Failure to Launch. Impairment of Primary Spermatogenesis & Optimization of Reproductive Techniques
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1 Failure to Launch. Impairment of Primary Spermatogenesis & Optimization of Reproductive Techniques October 16, 2013 Ryan Flannigan PGY3 UBC Urology Objectives } Introduce Azoospermic Infertility & Workup. } Epidemiology } Investigations } Diagnosis & Mechanisms of primary failure of spermatogenesis. } Primary } Discuss management of primary failure of spermatogenesis. } Surgical Techniques } Hormonal Optimization } ICSI & Dr. Beth Taylor s experience 1
2 Objectives } Introduce Azoospermic Infertility & Workup. } Epidemiology } Investigations } Diagnosis & Mechanisms of primary failure of spermatogenesis. } Primary } Discuss management of primary failure of spermatogenesis. } Surgical Techniques } Hormonal Optimization } ICSI & Dr. Beth Taylor s experience
3 Presentation } I have this friend. Presentation } I have this friend. } He and his wife are trying to get pregnant, without any success. 3
4 Presentation } I have this friend. } He and his wife are trying to get pregnant, without any success. } On exam his testis are small 10 cc s. } I did a semen analysis, and it shows no sperm what do I tell him? Infertility } 15% of couples fail to conceive in 12 months. } Male Factors 20% } Both Male & Female factors 30-40% } Azoospermia } WHO: no spermatozoa are found in the sediment of a centrifuged sample } AUA: no sperm after centrifugation at 3000 x g for 15 minutes and examination of the pellet } 1% in the general population } 15% among infertile males. 1. World Health Organization. WHO Laboratory Manual for the Examination and Processing of Human Semen. Geneva: World Health Organization; th ed. 2. Male Infertility Best Practice Policy Committee of the American Urological Association; Practice Committee of the American Society for Reproductive Medicine. Report on optimal evaluation of the infertile male. Fertil Steril. 2006;86(5 Suppl 1):S
5 Presentation } I have this friend. } He and his wife are trying to get pregnant, without any success. } On exam his testis are small 10 cc s. } I did a semen analysis, and it shows no sperm what do I tell him? } Where do we go from here? Azoospermia Work up } History 5
6 Azoospermia Associated Conditions } Varicocele } Cryptorchidism } Past GU Infections } Trauma } Torsion } Chemotherapies } Radiation } Past fertility } Familial problems with fertility Azoospermia Work up } History & Physical examination } Testi size } Palpable Vas Deferens } Secondary sexual characteristics 6
7 Azoospermia Work up } History & Physical examination } 2 nd Semen analysis } Semen centrifuged for 15 minutes at a minimum rate of 3000 x g à pellet examined at 400x magnification & confirms NO sperm. } Post Ejaculate Urinalysis Azoospermia Work up } History & Physical examination } 2 nd Semen analysis } Semen centrifuged for 15 minutes at a minimum rate of 3000 x g à pellet examined at 400x magnification & confirms NO sperm. } Post Ejaculate Urinalysis } Hormonal } FSH & LH +/- T, Prolactin } Genetic Testing } Karyotype } 47XXY, 45X / 46XY } Y-Microdeletions } (CF screen if absent Vas on PE) 7
8 Azoospermia Work up } Results } Semen Analysis - azoospermic } FSH & LH are elevated } Testis remain hypogonadal. } UA is negative for sperm } Genetic screen is (P) Azoospermia Work up } Results } Semen analysis - azoospermic } FSH & LH are elevated } Testis remain hypogonadal. } UA is negative for sperm } Genetic screen is (P) } PRIMARY HYPOGONADAL HYPOGONADISM 8
9 Objectives } Introduce Azoospermic Infertility & Workup. } Epidemiology } Investigations } Diagnosis & Mechanisms of primary failure of spermatogenesis. } Primary } Discuss management of primary failure of spermatogenesis. } Surgical Techniques } Hormonal Optimization } ICSI & Dr. Beth Taylor s experience 9
10 10
11 LH 11
12 LH Testosterone Testosterone 12
13 FSH Testosterone FSH Sertoli Cells Initiate and support Spermatogenesis Fuel for Germ Cells: lactate, transferrin, androgen binding protein Testosterone Regulatory factors: stem cell factor, TFG, IGF-I, FGF, EGF 13
14 FSH B l o o d T e s t i B a r r i e r Testosterone 14
15 Primary Failure Spermatogenesis } Defective spermatogenesis may fall within a spectrum of 4 histopathologic scenarios: Primary Failure Spermatogenesis } Defective spermatogenesis may fall within a spectrum of 4 histopathologic scenarios: } Hyalinization } Sertoli Cell Only } Hypospermatogenesis } Maturation Arrest 15
16 Hyalinization } Klinefelter s & Mixed Gonadal dysgenesis Sertoli Cell Only } Absence of germ cells in the seminiferous tubules 16
17 Hypospermatogenesis Maturation Arrest } Early } Only spermatogonia or spermatocytes present. } Late } Spermatids may be detected without spermatozoa Weedin JW, Bennett RC, Fenig DM, Lamb DJ, Lipshultz. Early Versus Late maturation Arrest: Reproductive Outcomes of Testicular Failure. J Urol. 2011; 186(2):
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20 Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Past GU Infections } Trauma } Torsion } Chemotherapies } Radiation 20
21 Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infection } Trauma/Torsion } Chemotherapies } Radiation Y-Microdeletions 21
22 Y-Microdeletions 22
23 Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infection } Trauma/Torsion } Chemotherapies } Radiation Klinefelters } Genotype } 47 XXY, or mosaic 46,XY/47,XXY } 1: births } Phenotype } Often do no come apparent until puberty. } Deficits in coordination } Less virilization } Gynecomastia } Hypogonadism } Infertility 23
24 Klinefelters Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infection } Trauma/Torsion } Chemotherapies } Radiation 24
25 45,X / 46, XY } Present with: } Ambiguous genitalia } Infertility } Hyalinized & disorganized cytoarchitecture 45,X / 46, XY 25
26 45,X / 46, XY Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infections } Trauma/Torsion } Chemotherapies } Radiation 26
27 Varicocele } Among patients with infertility 16.6% have a varicocele. } 1.6% of azoospermic patients. } Proposed Mechanisms? } Oxidative Stress } Increased temperature } Increased Venous pressure à reducing arterial flow? (Fuzisawa 1989) } Increased toxic metabolites, with venous stasis? (Sweeney 1995). } = increased germ cell apoptosis. (Simsek 1998) Varicocele } Varicocelectomy? } In Oligospermic patients: } Metanalysis demonstrated a non-significant effect of 1.10 (95% CI ) (Kantartzi et al. 2007). 27
28 Varicocele } Varicocelectomy? } In Oligospermic patients: } Metanalysis demonstrated a non-significant effect of 1.10 (95% CI ) (Kantartzi et al. 2007). } Azoospermic patients } 27 azoospermic patients treated with microsurgical varicocelectomy } 33% developed sperm in ejaculate } 55% became azoospermic again within 6 months. Varicocele } 31 men with azoospermia underwent microsurgical varicocelectomy } 22% had sperm reported in semen analysis } 9.6% were adequate for ICSI } No increased sperm retrieval with TESE 28
29 Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infections } Trauma/Torsion } Chemotherapies } Radiation Cryptorchidism } Prevalence is 1-3% } Sperm on semen analysis is as follows: Normal Oligospermia Azoospermia Unilateral 35-72% 21-27% 0-8% Bilateral <20% 23-36% 45-77% } No evidence than pexing impairs fertility } Discussion about Antisperm antibodies Werder EA, Illig R, Torresani T et al. Gonadal function in young adults after surgical treatment of cryptorchidism. Br J Med. 1976, 2: Okuyama A, Nonomura N, Maniki M, et al. Surgical management of epididymal and ductal anomalies associated with testicular maldescent. J Urol, 1989, 142:
30 Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infections } Trauma/Torsion } Chemotherapies } Radiation Orchitis & Infections } 45 men with Gonococcal urethritis and epididymoorchitis } Testi Biopsy } Extensive seminiferous tubular necrosis & inflammatory cell infiltration } Semen analysis at 2 years: } 27% had persistent azoospermia } 33% remained Oligospermia 30
31 Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Past GU Infections } Trauma/Torsion } Chemotherapies } Radiation Trauma & Torsion } Abnormal semen analysis observed up to 12 years post torsion. } Rare Azoospermia } 37% Oligospermia } 56% have asthenospermia } 53% have teratozoospermia } *Abnormal spermatogenesis bilaterally } Method of Rx } Orchidopexy for viable testi does not further decrease sperm count. } >24hr Torsion, orchidopexy vs orchiectomy? Anderson JB, Williamson RCN. Fertility after Torsion of the Spermatic Cord. Br J Uro. 1990; 65:
32 Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infection } Trauma/Torsion } Chemotherapies } Radiation Chemotherapy B l o o d T e s t i B a r r i e r 32
33 Chemotherapy Meistrich M. Effects of chemotherapy and radiotherapy on spermatogenesis in humans. J Fertil Steri. In press Chemotherapy } Biologics? } Tyrosine Kinase inhibitors normal to reduced sperm counts (Seshadri 2004; Breccia 2008). } Interferon therapy 1 report of azoospermia (Longo 2002). } mtor inhibitors have not yet been studies (Meistrich 2013). 33
34 Chemotherapy } Biologics? } Tyrosine Kinase inhibitors normal to reduced sperm counts (Seshadri 2004; Breccia 2008). } Interferon therapy 1 report of azoospermia (Longo 2002). } mtor inhibitors have not yet been studied (Meistrich 2013). } Recovery } IF spermatogonial stem cells still exist ~12weeks } IF no surviving stem cells (alkylating chemotherapies) possible permanent azoospermia Etiologies Among a Spectrum of Disease } Genetic Screen } Y-Microdeletions } Karyotype } Klinefelter s XXY } Mixed Gonadal Dysgenesis 45, X/ 46, XY } Associated Conditions } Varicocele } Cryptorchidism } Orchitis & Infection } Trauma/Torsion } Chemotherapies } Radiation 34
35 Radiotherapy } Impairs spermatogensis: } Direct gonadal toxicity resulting in immediate apoptosis in cycling cells, and apoptosis up to 21wks (Meistrich 2013). } Oxidative stress } Become azoospermic by 18 weeks } Due to Maturation Depletion Radiotherapy } Dose Dependent Recovery } 0.2Gy ~ 21 week recovery } 1 Gy ~ 7 months for appearance, 2 yrs for pre-tx levels. } 6 Gy ~ 24 months } 10 Gy ~ only 15% will ever recover spermatogenesis } Fractionated dose are more harmful Meistrich M. Effects of chemotherapy and radiotherapy on spermatogenesis in humans. J Fertil Steri. In press 35
36 Unifying Pathophysiology? } Oxidative stress and Free Radicals } Free radicals are compounds with unpaired electrons } Ie. O2. and OH. } Generation increased in pathologic conditions, or when inflammatory macrophages or neutrophils are recruited. } Exert damaging effects on cellular membranes, covalent crosslinking to biologically active molecules Superoxide O2. & OH. Detoxifying Destructive Glutathione peroxidase Superoxide Dismutase Catalase. Hydrogen Peroxide + Cu++ OR Fe++ Reduces ROS to non damaging molecules Damaging molecules to cellular membranes, DNA, RNA. Differential detoxifying capacity of Germ cells versus Leydig & Sertoli cells 36
37 ROS effects on Germ Cells Cytokine-induced-stress-kinase E-Selectin Increased ROS Peroxidase damage to cellular membranes Initiates Germ Cell Apoptosis Germ Cell Apoptosis Bcl-2 mediated Caspace Cascade DNA Fragmentation Apoptosis Sertoli Cells activated to phagocytose apoptotic GC s Release Nuclear Factor Kappa Beta & IL-1, IL-2 37
38 Varicocele -Ronco Santoro & Romeo Romeo 2003 Cryptorchidism -Ikeda Ishikawa 2005 Petola 1995 Misro 2005 Radiotherapy -Sohal Marda Hasegawa 1997 De Rooij 2002 Oxidative Stress Orchitis/Infection -Reddy Spiess 2007 Chemotherapy -Arnon Tilgada Asmis Wolf & Baynes Sudharsan Santos 2008 Torsion -Turner Da Ros Lysiak 2001/2007 -Anim Rodriguez Ozkurkrugil 2004 Objectives } Introduce Azoospermic Infertility & Workup. } Epidemiology } Investigations } Diagnosis & Mechanisms of primary failure of spermatogenesis. } Primary } Discuss management of primary failure of spermatogenesis. } Surgical Techniques } Hormonal Optimization } ICSI & Dr. Beth Taylor s experience 38
39 Sperm Retrieval } Fine Needle aspiration (FNA) } Testi Biopsy } Testicular sperm extraction (TESE) } Microdissection testicular sperm extraction (micro-tese) 39
40 Micro-TESE 40
41 Ø Overall: 32% versus 57% successful sperm retrieval p< Post TESE Endocrine changes No difference between groups Mean decrease of testosterone ~310à 250ng/dL 41
42 Post TESE Endocrine changes No difference between groups Mean decrease of testosterone ~310à 250ng/dL Optimizing Sperm Retrivals } Non-steroidal Antiestrogens } Aromatase inhibitors } Antioxidant Therapy 42
43 Non-steroidal Antiestrogen } Ie. Clomiphene } Increases FSH & LH release thus, increasing testosterone Non-steroidal Antiestrogen } 42 men with azoospermia } Clomiphene dosed to achieve testosterone ng/dL } 64% achieved sperm in semen } ALL remaining patients had successful sperm retrieval. 43
44 Aromatase inhibitors } Aromatase converts testosterone to estradiol Aromatase Inhibitor Steroidal -Testolactone Non-steroidal -Anastrozole -Letrozole Aromatase Inhibitors } 140 men with azoospermia & oligospermia } Treated with either Testolactone or anastrozole daily } Both had improved T:E. 5.3à 12.4 & 7.2à 18.1 } Oligospermic men improved semen parameters: motility, concentration & morphology. } Azoospermic men did not develop sperm in semen. } Benefit during sperm retrieval? 44
45 Specifics in Klinefelter s } Sperm retrieval rate is correlated to Testosterone levels } Sperm retrieval 68% } Pregnancy rate 57% } Live birth rate 45% Anti-Oxidant Therapies } Antioxidants used: } Vitamin C & E, zinc, selenium, folic acid, carnitine, N-acetyl cysteine, astaxanthin, & co-enzyme Q10 45
46 Anti-Oxidant Therapy } 35 azoospermic men } 11 controls } 24 received, multivitamins, Coenzyme Q10 & 2 lemons per day Following Sperm Retrieval } Sperm is used in Intracytoplasmic Sperm Injection (ICSI) } Please welcome Dr. Beth Taylor. 46
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