seeking fertility.2. The method of choice is based primarily upon the type of azoospermia being obstructive or nonobstructive.3. Retrieved sperm should be used for ICSI or cryopreserved for a future ICSI attempt.
Microsurgical vs Single-Biopsy TESE in Nonobstructive Azoospermia Controlled series of 60 patients Sperm Retrieval Success Rates Micro-TESE single-biopsy TESE 93% Method Histology categories P=0.0005 64% 64% pairwise 45% comparisons P<0.0001 25% 20% 9% 6% Overall Hypospermatogenesis Maturation Arrest Sertoli-cell Only Microsurgical versus conventional single-biopsy testicular sperm extraction in nonobstructive azoospermia: a prospective controlled studyEsteves, 33 Verza Jr S, Esteves SC. Fertil Steril 2011; 96 (3): S53
Hypospermatogenesis Sertoli cell-onlyVerza Jr & Esteves, Atlas of Human Reproduction, SBRH 2012
Conventional TESE Micro-TESE Microsurgical vs Single- Biopsy TESE in Nonobstructive Azoospermia Fragment weight Fragment weight Tissue Removal Open Large Micro- P- Single-Biopsy TESE value TESE Tissue Removed (mg) 65 ± 25 8.9 ± 2.5 <0.01Esteves, 36 Verza Jr & Esteves. Fertil Steril 2011; 96 (3): S53
Success of Sperm Retrieval by Cause of NOACryptorchidism 52-74%Varicocele 63-68%Post-infection 67%Torsion >50%Post-chemotherapy/RT 25-75%Genetic (Klinefelter, AZFc Yq microdeletions) 25-70%Idiopathic 50-60% OVERALL 40-60% Esteves et al., Fertil Steril 94; 2010; Raman and Schlegel. J Urol.170; 2003; Hopps et al. Hum Reprod. 180, 2003; Damani et al. JCO. 15; 2002
No. of Patients 255 % Retrieval Rate 51.1 No. ICSI cycles 328 Mean ± SD Patient Age Male 37.0 ± 7.6 Female 32.4 ± 4.7 Mean ± No. Injected Oocytes 9.8 ± 7.2 Mean ± %2PN Fertilization 43.7 ± 27.9 No. Transfer 298 Mean ± No. Embryos Transferred 2.4 No. Clinical Pregnancy (%) 86 (28.9) No. Live birth (%) 64 (21.5)Esteves, 38
No. of Babies Born 102 No. Multiple Deliveries (%) 29 (28.4) Mean ± SD Gestational Age 35.5 ± 2.7 Mean ± Birth Weight 2532 ± 601 Gender ratio; boy/girl 1.0/1.3 No. Perinatal Deaths 6 (5.9) No. Birth Defects (%) 3 (2.9)Esteves, 39
Sperm Retrieval Success Rates andReproductive Potential of Azoospermic Men undergoing ICSI Obstructive (N=142) Nonobstructive (N=172) 97.9% 55.2% 38.2% 25.0% Successful Sperm Retrieval Live Birth rateOdds-ratio 43.0 1.8695% CI 10.3 – 179.5 1.03 – 2.89P-value <0.01 0.03 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):S232-3.
1. Nowadays, the use of surgically-retrieved sperm and ICSI has become an established procedure for couples wishing to obtain a biological offspring in whom the male partner have azoospermia.2. So far, the post-natal outcomes of babies born from such fathers are reassuring.
Learning Objectives Understand azoospermia and the differences between obstructive and nonobstructive subtypes Azoospermia is the lack of sperm in the ejaculate Should be confirmed by semen analysis with centrifugation and pellet examination Found in ~10% of the male infertility population Types are Obstructive and Nonobstructive Congenital, acquired and unknown etiologies AO: sperm production is normal NOA: sperm production severely abnormal or absentEsteves, 44
Learning Objectives Learn the available methods for sperm acquisition in azoospermia and their indications Percutaneous (PESA, TESA) and open (MESA, TESE, micro-TESE) techniques are available Epididymides and testicles are the target organs Epididymal retrievals: Obstructive azoospermia Testicular retrievals: AO and NOA Microdissection TESE for the most difficult cases of NOAEsteves, 45
Learning Objectives Learn the success rates of sperm retrieval in different azoospermia scenarios Sperm retrieved in virtually all cases of OA Not related to collection method or cause of obstruction Sperm retrieved in 40-60% of NOA cases Current testing not reliable to predict SR success Success not related to the cause of NOA Men with AZF a or b microdeletions not candidates Higher SRR with micro-TESEEsteves, 46
Learning Objectives Understand the reproductive potential of azoospermic men undergoing assisted conception Success of ICSI Not related to collection method Related to the type of azoospermia Follow-up of children born similar outcomes (few data) Obstructive Azoospermia Similar (or better) results than ejaculated sperm ~40% live birth rates Nonobstructive Azoospermia Lower results than other infertility causes ~25% live birth ratesEsteves, 47