Effect of Letrozole on sperm parameters, chromatin status and ROS level in idiopathic Oligo/Astheno/Teratozoospermia.

BACKGROUND: This study investigates the effect of letrozole on hormone profiles, semen parameters, body mass index (BMI), degree of oxidative stress and sperm chromatin integrity in men with idiopathic oligo/astheno/teratozoospermia (iOAT) and T:E2 ratio ≤ 10. MATERIALS AND METHODS: This study is a longitudinal, prospective, interventional and open-labelled clinical trial. Semen samples were collected from 20 iOAT men with low serum testosterone (T) to estradiol (E2) ratio (T:E2 ratio ≤ 10). The participants were treated with 2.5 mg letrozole orally per day for 3 months. Then, sperm parameters, hormone profiles, BMI, chromatin integrity and intracellular reactive oxygen species (ROS) level were assessed pre- and post- treatment. The chromatin integrity was evaluated by assessment of DNA fragmentation (with TUNEL assay) and protamine deficiency (with Chromomycin A3, CMA3). Also, the intracellular ROS levels were investigated by 2', 7'-dichlorodihydrofluorescein diacetate (DCFH-DA) staining. Finally, the differences between the parameters evaluated before and after letrozole treatment were analyzed with the t-test and the Wilcoxon signed-rank test. RESULTS: Sperm concentration, percentage of sperm motility and its normal morphology increased significantly after letrozole treatment. Moreover, serum testosterone level increased but estradiol level decreased significantly following treatment. The mean of T:E2 ratio improved 1600%. Also, letrozole treatment significantly reduced the percentage of sperm TUNEL positivity and sperm CMA3 positivity. While no significant difference was observed between intracellular ROS levels and BMI before and after treatment. Finally, as a notable result, four spontaneous pregnancies (20%) were achieved after treatment. CONCLUSIONS: Letrozole treatment can effectively increase spontaneous pregnancies by improving sperm parameters and sperm chromatin integrity in men with iOAT and T:E2 ratio ≤ 10. TRIAL REGISTRATION: Trial registration: IRCT, IRCT20191030045283N1. Registered 16 November 2019 - Retrospectively registered, https://fa.irct.ir/user/trial/43484/view.

Does omitting teratospermia as a selection criterion for ICSI change pregnancy rates?

PURPOSE: There is controversy whether teratospermia is associated with poorer IVF outcomes and if ICSI may overcome this deficit. The debate likely lies in study heterogeneity, poor adjustment for confounders, and inter-observer variation in sperm morphology assessment. Given the current literature, a shift in practice was implemented at our center in February 2017, whereby teratospermia was no longer a criterion for ICSI. We hypothesized that, despite decreasing ICSI rates, we would see no change in ART outcomes. METHODS: A retrospective study was performed including 1821 couples undergoing IVF/ICSI at a single center from January 2016 to December 2018, divided into cohorts before and after the practice change. The primary outcome of clinical pregnancy and secondary outcomes of fertilization, fertilization failure, good quality blastocyst formation, embryo utilization, positive hCG, and miscarriage rates was compared, adjusting for potential confounders. Subgroup analysis was performed evaluating teratospermia as the only reason for a male factor infertility diagnosis. RESULTS: Despite a decrease in ICSI rate of 30.3%, we found no significant difference in clinical intrauterine pregnancy rate, with an adjusted relative risk of 0.93 (0.81, 1.07, P = 0.3008). There were no significant differences in other secondary outcomes after multivariate adjustment. Subgroup analysis for those with male factor infertility due to teratospermia showed no difference in outcomes. CONCLUSION: This study concurs with the recent data suggesting that employing ICSI solely for teratospermia is unnecessary. This may allow clinics to decrease ICSI rates without sacrificing success rates, leading to lower cost and risk associated with treatment.

Insight on multiple morphological abnormalities of sperm flagella in male infertility: what is new?

The syndrome of multiple morphological abnormalities of the sperm flagella (MMAF) is a specific kind of asthenoteratozoospermia with a mosaic of flagellar morphological abnormalities (absent, short, bent, coiled, and irregular flagella). MMAF was proposed in 2014 and has attracted increasing attention; however, it has not been clearly understood. In this review, we elucidate the definition of MMAF from a systematical view, the difference between MMAF and other conditions with asthenoteratozoospermia or asthenozoospermia (such as primary mitochondrial sheath defects and primary ciliary dyskinesia), the knowledge regarding its etiological mechanism and related genetic findings, and the clinical significance of MMAF for intracytoplasmic sperm injection and genetic counseling. This review provides the basic knowledge for MMAF and puts forward some suggestions for further investigations.

Is there any relationship between human sperm parameters and protamine deficiency in different groups of infertile men?

OBJECTIVE: Abnormality in Histone-Protamine replacements has been indicated to cause sperm DNA damage and infertility. The aim of the present study was to investigate the relationships between sperm parameters in oligospermia, asthenospermia, and teratospermia with protamine deficiency in infertile men. MATERIAL AND METHOD: In this case-control study, we had three experimental groups including oligospermia (n=100), asthenospermia (n=100), and teratospermia (n=100) as well as normospermia (n=100) as controls. Sperm analyses were performed according to the recommendations of the World Health Organization (WHO, 2010) and sperm chromatin quality was assessed using Chromomycin A3 (CMA3) staining for each sample. RESULTS: The comparison of the data between groups indicated that the percentage of spermatozoa with protamine deficiency was significantly different in patients with oligospermia, asthenospermia, and teratospermia when compared with control ones. However, there was no significant correlation between sperm nuclear protamine deficiency and their parameters of the men with teratospermia using CMA3 test. Regarding the oligospermia and asthenospermia semen samples, the findings showed the negative correlations between the sperm nuclear protamine deficiency and progressive motility as well as immobility (p<0.001). CONCLUSION: The higher proportion of spermatozoa with abnormal chromatin packaging was observed in asthenospermic samples than those from other experimental groups as well as controls. It seems that normal morphology cannot have a valuable predictive value for good chromatin quality of spermatozoa, as much as normal motility characteristics, since samples with high mobility rates often have lower protamine deficiencies. The findings may provide a supportable promoting the future wider clinical application of chromatin/DNA integrity testing along with the semen analysis in male infertility.

The association between sperm head elongation and semen quality.

BACKGROUND: Previous studies suggested that sperm head shape may serve as an effective indicator of semen quality. However, there lacks research with large sample and quantitative measurement. OBJECTIVES: The objective of this retrospective study was to explore the association between sperm head elongation (Width/Length ratio) and routine semen parameters. MATERIALS AND METHODS: From January 2012 to December 2017, 63 866 semen samples were collected from male subjects at 18-60 years of age. Sperm head elongation and routine semen parameters (semen volume, sperm concentration, motility, etc.) were examined with computer-assisted semen analysis (CASA) systems in order to evaluate the association between elongation and semen quality. RESULTS: Logistic and linear regression models showed that the value of elongation is negatively correlated with sperm concentration, total sperm count, progressive motility, total motility, percentage of morphologically normal spermatozoa, and acrosin activity (all p < 0.001). DISCUSSION: The results suggested that higher value of elongation is generally associated with higher risks of abnormality in semen quality. The importance of elongation may be explained by abnormal acrosin activity in the round-headed spermatozoa, which has been reported to cause failure of natural pregnancy. CONCLUSIONS: This study provides a new insight into the sperm head shape, which may be used as a complementary parameter in clinical semen examination and academic research.

Changes of Sperm Parameters Along Time Among Groups of Different Qualities.

BACKGROUND: Male infertility is solely responsible for approximately 20% of all infertility in couples. Various factors have been proposed as having a negative effect on sperm quality; however, the reasons for the global decline in sperm parameters during the last few decades are still controversial. OBJECTIVES: To investigate the fluctuations of semen parameters (sperm concentration, motility, and morphology) in three sperm quality groups and to examine the trends of those parameters in the same men over time. RESULTS: Our data showed deterioration in all semen parameters assessed in the group of men originally considered as having normal semen values according to the 2010 criteria of the World Health Organization. In contrast, we found significant improvement over time in all semen parameters in the group of men with severe oligo-terato-asthenozoospermia. CONCLUSIONS: Our results suggest that, although there were changes in sperm quality over time in the groups assessed, the clinical significance is negligible and does not necessarily justify a change in the therapeutic approach to infertility or sperm cryopreservation.

TSGA10 is a novel candidate gene associated with acephalic spermatozoa.

Acephalic spermatozoa is a rare teratozoospermia associated with male infertility. However, the pathogenesis of this disorder remains unclear. Here, we report a 27 years old infertile male from a consanguineous family, who presented with 99% headless sperm in his ejaculate. Electron microscopic and immunofluorescence analysis suggested breakage at the midpiece of the patient's sperm cells. Subsequent whole-exome sequencing analysis identified a homozygous deletion within TSGA10 (c.211delG; p.A71Hfs*12), which resulted in the production of truncated TSGA10 protein. TSGA10 is a testis-specific protein that localized to the midpiece in the spermatozoa of a normal control; however, immunostaining failed to detect TSGA10 protein in the patient's sperm. Western blot analysis also showed complete absence of TSGA10 protein in the patient. One cycle of in vitro fertilization-assisted reproduction was conducted, but pregnancy was not achieved after embryo transfer, possibly due to poor embryo quality. Therefore, we speculate that the presence of rare sequence variants within TSGA10 may be associated with acephalic spermatozoa in humans.

Fluorescence in situ hybridisation sperm examination is significantly impaired in all categories of male infertility.

To study the outcome of FISH sperm examination in cases with sperm pathology and outline the potential correlation with certain chromosomal defects. A retrospective study of prospectively collected data was performed in IAKENTRO, Infertility Treatment Center. Rates of abnormal FISH semen examination were compared between male infertility patients and fertile controls. Detection of abnormal FISH semen examination as well as each chromosomal abnormality detected was correlated with each sperm deficiency (asthenozoospermia, oligozoospermia and teratozoospermia) in a univariate regression model. There were 72 male partners included, of which 52 male infertility patients and 20 controls. The rate of abnormal sperm FISH examination was significantly higher in patients' group (55.8% vs. 15.0% for controls, p = .002). Asthenozoospermia, oligozoospermia and teratozoospermia were significantly correlated with detection of abnormal FISH examination (p = .004, p = .01 and p < .001 respectively). Teratospermia was significantly correlated with increased aneuploidy rate for chromosome 17 (p = .005), chromosome X (p = .05) and Y (p = .03). FISH examination reveals pathology in a significant proportion of patients with sperm defects and should be recommended to achieve early detection of chromosomal defects that may postpone favourable reproductive outcome.

Disruption of Ssp411 causes impaired sperm head formation and male sterility in mice.

BACKGROUND: We previously cloned the Ssp411 gene. We found that the Ssp411 protein is predominantly expressed in elongated spermatids in the rat testis in a stage-dependent manner. Although our findings strongly suggested that Ssp411 might play an important role in mammalian spermatogenesis, this hypothesis has not been studied. METHODS: We first used real-time PCR, Western blotting and immunohistochemistry to confirm that the expression pattern of Ssp411 in several murine tissues is similar to its expression pattern in corresponding rat tissues. To better understand the roles of Ssp411 in male reproduction in vivo, we identified and characterized an Ssp411 expression-disrupted murine strain (Ssp411PB/PB) that was generated by piggyBac (PB) transposon insertion. We studied Ssp411-interacting proteins using proteome microarray, co-IP and GST pull-down assay. RESULTS: Both Ssp411 mRNA and protein were detected exclusively in spermatids after step 9 during spermiogenesis in testis. Phenotypic analysis suggested that only Ssp411PB/PB males are sterile. These males have smaller testes, reduced sperm counts, decreased sperm motility and deformed spermatozoa. Microscopy analysis indicated that the manchette, a structurally reshaped sperm head, is aberrant in Ssp411PB/PB spermatids. The results of proteome microarray analysis and GST pull-down assays suggested that Ssp411 participates the ubiquitin-proteasome system by interacting with PSMC3. This has been reported to be manchette-associated and important for the head shaping of spermatids. CONCLUSIONS: Our study suggested that Ssp411 is required for spermiogenesis. It seems to play a role in sperm head shaping. The lack of Ssp411 causes sperm deformation and results in male infertility. GENERAL SIGNIFICANCE: Ssp411PB/PB mouse strain is an animal model of idiopathic oligoasthenoteratozoospermia (iOAT), and the gene may represent a therapeutic target for iOAT patients.

Comprehensive functional enrichment analysis of male infertility.

Spermatogenesis is a multifactorial process that forms differentiated sperm cells in a complex microenvironment. This process involves the genome, epigenome, transcriptome, and proteome to ensure the stability of the spermatogonia and supporting cells. The identification of signaling pathways linked to infertility has been hampered by the inherent complexity and multifactorial aspects of spermatogenesis. Systems biology is a promising approach to unveil underlying signaling pathways and genes and identify putative biomarkers. In this study, we analyzed thirteen microarray libraries of infertile humans and mice, and different classes of male infertility were compared using differentially expressed genes and functional enrichment analysis. We found regulatory processes, immune response, glutathione transferase and muscle tissue development to be among the most common biological processes in up-regulated genes, and genes involved in spermatogenesis were down-regulated in maturation arrest (MArrest) and oligospermia cases. We also observed the overexpression of genes involved in steroid metabolism in post-meiotic and meiotic arrest. Furthermore, we found that the infertile mouse model most similar to human MArrest was the Dazap1 mutant mouse. The results of this study could help elucidate features of infertility etiology and provide the basis for diagnostic markers.

Oxidative damage of the male reproductive system induced by paraquat.

The effects of paraquat (PQ) on the male reproductive system are unclear. In this study, male rats were divided into four groups (0, 0.5, 2, and 8 mg/kg) and treated with PQ by oral gavage for 8 weeks. At the end of the experiment, a significant decline in sperm count, motility, and viability and an increase in teratospermia were observed in the PQ-treated group (P < 0.05). Further investigation found that PQ resulted in enhanced lipid peroxidation and more apoptosis in the testis tissues, and apoptosis was likely to be associated with activation of the mitochondrial pathway. In summary, our study demonstrated oxidative damage due to PQ on the male reproductive system.

Men with a complete absence of normal sperm morphology exhibit high rates of success without assisted reproduction.

In couples with infertility, abnormal strict morphology of 0% normal forms (NF) is a criterion to proceed rapidly to in vitro fertilization (IVF). Since no data currently exist, we investigated the outcomes for men with 0% NF to determine reproductive success without the use of assisted reproductive technologies (ART). A cohort of 24 men with 0% NF were identified (2010-2013) with 27 randomly selected men with ≥4% NF as controls. Patient charts were reviewed with men contacted and administered an Institutional Review Board (IRB)-approved telephone questionnaire to ascertain outcomes. After a median follow-up time of 2.5 years, 29.2% of men with 0% NF did not require ART for their first pregnancy (controls = 55.6%, P ≤ 0.05). When all pregnancies were analyzed together, men with 0% NF achieved twenty pregnancies of which 75% did not require IVF (controls = thirty pregnancies; 76.7% did not require IVF). The average age of men and female partners was similar between men with 0% NF and ≥4% NF. All men had normal follicle-stimulating hormone (FSH), testosterone, prolactin, sex hormone-binding globulin (SHBG), and estradiol. Although, global semen parameters were worse in men with 0% NF, when a first pregnancy was a natural conception (NC), 100% of men with 0% NF (n = 7/7) and 37.5% of controls (n = 3/8) went on to have a subsequent pregnancy via NC. Men with 0% NF conceived without IVF in 29.2% of cases compared to 55.6% of controls. Strict morphology should not be used to predict fertilization, pregnancy, or live birth potential. In men with 0% NF, alternative modalities should be considered before immediate IVF.

Normozoospermic versus teratozoospermic domestic cats: differential testicular volume, sperm morphometry, and subpopulation structure during epididymal maturation.

Teratozoospermia (<40% morphologically normal spermatozoa/ejaculate) is a frequent phenomenon in feline species. This research was carried out to study the possible differences in testicular volume, differential sperm morphometric traits, and potential differences regarding the sperm subpopulational structure during epididymal sperm maturation in teratozoospermic feline donors. Epididymal sperm samples were collected from the caput (R1), corpus (R2), and cauda (R3) epididymidis in two donor groups (N: normozoospermic; T: teratozoospermic). Aliquots were assessed for concentration, viability, motility, and acrosomal integrity. Sperm morphometric descriptors from CASA-Morph analysis were analyzed by the Principal Component Analysis (PCA) and clustering analyses. Irrespective of the group analyzed, PCA revealed two Principal Components (PCs) for each epididymal region explaining more than the 93% of the variance. Surprisingly, the number of subpopulations remained constant in regions R1-R2-R3 irrespective of the donor group analyzed. However, the distribution of these subpopulations was found to be structurally different and strongly influenced by the epididymal region and the donor group. In conclusion, testicular morphometry and the sperm subpopulation structure were different in N and T donors. The alterations in subpopulations during epididymal maturation could be used as a potential clinical indicator of teratozoospermic individuals since an important influence of teratozoospermia on sperm subpopulation structure has been demonstrated.

[Stimulation of a spermatogenesis in men at a pathospermia and infertility].

THE PURPOSE: to investigate of the methods of treatment, directed on increase in quantity of spermatozoa in an ejaculate. MATHERIALS AND METHODS: for this purpose used clomifene and combinations of recombinant FSH with chorionic gonadotrophin (HCG) in 60 men with infertility. RESULTS: Efficiency of monotherapy by clomiphene was higher and made 20% for conception, and 63% for oligoteratozoospermia. Efficiency of the combined therapy of HCG in combination with recombinant FSH was 40% for conception, and 87% for oligoteratozoospermia. The efficiency of the combined therapy by recombinant FSH and HCG in cases of the previous inefficiency monotherapy HCG and clomifene for oligoteratozoospermia made 65%. SUMMARY: The combined therapy of HCG in combination with recombinant FSH is al most effective. At the same time the studied types of the stimulating therapy are safe and don't lead to development of side effects.