This study evaluated the predictive power of spermatid count and cytology for assisted reproduction outcome after FSH therapy. A total of 174 men with severe oligozoospermia and normal plasma FSH concentration underwent semen analysis including spermatid count, TUNEL test, FISH analysis for sperm aneuploidies and testicular fine-needle aspiration cytology. Ninety-two men with hypospermatogenesis received FSH therapy for 3months and 82 patients with maturative disturbance or partial obstruction served as controls. Semen was analysed at baseline, after FSH therapy and after 3- and 9-month follow up, and pregnancies were recorded. Subjects not reaching pregnancy at 3-month follow up were recommended assisted reproduction treatment. Spermatid count was related to testicular cytology: spermatid concentrations <0.01, 0.01–0.3 and >0.3 × 106/ml were predictive of partial obstruction, hypospermatogenesis and maturative disturbance. FSH therapy patients showed increases in sperm number and motility (both P < 0.001), allowing some couples to undergo intrauterine insemination instead of IVF. Cumulative pregnancy rate after 12 months was higher with FSH therapy (44.6%) than without (22.0%; P = 0.002). FSH therapy improved pregnancy rate and sometimes allowed less invasive assisted reproduction treatment in well-selected patients. Spermatid count could represent a new parameter to predict response to FSH therapy.
One-hundred seventy-four patients with severe reduction of sperm count and normal sex hormones plasma levels underwent semen analysis with spermatid count, and testicular fine needle aspiration cytologiy (FNAC). Ninety-two men infertile men with reduced sperm production (hypospermatogenesis) were treated with highly purified urofollitropin and 82 patients with sperm maturative defects or partial obstruction of the seminal tract served as controls. After treatment and after the following 3 and 9months all subjects performed a new semen analysis and pregnancies were recorded. Subjects who had not reached spontaneous pregnancy were suggested to undergo assisted reproductive techniques (ARTs). Spermatid count was strongly related to testicular cytology: spermatid concentrations were predictive of partial obstruction, hypospermatogenesis and maturative disturbance respectively. Treated patients showed significant increase in sperm number and motility allowing some couples to undergo easier and less invasive assisted reproductive techniques. The number of pregnancies was significantly higher among treated (44.6%) than untreated couples (22.0%). Our data confirmed that FSH treatment can induce a significant improvemet of pergnancy rate and sometimes allows less invasive ARTs use in well selected severe oligozoospermic patients. Moreover, we suggest that spermatid count can be useful to define tubular status and could represent a new parameter to predict response to FSH therapy.
Introduction
Normal functioning of the gonadotrophic axis, in particular normal FSH secretion, plays crucial roles in spermatogenesis (Matsumoto, 1989, Sharpe, 1989), and several experimental and clinical studies have demonstrated the importance of FSH in regulating a normal quantitative spermatogenic process in animals and humans (Marshall et al., 1995, McLachlan et al., 1995, Moudgal et al., 1997). It has previously been demonstrated that FSH administration is able to increase sperm concentration, spermatogonal population and pregnancy rate in oligozoospermic subjects with normal plasma concentrations of gonadotrophins (Acosta et al., 1992, Bartoov et al., 1994, Glander and Kratzsch, 1997, Merino et al., 1996, Strehler et al., 1997). In addition, some authors reported that a significant improvement of sperm quality was evident after therapy. FSH therapy is useful in reducing the apoptotic process of spermatozoa and improving the qualitative properties of the axoneme, chromatin and acrosome (Baccetti et al., 1997).
Recently Colacurci et al. (2012) showed that the degree of sperm DNA fragmentation significantly decreases after 90 days of FSH therapy. However, reducing fragmentation cannot guarantee fertilization and embryo development success systematically. Some authors reported that genetic, in particular epigenetic causes, can lead to abnormal spermatogenesis, with potential implications for sperm quality, fertilization and embryo development (Jenkins and Carrell, 2012, Lalancette et al., 2009). In particular, Montjean et al. (2013) evaluated methylation profiles in mature sperm DNA from normozoospermic and oligozoospermic men by observing the presence of methylation changes in spermatozoa of the latter group.
Another potential mechanism of DNA damage that could be able to reduce the capability of spermatozoa to fertilize is related to DNA denaturation (Evenson and Wixon, 2008). It has been postulated that defective chromatin condensation during spermatogenesis could lead to spermatozoa unable to mature completely during spermiogenesis and therefore unable to repair DNA breaks during chromatin rearrangement occurring when histones are replaced by protamines (O’Brien and Zini, 2005, Zini and Libman, 2006). A positive effect on sperm DNA condensation after FSH therapy has been demonstrated in idiopathic infertile men (Kamischke et al., 1998).
These findings may explain the increase in oocyte fertilization and pregnancy rate, sometimes in absence of improved classic seminal parameters, that has been observed in couples undergoing assisted reproduction treatment in which the male partner received FSH (Acosta et al., 1991, Acosta et al., 1992, Ashkenazi et al., 1999). However, the improvement in seminal characteristics is variable and not much evident in many cases, and a proportion of patients do not respond to FSH therapy.
Oligozoospermia may be sustained by various alterations of the seminiferous epithelium that could explain the failure of FSH therapy reported by other studies (Attia et al., 2007, Bartoov et al., 1994, Foresta et al., 1995). Fine-needle aspiration cytology (FNAC) is a minimally invasive, rapid and effective procedure to evaluate the tubular status in oligozoospermic patients (Adhikari, 2009). This study group demonstrated that FSH therapy of oligozoospermic subjects with normal FSH plasma concentration, moderate hypospermatogenesis and absence of maturation disturbance is able to induce both a significant increase in sperm count (Foresta et al., 2002) and an improvement in pregnancy rate (Foresta et al., 2005). In contrast, when hypospermatogenesis is associated with maturative disturbance, FSH therapy shows no effect (Foresta et al., 2009). In the light of this consideration, selection criteria should be used to derive predictive information on the response to FSH therapy.
Some polymorphisms in the FSH receptor gene (FSHR) are able to influence the sensitivity of the receptor for the hormone and the reproductive parameters both in men and women (Aittomaki et al., 1995, Behre et al., 2005, Greb et al., 2005, Tüttelmann et al., 2012). On this basis, the current study group tested the hypothesis that FSHR sensitivity might be important in the response to FSH therapy and demonstrated that this treatment induces diverse improvement in seminal parameters according to defined FSHR genotypes; only subjects with at least one serine in position 680 (homozygotic AS/AS and heterozygotic TN/AS) had an improvement of seminal parameters (Selice et al., 2011). Again, molecular studies on the FSHB gene (coding for the β-subunit of FSH) showed that a G/T single-nucleotide polymorphism located in the FSHB gene promoter (−211 bp from the mRNA transcription start site; rs10835638) is responsible for the relative activity of endogenous FSH (Grigorova et al., 2007, Grigorova et al., 2008). Finally, research showed that oligozoospermic TT homozygotes for the FSHB −211 variant have a better response to FSH therapy in terms of sperm count and motility than patients with variants GG and GT (Ferlin et al., 2011).
Although the pharmacogenomic approach to male infertility seems to represent an important tool to predict the response to FSH therapy, it is still very expensive, time consuming in clinical situations and requires more and larger studies to determine tailored FSH dosage. This prospective controlled clinical study aimed to identify a new, simple, cheap and effective parameter to predict the response to FSH therapy in infertile oligozoospermic patients with normal FSH plasma concentrations in terms of sperm parameters and fertility outcome.
Section snippets
Patients and setting
This study was approved by the ethics committee of the Hospital-University of Padova (protocol number 2591P, approved 21 March 2010). Informed consent was obtained from each subject. All phases of the study were performed at the Infertility Centre of the University of Padova from January 2011 to December 2012. All men were informed about the FSH therapy protocol and in particular about the different phases of the study and the possibility of undergoing assisted reproduction treatment during
Results
Testicular cytology obtained by FNAC and seminal smears for spermatid count from patients in each group are shown in Figure 3.
Table 1 shows the baseline clinical, hormonal and seminal characteristics observed in 174 oligozoospermic patients divided into three groups on the basis of FNAC. There were 92 subjects who showed only hypospermatogenesis (group A), 61 who showed hypospermatogenesis with maturative disturbance (44 round and 17 elongated spermatidic arrest) (group B) and 21 who showed
Discussion
Many pathological conditions can lead to oligozoospermia, such as varicocele, cryptorchidism, orchitis, testicular trauma, karyotype abnormalities, Y chromosome microdeletions, cystic fibrosis gene abnormalities, triplet amplification or polymorphism or mutation in the LH or androgen receptor genes and idiopathic causes. Indeed, reduced sperm production may be associated with different testicular alterations, such as hypospermatogenesis and maturation disturbance at different levels, most
Acknowledgements
The authors thank Alberto Bottacin, Barbara Sartini and Laura Marino for technical assistance and all the staff of the Unit for Human Reproduction Pathology for helpful discussion. Special thanks to Karyn Ashley Cecchini for her kind and careful editing of English.
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To study the possible role of serum 17α-hydroxy-progesterone (17αOH-P) levels in predicting favorable responses to follicle-stimulating hormone (FSH) administration in patients with normal serum FSH levels and idiopathic abnormal sperm parameters.
Prospective cohort study.
University-affiliated fertility center.
Fifty patients with oligozoospermia, asthenozoospermia, and/or teratozoospermia and normal serum levels of gonadotropins and total testosterone (TT).
Treatment with exogenous FSH is administered subcutaneously at a dose of 150 IU 3 times a week for 3 consecutive months.
Luteinizing hormone levels, FSH levels, TT levels, 17αOH-P levels, testicular volume, conventional sperm parameters, and seminal spermatid concentration were evaluated before and after therapy. To evaluate the predictive role of pretreatment serum 17αOH-P levels on FSH responsiveness, the doubling of sperm concentration at the end of the FSH administration was considered a positive outcome.
After therapy, patients showed a significant increase in sperm concentration, total sperm count (TSC), progressive motility, percentage of normal forms, FSH levels, TT levels, and testicular volume. There was a negative correlation between pretreatment 17αOH-P levels and the posttreatment increase in sperm concentration, TSC, progressive motility, and normal morphology, and a positive correlation with the posttreatment increase in spermatids. Predictive analysis showed that 17αOH-P levels (<1.18 ng/mL) foretold a doubling of sperm concentration with a sensitivity of 90.0% and a specificity of 73.3%, and of TSC with a sensitivity of 91.3% and a specificity of 81.48%.
The results of this study suggest that pretreatment serum levels of 17αOH-P, a marker of steroidogenic function, appear to be able to predict the success of subcutaneous administration of exogenous FSH in terms of spermatogenesis improvement. Receiver operating characteristic curves indicated that 17αOH-P levels (<1.18 ng/mL) predict a doubling of sperm concentration and TSC after exogenous FSH administration to patients with idiopathic abnormal sperm parameters and normal gonadotropin levels.
Rol predictivo de los niveles séricos de 17 alfa hidroxi progesterona antes y despúes de la administración de hormona foliculo estimulante en pacientes con parámetros espermáticos anormales
Estudiar el posible rol de los niveles séricos de 17α-hidroxi-progesterona (17αOH-P) en la predicción de respuestas favorables a la administración de hormona folículo estimulante (FSH) en pacientes con niveles séricos normales de FSH y parámetros espermáticos anormales idiopáticos.
Estudio de corte prospectivo.
Centro de fertilidad afiliado a la universidad.
Cincuenta pacientes con oligozoospermia, astenozoospermia y niveles séricos normales de gonadotrofinas y testosterona total (TT).
Tratamiento con FSH exógena es administrada en forma subcutánea a una dosis de 150 UI 3 veces a la semana por 3 meses consecutivos.
antes y después del tratamiento, se evaluaron los niveles séricos de hormona luteinizante, FSH, TT, 17αOH-P, el volumen testicular, parámetros espermáticos convencionales y concentración de espermatozoides seminales. Evaluar el rol predictivo de los niveles séricos pre tratamiento de 17αOH-P sobre la capacidad de respuesta de la FSH, la duplicación de la concentración espermática y el fin de la administración de FSH fue considerado un resultado positivo.
Luego del tratamiento, los pacientes mostraron un aumento significativo de la concentración espermática, conteo espermático total (TSC), motilidad progresiva, porcentaje de formas normales, niveles de FSH, niveles de TT y volumen testicular. Hubo una correlación negativa entre niveles pretratamiento de 17αOH-P y el incremento postratamiento de la concentración espermática, TSC, motilidad progresiva, y morfología normal, y una correlación positiva con el incremento de espermatozoides post tratamiento. El análisis predictivo mostró que los niveles de 17αOH-P (menores 1.18 ng/ml) predecían una duplicación de la concentración espermática con una sensibilidad de 90% y una especificidad de 73.3%y una TSC con una sensibilidad de 91.3% y una especificidad de 81.48%.
Los resultados de este estudio sugieren que los niveles séricos pre tratamiento de 17αOH-P, un marcador de la función esteroideogénica, parecen capaces de predecir el éxito de la administración subcutánea de FSH exógena en términos de mejora de la espermatogénesis. Las curvas características de funcionamiento indicaron que los niveles de 17αOH-P (<1,18 ng/ml) predicen una duplicación de la concentración de espermatozoides y del TSC después de la administración de FSH exógena a pacientes con parámetros espermáticos anormales idiopáticos y niveles normales de gonadotropina.
2023, Best Practice and Research: Clinical Obstetrics and Gynaecology
Citation Excerpt :
According to the latest WHO manual for examining human semen, published in 2021, oligozoospermia is defined as a sperm concentration below the lower reference limit of 16 million/mL [1]. Several studies have explored the use of hormonal treatment with FSH for these patients (133,137-156). The characteristics of the most relevant trials published are summarized in Table 2.
Gonadotropin therapy to treat specific male infertility disorders associated with hypogonadotropic hypogonadism is evidence-based and effective in restoring spermatogenesis and fertility. In contrast, its use to improve fertility in men with idiopathic oligozoospermia or nonobstructive azoospermia remains controversial, despite being widely practiced. The existence of two major inter-related pathways for spermatogenesis, including FSH and intratesticular testosterone, provides a rationale for empiric hormone stimulation therapy in both eugonadal and hypogonadal males with idiopathic oligozoospermia or nonobstructive azoospermia. Real-world data (RWD) on gonadotropin stimulating for these patient subsets, mainly using human chorionic gonadotropin and follicle-stimulating hormone, accumulated gradually, showing a positive therapeutic effect in some patients, translated by increased sperm production, sperm quality, and sperm retrieval rates. Although more evidence is needed, current insights from RWD research indicate that selected male infertility patients might be managed more effectively using gonadotropin therapy, with potential gains for all parties involved.
Overall, 383 men with idiopathic infertility or with abnormal semen analyses were treated with FSH for a maximum of 3 months (Table 2). Recombinant FSH (rFSH) was used in three studies and urinary-derived FSH (uFSH) (Colacurci et al., 2012; Ruvolo et al., 2013; Simoni et al., 2016) in the other three (Garolla et al., 2014, 2017; Palomba et al. 2011) (Table 2). FSH administration significantly improved sDF in infertile men by 4.24% (95% CI: 0.24–8.25%; P = 0.04) after 3 months of treatment (Figure 4), although no sDF differences were observed between study and control groups both before and after FSH administration (Supplementary Figures 6A and 6B).
Conventional semen analyses have limitations in male infertility diagnosis and prognosis. Assessment of sperm DNA fragmentation (sDF) has been proposed to discriminate fertile from infertile men and predict FSH treatment response in infertile men, although a comprehensive evaluation of this is not available. The aims of these meta-analyses were to assess the power of sDF in male infertility diagnosis and its role in predicting FSH therapy response in infertile men. Two literature searches were conducted in MEDLINE (PubMed), Embase, the Cochrane Library, Scopus and UpToDate. First, interventional/observational clinical trials comparing fertile to infertile/subfertile men were included. Second, interventional/observational clinical trials evaluating FSH-treated infertile men were assessed. sDF levels were significantly higher in infertile men considering 28 studies (P < 0.001), independently of the sDF method applied. Receiver operator characteristics curves identified an sDF threshold of 20%, with sensitivity of 79% and specificity of 86%. Six studies showed significant sDF improvement of 4.24% (95% confidence interval: 0.24–8.25%) after 3 months of FSH treatment. These meta-analyses demonstrate the sDF relevance in male infertility, suggesting a higher accuracy in detecting sperm function than conventional semen parameters. Although larger prospective trials are needed, sDF represents a promising tool for clinical and research practice.
Carlo Foresta is full Professor of Endocrinology and Director of Human Reproduction Pathology Unit at the University of Padova, Italy. During the last 30 years his research activities included different aspects related to male reproduction and spermatogenesis. Current research and clinical interests are in the genetics of male infertility, cryptorchidism and testicular tumours, endocrine regulation of spermatogenesis, sexual transmittable diseases and sexual disorders. He is author of more than 250 papers in International peer-reviewed journals, and invited speaker to many international and national scientific congresses. He is Past President of the Italian Society of Andrology and Sexual Medicine and member of the executive board of many scientific societies.
