Resolving Male Infertility: A Case Report on Treating Obstructive Azoospermia Using SpermMobil in Intracytoplasmic Sperm Injection Procedure.

Infertility affects couples worldwide. Among these, obstructive azoospermia (OA) is a common cause. In some cases, the lack of spermatozoa in ejaculation results from blockages in the male reproductive tract. In this case study, we discuss an infertile male diagnosed with OA following three years of unsuccessful attempts at conception. The male had a history of bilateral inguinal hernia repair due to congenital bilateral absence of the vas deferens. Diagnostic assessments confirmed azoospermia. Microscopic epididymal sperm aspiration (MESA) was performed for sperm retrieval due to its efficacy and reduced postoperative pain, testicular atrophy, and decreased testosterone levels. The retrieved sperm was processed using SpermMobil media for intracytoplasmic sperm injection. Following successful fertilization, embryo transfers resulted in a positive pregnancy test. This case highlights the significance of specific treatment approaches for OA, specifically the effectiveness of MESA and SpermMobil in achieving successful outcomes in assisted reproduction technology for male infertility.

The human infertility single-cell testis atlas (HISTA): an interactive molecular scRNA-Seq reference of the human testis.

BACKGROUND: Single-cell RNA-seq (scRNA-Seq) has been widely adopted to study gene expression of the human testis. Several datasets of scRNA-Seq from human testis have been generated from different groups processed with different informatics pipelines. An integrated atlas of scRNA-Seq expression constructed from multiple donors, developmental ages, and fertility states would be widely useful for the testis research community. OBJECTIVE: To describe the generation and use of the human infertility single-cell testis atlas (HISTA), an interactive web tool for understanding human spermatogenesis through scRNA-Seq analysis. METHODS: We obtained scRNA-Seq datasets derived from 12 donors, including healthy adult controls, juveniles, and several infertility cases, and reprocessed these data using methods to remove batch effects. Using Shiny, an open-source environment for data visualization, we created numerous interactive tools for exploring the data, some of which support simple statistical hypothesis testing. We used the resulting HISTA browser and its underlying data to demonstrate HISTA's value for testis researchers. RESULTS: A primary application of HISTA is to search by a single gene or a set of genes; thus, we present various analyses that quantify and visualize gene expression across the testis cells and pathology. HISTA also contains machine-learning-derived gene modules ("components") that capture the entire transcriptional landscape of the testis tissue. We show how the use of these components can simplify the highly complex data in HISTA and assist with the interpretation of genes with unknown functions. Finally, we demonstrate the diverse ways HISTA can be used for new data analysis, including hypothesis testing. DISCUSSION AND CONCLUSIONS: HISTA is a research environment that can help scientists organize and understand the high-dimensional transcriptional landscape of the human testis. HISTA has already contributed to published testis research and can be updated as needed with input from the research community or downloaded and modified for individual needs.

A novel cancer-germline gene DAZL promotes progression and cisplatin resistance of non-small cell lung cancer by upregulating JAK2 and MCM8.

Germline-specific genes are usually activated in cancer cells and drive cancer progression; such genes are called cancer-germline or cancer-testis genes. The RNA-binding protein DAZL is predominantly expressed in germ cells and plays a role in gametogenesis as a translational activator or repressor. However, its expression and role in non-small cell lung cancer (NSCLC) are unknown. Here, mining of RNA-sequencing data from public resources and immunohistochemical analysis of tissue microarrays showed that DAZL was expressed exclusively in testis among normal human tissues but ectopically expressed in NSCLC tissues. Testis and NSCLC cells expressed the shorter and longer transcript variants of the DAZL gene, respectively. Overexpression of the longer DAZL transcript promoted tumor growth in a mouse xenograft model. Silencing of DAZL suppressed cell proliferation, colony formation, migration, invasion, and cisplatin resistance in vitro and tumor growth in vivo. Quantitative proteomic analysis based on tandem mass tag and Western blot analysis showed that DAZL upregulated the expression of JAK2 and MCM8. RNA-binding protein immunoprecipitation assays showed that DAZL bound to the mRNA of JAK2 and MCM8. The JAK2 inhibitor fedratinib attenuated the oncogenic outcomes induced by DAZL overexpression, whereas silencing MCM8 counteracted the effects of DAZL overexpression on cisplatin-damaged DNA synthesis and half-maximal inhibitory concentration of cisplatin. In conclusion, DAZL was identified as a novel cancer-germline gene that enhances the translation of JAK2 and MCM8 to promote NSCLC progression and resistance to cisplatin, respectively. These findings suggest that DAZL is a potential therapeutic target in NSCLC.

Mesenchymal Stem Cells in Regenerative Medicine, Possible Applications in The Restoration of Spermatogenesis: A Review.

Infertility is a common clinical condition and about half of the major causes are due to male-related infertility. Pathogenesis of this abnormality is generally undefined; so establishing a proper treatment option is relatively uncertain. In recent years, several evidences demonstrated that mesenchymal stem cells (MSCs) can be a hope for innovative and efficient treatment of male infertility. This study reviews possible applications of MSCs in the restoration of spermatogenesis in male infertility of both humans and animals to suggest new avenues for future clinical practices. Articles published in "PubMed" and "Google Scholar" from January 1, 2000, to August 1, 2023, were investigated by searching items of "mesenchymal stem cells", "cell therapy", "cell transplantation", and, "regenerative medicine" keywords, in addition to the "urology", "andrology", "reproductive medicine", "male infertility", "azoospermia", and "spermatogenesis". The results obtained from the transplantation of MSCs in the treatment of male infertility seemed encouraging and they revealed the safety and efficacy of these cells to recover spermatogenesis; eventhough further stem cell research is still required before recruiting clinical application of MSCs in the treatment of human male infertility. Undertaking more well-defined, standardized, and reproducible protocols and enrolling larger sample sizes during a longer follow-up period can benefit the relevance of MSC transplantation in the restoration of spermatogenesis and treatment of male infertility. It seems that developing and utilizing stem cell transplantations, exosomes, scaffold delivery systems, and three dimensional (3D) culture methods may open a new window to getting more benefits from cell therapy in the treatment of men infertility.

Safety evaluation of single-sperm cryopreservation technique applied in intracytoplasmic sperm injection.

Intracytoplasmic sperm injection (ICSI) is a technique that directly injects a single sperm into the cytoplasm of mature oocytes. Here, we explored the safety of single-sperm cryopreservation applied in ICSI. This retrospective study enrolled 186 couples undergoing ICSI-assisted pregnancy. Subjects were allocated to the fresh sperm (group A)/single-sperm cryopreservation (group B) groups based on sperm type, with their clinical baseline/pathological data documented. We used ICSI-compliant sperm for subsequent in vitro fertilization and followed up on all subjects. The recovery rate/cryosurvival rate/sperm motility of both groups, the pregnancy/outcome of women receiving embryo transfer, and the delivery mode/neonatal-related information of women with successful deliveries were recorded. The clinical pregnancy rate, cumulative clinical pregnancy rate, abortion rate, ectopic pregnancy rate, premature delivery rate, live birth delivery rate, neonatal birth defect rate, and average birth weight were analyzed. The two groups showed no significant differences in age, body mass index, ovulation induction regimen, sex hormone [anti-Müllerian hormone (AMH)/follicle-stimulating hormone (FSH)/luteinizing hormone (LH)] levels, or oocyte retrieval cycles. The sperm recovery rate (51.72%-100.00%) and resuscitation rate (62.09% ± 16.67%) in group B were higher; the sperm motility in the two groups demonstrated no significant difference and met the ICSI requirements. Group B exhibited an increased fertilization rate, decreased abortion rate, and increased safety versus group A. Compared with fresh sperm, the application of single-sperm cryopreservation in ICSI sensibly improved the fertilization rate and reduced the abortion rate, showing higher safety.

The relationship of testicular stiffness with Johnsen score and sperm retrieval outcome in men with non-obstructive azoospermia.

Background: The pathological Johnsen score (JS) is a quantitative histological scoring system used to assess spermatogenesis in men with nonobstructive azoospermia (NOA), while elastic modulus derived from shear wave elastography (SWE) is a diagnostic tool for evaluating spermatogenic dysfunction. In this prospective observational study, we aimed to investigate whether testicular stiffness measured by SWE could serve as a substitute for JS in predicting sperm retrieval outcomes in men with NOA. Methods: This prospective cohort study analyzed 140 testes from 115 consecutive outpatient participants with NOA who had sought treatment at the reproductive medical center of a tertiary care hospital between January 2018 and October 2021. Testicular volume, elastic modulus, JS, and sperm retrieval outcomes were calculated. Statistical differences in parameters between the positive and negative sperm retrieval groups were determined using the Mann-Whitney test. Spearman rank correlation analysis was performed to determine the correlations between JS and either testicular volumes or elastic modulus. Receiver operating characteristic (ROC) curves were drawn to evaluate the diagnostic performance of the testicular elastic modulus and testicular volume. Results: The JS correlated positively with testicular volume and negatively with the maximum elastic modulus (Emax) and mean elastic modulus (Emean), with correlation coefficients of 0.804, -0.686, and -0.456, respectively (P<0.01). There were significant differences in JS, testicular volume, and Emax between participants with positive and negative sperm retrieval of microdissection testicular sperm extraction (micro-TESE) (P<0.01). ROC curves were plotted for JS, testicular volume, and Emax to distinguish between participants with positive and negative sperm retrieval. The areas under the ROC curves (AUCs) were 0.783 [95% confidence interval (CI): 0.707-0.859; P<0.01], 0.737 (95% CI: 0.651-0.823; P<0.01), and 0.729 (95% CI: 0.643-0.814; P<0.01), respectively. When the cutoff value of JS was 4.5, its sensitivity and specificity were 60.3% and 89.6%, respectively. When the cutoff value of Emax was 3.75 kPa, its sensitivity and specificity were 79.1% and 64.4%, respectively. The sensitivity and specificity were 68.5% and 83.6%, respectively when the cutoff value of testicular volume was 8.17 mL. Emax combined with testicular volume improved this diagnostic value, with an AUC of 0.742 (95% CI: 0.657-0.828; P<0.01), and sensitivity and specificity were 83.6% and 68.5%, respectively. Conclusions: Our study suggests that the combination of testicular stiffness and volume measurements may serve as a viable alternative to pathological JS in predicting the likelihood of successful sperm retrieval prior to micro-TESE procedures.

Global Practice Patterns in the Evaluation of Non-Obstructive Azoospermia: Results of a World-Wide Survey and Expert Recommendations.

PURPOSE: Non-obstructive azoospermia (NOA) represents the persistent absence of sperm in ejaculate without obstruction, stemming from diverse disease processes. This survey explores global practices in NOA diagnosis, comparing them with guidelines and offering expert recommendations. MATERIALS AND METHODS: A 56-item questionnaire survey on NOA diagnosis and management was conducted globally from July to September 2022. This paper focuses on part 1, evaluating NOA diagnosis. Data from 367 participants across 49 countries were analyzed descriptively, with a Delphi process used for expert recommendations. RESULTS: Of 336 eligible responses, most participants were experienced attending physicians (70.93%). To diagnose azoospermia definitively, 81.7% requested two semen samples. Commonly ordered hormone tests included serum follicle-stimulating hormone (FSH) (97.0%), total testosterone (92.9%), and luteinizing hormone (86.9%). Genetic testing was requested by 66.6%, with karyotype analysis (86.2%) and Y chromosome microdeletions (88.3%) prevalent. Diagnostic testicular biopsy, distinguishing obstructive azoospermia (OA) from NOA, was not performed by 45.1%, while 34.6% did it selectively. Differentiation relied on physical examination (76.1%), serum hormone profiles (69.6%), and semen tests (68.1%). Expectations of finding sperm surgically were higher in men with normal FSH, larger testes, and a history of sperm in ejaculate. CONCLUSIONS: This expert survey, encompassing 367 participants from 49 countries, unveils congruence with recommended guidelines in NOA diagnosis. However, noteworthy disparities in practices suggest a need for evidence-based, international consensus guidelines to standardize NOA evaluation, addressing existing gaps in professional recommendations.

Evaluation of an artificial intelligence-facilitated sperm detection tool in azoospermic samples for use in ICSI.

RESEARCH QUESTION: Can artificial intelligence (AI) improve the efficiency and efficacy of sperm searches in azoospermic samples? DESIGN: This two-phase proof-of-concept study began with a training phase using eight azoospermic patients (>10,000 sperm images) to provide a variety of surgically collected samples for sperm morphology and debris variation to train a convolutional neural network to identify spermatozoa. Second, side-by-side testing was undertaken on two cohorts of non-obstructive azoospermia patient samples: an embryologist versus the AI identifying all the spermatozoa in the still images (cohort 1, n = 4), and a side-by-side test with a simulated clinical deployment of the AI model with an intracytoplasmic sperm injection microscope and the embryologist performing a search with and without the aid of the AI (cohort 2, n = 4). RESULTS: In cohort 1, the AI model showed an improvement in the time taken to identify all the spermatozoa per field of view (0.02 ± 0.30  ×  10-5s versus 36.10 ± 1.18s, P < 0.0001) and improved recall (91.95 ± 0.81% versus 86.52 ± 1.34%, P < 0.001) compared with an embryologist. From a total of 2660 spermatozoa to find in all the samples combined, 1937 were found by an embryologist and 1997 were found by the AI in less than 1000th of the time. In cohort 2, the AI-aided embryologist took significantly less time per droplet (98.90 ± 3.19 s versus 168.7 ± 7.84 s, P < 0.0001) and found 1396 spermatozoa, while 1274 were found without AI, although no significant difference was observed. CONCLUSIONS: AI-powered image analysis has the potential for seamless integration into laboratory workflows, to reduce the time to identify and isolate spermatozoa from surgical sperm samples from hours to minutes, thus increasing success rates from these treatments.

Toward clinical exomes in diagnostics and management of male infertility.

Infertility, affecting ∼10% of men, is predominantly caused by primary spermatogenic failure (SPGF). We screened likely pathogenic and pathogenic (LP/P) variants in 638 candidate genes for male infertility in 521 individuals presenting idiopathic SPGF and 323 normozoospermic men in the ESTAND cohort. Molecular diagnosis was reached for 64 men with SPGF (12%), with findings in 39 genes (6%). The yield did not differ significantly between the subgroups with azoospermia (20/185, 11%), oligozoospermia (18/181, 10%), and primary cryptorchidism with SPGF (26/155, 17%). Notably, 19 of 64 LP/P variants (30%) identified in 28 subjects represented recurrent findings in this study and/or with other male infertility cohorts. NR5A1 was the most frequently affected gene, with seven LP/P variants in six SPGF-affected men and two normozoospermic men. The link to SPGF was validated for recently proposed candidate genes ACTRT1, ASZ1, GLUD2, GREB1L, LEO1, RBM5, ROS1, and TGIF2LY. Heterozygous truncating variants in BNC1, reported in female infertility, emerged as plausible causes of severe oligozoospermia. Data suggested that several infertile men may present congenital conditions with less pronounced or pleiotropic phenotypes affecting the development and function of the reproductive system. Genes regulating the hypothalamic-pituitary-gonadal axis were affected in >30% of subjects with LP/P variants. Six individuals had more than one LP/P variant, including five with two findings from the gene panel. A 4-fold increased prevalence of cancer was observed in men with genetic infertility compared to the general male population (8% vs. 2%; p = 4.4 × 10-3). Expanding genetic testing in andrology will contribute to the multidisciplinary management of SPGF.

Global Practice Patterns and Variations in the Medical and Surgical Management of Non-Obstructive Azoospermia: Results of a World-Wide Survey, Guidelines and Expert Recommendations.

PURPOSE: Non-obstructive azoospermia (NOA) is a common, but complex problem, with multiple therapeutic options and a lack of clear guidelines. Hence, there is considerable controversy and marked variation in the management of NOA. This survey evaluates contemporary global practices related to medical and surgical management for patients with NOA. MATERIALS AND METHODS: A 56-question online survey covering various aspects of the evaluation and management of NOA was sent to specialists around the globe. This paper analyzes the results of the second half of the survey dealing with the management of NOA. Results have been compared to current guidelines, and expert recommendations have been provided using a Delphi process. RESULTS: Participants from 49 countries submitted 336 valid responses. Hormonal therapy for 3 to 6 months was suggested before surgical sperm retrieval (SSR) by 29.6% and 23.6% of participants for normogonadotropic hypogonadism and hypergonadotropic hypogonadism respectively. The SSR rate was reported as 50.0% by 26.0% to 50.0% of participants. Interestingly, 46.0% reported successful SSR in <10% of men with Klinefelter syndrome and 41.3% routinely recommended preimplantation genetic testing. Varicocele repair prior to SSR is recommended by 57.7%. Half of the respondents (57.4%) reported using ultrasound to identify the most vascularized areas in the testis for SSR. One-third proceed directly to microdissection testicular sperm extraction (mTESE) in every case of NOA while others use a staged approach. After a failed conventional TESE, 23.8% wait for 3 months, while 33.1% wait for 6 months before proceeding to mTESE. The cut-off of follicle-stimulating hormone for positive SSR was reported to be 12-19 IU/mL by 22.5% of participants and 20-40 IU/mL by 27.8%, while 31.8% reported no upper limit. CONCLUSIONS: This is the largest survey to date on the real-world medical and surgical management of NOA by reproductive experts. It demonstrates a diverse practice pattern and highlights the need for evidence-based international consensus guidelines.

Proteomic biomarkers in seminal plasma as predictors of reproductive potential in azoospermic men.

Introduction: Azoospermia, characterized by an absence of sperm in the ejaculate, represents the most severe form of male infertility. While surgical sperm retrieval in obstructive azoospermia (OA) is successful in the majority of cases, patients with non-obstructive azoospermia (NOA) show retrieval rates of only about 50% and thus frequently have unnecessary surgery. Surgical intervention could be avoided if patients without preserved spermatogenesis are identified preoperatively. This prospective study aimed to discover biomarkers in seminal plasma that could be employed for a non-invasive differential diagnosis of OA/NOA in order to rationalize surgery recommendations and improve success rates. Methods: All patients signed written informed consent, underwent comprehensive andrological evaluation, received human genetics to exclude relevant pathologies, and patients with azoospermia underwent surgical sperm retrieval. Using label-free LC-MS/MS, we compared the proteomes of seminal plasma samples from fertile men (healthy controls (HC), n=8) and infertile men diagnosed with 1) OA (n=7), 2) NOA with successful sperm retrieval (mixed testicular atrophy (MTA), n=8), and 3) NOA without sperm retrieval (Sertoli cell-only phenotype (SCO), n=7). Relative abundance changes of two candidate markers of sperm retrieval, HSPA2 and LDHC, were confirmed by Western Blot. Results: We found the protein expression levels of 42 proteins to be significantly down-regulated (p ≤ 0.05) in seminal plasma from SCO NOA patients relative to HC whereas only one protein was down-regulated in seminal plasma from MTA patients. Analysis of tissue and cell expression suggested that the testis-specific proteins LDHC, PGK2, DPEP3, and germ-cell enriched heat-shock proteins HSPA2 and HSPA4L are promising biomarkers of spermatogenic function. Western blotting revealed a significantly lower abundance of LDHC and HSPA2 in the seminal plasma of men with NOA (SCO and MTA) compared to controls. Discussion: The results indicate that certain testis-specific proteins when measured in seminal plasma, could serve as indicators of the presence of sperm in the testis and predict the success of sperm retrieval. Used in conjunction with conventional clinical assessments, these proteomic biomarkers may assist in the non-invasive diagnosis of idiopathic male infertility.

Establishment and validation of a 5-factor diagnostic model for obstructive and non-obstructive azoospermia based on routine clinical parameters.

Azoospermia is a serious leading male-factor cause of infertility in couples of childbearing age. The two main azoospermia types, obstructive (OA) and non-obstructive (NOA) azoospermia, differ in their treatment approaches. Therefore, their clinical diagnosis is extremely important, requiring an accurate, efficient, and easy-to-use diagnostic model. This retrospective observational study included 707 patients with azoospermia treated between 2017 and 2021, 498 with OA, and 209 with NOA. Hematological and seminal plasma parameters, hormone levels, and testicular volume were used in logistic regression analysis to evaluate and compare their diagnostic performance, results showed that the optimal diagnostic model is constructed by five variables including semen volume, semen pH, seminal plasma neutral α-glucosidase activity, follicle-stimulating hormone in the serum, and testicular volume, compared with follicle-stimulating hormone-based and testicular volume-based models. The 5-factor diagnostic model had an accuracy of 90.4%, sensitivity of 96.4%, positive predictive value of 90.6%, negative predictive value of 89.8%, and area under the curve of 0.931, all higher than in the other two models. However, its specificity (76.1%) was slightly lower than in the other models. Meantime, the internal 5-fold cross-validation results indicated that the 5-factor diagnostic model had a good clinical application value. This study established an accurate, efficient, and relatively accessible 5-factor diagnostic model for OA and NOA, providing a reference for clinical decision-making when selecting an appropriate treatment.

[Tumor necrosis factor in the ejaculate as an indicator of reduced fertility].

INTRODUCTION: Pro-inflammatory cytokine - tumor necrosis factor-alpha (TNF) is one of the components of the seminal plasma proteome; its meaning has not been definitively revealed. A comparative analysis of the concentration of this protein in the blood serum and in the ejaculate and changes in its level in the semen of men with infertility is f scientific interest. THE PURPOSE OF THE STUDY: determination of TNF- level in the blood serum and seminal plasma of healthy men and patients with reduced fertility. MATERIALS AND METHODS: 70 men of reproductive age with azoospermia (main group, n=18), with oligoastenozoospermia (comparison group, n=18) and with normal spermogram parameters (control group, n=34) were examined. The ejaculate was examined using an SQA-V semen analyzer (MES, Israel). In seminal plasma samples, the concentration of TNF was determined using the alpha-TNF-ELISA-BEST test system (A-8756, Vector-Best LL, Russia). RESULTS: The concentration of TNF- in blood serum had a significant variation (CV=85.31%) and amounted to 2.75+/-2.18 pg/ml, which is 2.55 times lower than the same indicator in seminal plasma (7.01+/-5.98 pg/ml, CV=126.15%, p<0.00001). When comparing the content of TNF- in seminal plasma, significant differences were found in the examined patients (Kruskal-Wallis test H=24.75991; p<0.00001). Pairwise comparison revealed a statistically significant difference in the level of TNF- in seminal plasma between the comparison and control groups (p2-3=0.000023), as well as between the main group and the comparison group (p1-2=0.000043); there were no significant differences between the main and control groups (p>0.05). When determining the content of TNF- in the blood serum, there was no statistically significant difference between the groups (p>0.05). There were no correlations between the concentration of TNF- in blood serum and in seminal plasma (R=0.295374), and the total number of spermatozoa in the ejaculate (R=-0.027945); and the concentration of spermatozoa in the ejaculate (R=-0.042902). DISCUSSION: It is unlikely that TNF crosses into seminal plasma from serum against a concentration gradient. It is most likely that TNF is produced locally in the organs of the reproductive system by resident immune cells or cells involved in spermatogenesis. An increased content of TNF- in seminal plasma in patients of the comparison group may indicate the presence of an inflammatory process in the reproductive system and a reduced fertility of the ejaculate. CONCLUSION: The physiological role of TNF in sperm, its sources in the organs of the male reproductive system, and the pathogenetic mechanisms of the participation of the TNF in pathological processes in male reproductive system still remain unclear. All this justifies the need for further study of the TNF level in seminal plasma in normal conditions and in diseases of the urogenital tract in men.

Association of the minimal cyclophosphamide equivalent dose and outcome of microdissection testicular sperm extraction in patients with persistent azoospermia after chemotherapy.

Objective: To investigate whether the minimal cyclophosphamide equivalent dose (mCED), a novel approach for estimating alkylating agent exposure, is associated with the sperm retrieval rates by microdissection testicular sperm extraction (mTESE) in azoospermic postchemotherapy cancer survivors. Design: A retrospective cohort study conducted between 2002 and 2017. Setting: An academic medical center. Patients: A total of 28 azoospermic postchemotherapy cancer survivors who underwent mTESE. Interventions: Chemotherapy exposure and mCED calculation. Main Outcome Measures: The primary outcome was the association between the mCED and sperm retrieval rate using mTESE. The mCED value for each patient's regimen received was estimated using the lowest recommended dosing regimen from the range of recommended doses at the time of administration. Results: Spermatozoa were successfully retrieved in 11 (39.3%) of the patients. Age at the time of receiving chemotherapy and mCED were significant factors associated with sperm retrieval. An mCED of <4,000 mg/m2 had a higher sperm retrieval rate (10/14, 71.4%) than an mCED of >4,000 mg/m2 (0/8, 0). The hormone levels were not significantly different when comparing patients with and without successful sperm retrieval. Seminoma, nonseminomatous germ cell tumor, and acute lymphoblastic leukemia had favorable sperm retrieval rates-100% (2/2), 66.7% (2/3), and 66.7% (2/3), respectively-although the numbers of patients in each group were small. Conclusion: Among this cohort of patients with cancer who required chemotherapy regimens, successful sperm retrieval by mTESE was only noted among cancer survivors receiving an mCED of <4,000 mg/m2.

Prevalence and clinical implications of biochemical hypogonadism in patients with nonobstructive azoospermia undergoing infertility evaluation.

Objective: To investigate the prevalence and clinical implications of biochemical hypogonadism in infertile men with nonobstructive azoospermia (NOA). Design: Cohort study. Setting: University-affiliated tertiary center for male reproductive health. Patients: 767 consecutive normogonadotropic or hypergonadotropic patients with NOA undergoing infertility evaluation from 2014 to 2021. Intervention: Patients aged 23-55 years underwent comprehensive clinical, hormonal, genetic, semen analysis, and histopathology evaluations and were classified on the basis of predefined baseline follicle-stimulating hormone (12 IU/L) and total testosterone (350 ng/dL) serum levels cutpoints into four groups: hypergonadotropic hypogonadal, hypergonadotropic eugonadal, normogonadotropic hypogonadal, and normogonadotropic eugonadal. All patients were naïve regarding previous sperm retrieval (SR) or hormonal therapy use. Main Outcome Measures: The period prevalence of biochemical hypogonadism, defined as testosterone levels of <350 ng/dL, and the distribution of patients per group were computed. The associations between hypogonadism, clinical factors, and SR success were evaluated using multivariable logistic regression analyses. Adjusted relative risks (aRRs) and 95% confidence intervals (CIs) were estimated to assess the association between SR and patient classification. Results: The overall period prevalence of biochemical hypogonadism was 80.8% (95% CI 77.9%-83.4%). The prevalence of patients by group was hypergonadotropic hypogonadal (42.4%, 38.9%-45.9%), normogonadotropic hypogonadal (38.5%; 35.1%-41.9%), hypergonadotropic eugonadal (8.3%; 6.6%-10.5%), and normogonadotropic eugonadal (10.8%; 8.8%-13.2%). Reduced testicular volume and lower estradiol levels were associated with an increased likelihood of hypogonadism. Paternal age was also an independent predictor, with higher age linked to an increased likelihood of hypogonadism. Hypogonadism was less likely in patients with germ cell maturation arrest and more likely in those with Sertoli cell-only. Patients with hypergonadotropic hypogonadism had lower SR success than normogonadotropic eugonadal counterparts (aRR 0.611; 95% CI 0.398-0.855). In the subset of hypogonadal men, hypergonadotropic patients had lower SR success than normogonadotropic participants (aRR 0.632; 0.469-0.811). Conclusion: The prevalence of biochemical hypogonadism among men with NOA is substantial. Hypogonadism is associated with testicular volume, estradiol levels, age, and histopathology patterns. This condition impacts SR success and emphasizes the need for improved care for men with NOA.

Systematic molecular analyses for 115 karyotypically normal men with isolated non-obstructive azoospermia.

STUDY QUESTION: Do copy-number variations (CNVs) in the azoospermia factor (AZF) regions and monogenic mutations play a major role in the development of isolated (non-syndromic) non-obstructive azoospermia (NOA) in Japanese men with a normal 46, XY karyotype? SUMMARY ANSWER: Deleterious CNVs in the AZF regions and damaging sequence variants in eight genes likely constitute at least 8% and approximately 8% of the genetic causes, respectively, while variants in other genes play only a minor role. WHAT IS KNOWN ALREADY: Sex chromosomal abnormalities, AZF-linked microdeletions, and monogenic mutations have been implicated in isolated NOA. More than 160 genes have been reported as causative/susceptibility/candidate genes for NOA. STUDY DESIGN, SIZE, DURATION: Systematic molecular analyses were conducted for 115 patients with isolated NOA and a normal 46, XY karyotype, who visited our hospital between 2017 and 2021. PARTICIPANTS/MATERIALS, SETTING, METHODS: We studied 115 unrelated Japanese patients. AZF-linked CNVs were examined using sequence-tagged PCR and multiplex ligation-dependent probe amplification, and nucleotide variants were screened using whole exome sequencing (WES). An optimized sequence kernel association test (SKAT-O), a gene-based association study using WES data, was performed to identify novel disease-associated genes in the genome. The results were compared to those of previous studies and our in-house control data. MAIN RESULTS AND THE ROLE OF CHANCE: Thirteen types of AZF-linked CNVs, including the hitherto unreported gr/gr triplication and partial AZFb deletion, were identified in 63 (54.8%) cases. When the gr/gr deletion, a common polymorphism in Japan, was excluded from data analyses, the total frequency of CNVs was 23/75 (30.7%). This frequency is higher than that of the reference data in Japan and China (11.1% and 14.7%, respectively). Known NOA-causative AZF-linked CNVs were found in nine (7.8%) cases. Rare damaging variants in known causative genes (DMRT1, PLK4, SYCP2, TEX11, and USP26) and hemizygous/multiple-heterozygous damaging variants in known spermatogenesis-associated genes (TAF7L, DNAH2, and DNAH17) were identified in nine cases (7.8% in total). Some patients carried rare damaging variants in multiple genes. SKAT-O detected no genes whose rare damaging variants were significantly accumulated in the patient group. LIMITATIONS, REASONS FOR CAUTION: The number of participants was relatively small, and the clinical information of each patient was fragmentary. Moreover, the pathogenicity of identified variants was assessed only by in silico analyses. WIDER IMPLICATIONS OF THE FINDINGS: This study showed that various AZF-linked CNVs are present in more than half of Japanese NOA patients. These results broadened the structural variations of AZF-linked CNVs, which should be considered for the molecular diagnosis of spermatogenic failure. Furthermore, the results of this study highlight the etiological heterogeneity and possible oligogenicity of isolated NOA. STUDY FUNDING/COMPETING INTEREST(S): This study was supported by Grants from the Japan Society for the Promotion of Science (21K19283 and 21H0246), the Japan Agency for Medical Research and Development (22ek0109464h0003), the National Center for Child Health and Development, the Canon Foundation, the Japan Endocrine Society, and the Takeda Science Foundation. The results of this study were based on samples and patient data obtained from the International Center for Reproductive Medicine, Dokkyo Medical University Saitama Medical Center, Koshigaya, Japan. The authors have no conflicts of interest to disclose. TRIAL REGISTRATION NUMBER: N/A.

Telomeric function and regulation during male meiosis in mice and humans.

BACKGROUND: Telomeres are unique structures situated at the ends of chromosomes. Preserving the structure and function of telomeres is essential for maintaining genomic stability and promoting genetic diversity during male meiosis in mammals. MATERIAL-METHODS: This review compiled recent literature on the function and regulation of telomeres during male meiosis in both mice and humans, and also highlighted the critical roles of telomeres in reproductive biology and medicine. RESULTS-DISCUSSION: Various structures, consisting of the LINC complex (SUN-KASH), SPDYA-CDK2, TTM trimer (TERB1-TERB2-MAJIN), and shelterin, are critical in controlling telomeric activities, such as nuclear envelope attachment and bouquet formation. Other than telomere-related proteins, cohesins and genes responsible for regulating telomere function are also highlighted, though the exact mechanism remains unclear. The gene-mutant mouse models with meiotic defects directly reveal the essential roles of telomeres in male meiosis. Recently reported mutant genes associated with telomere activity in clinical practice have also been illustrated in detail. CONCLUSIONS: Proper regulation of telomere activities is essential for male meiosis progression in mice and humans.

Technology for Biobanking of Epididymal Spermatozoa from Patient with Obstructive Azoospermia: Case Report about Baby Born after Conventional Freezing Only with a Nonpermeable Cryoprotectant 360 kDa Polyvinylpyrrolidone.

This publication reports, for the first time, the birth of a healthy child after intracytoplasmic sperm injection (ICSI) of motile spermatozoa after conventional ("slow") freezing of epididymal spermatozoa using 5% polyvinylpyrrolidone (PVP) of high molecular weight (360 kDa). Cryopreservation solution with 10% PVP was added to 30 µL of spermatozoa suspension in a 1:1 ratio, with a final PVP concentration of 5%. Then, polycarbonate capillaries for oocyte denudation with a diameter of 170 µm were filled with 60 µL of the resulting sperm suspension. After that, the capillaries were placed for 10 minutes at a height of 15 cm above liquid nitrogen and immersed into liquid nitrogen. To warm the spermatozoa, the capillaries were immersed in a water bath at a temperature of 40°C for 30 seconds. Oocyte fertilization was performed by ICSI. Zygotes were cultured in vitro for 5 days to the blastocyst stage. More than 100 spermatozoa were obtained after percutaneous epidydimal sperm aspiration, of which 80% were motile. After cryopreservation, storage for 3 months in liquid nitrogen, and thawing, 72% of the total sperm cells remained motile. Ten oocyte-cumulus complexes were found after follicle puncture, and eight metaphase II stage oocytes were fertilized using ICSI. After 18 hours, two pronuclei were found in seven (88%) of the oocytes. An analysis of the morphological characteristics of 5-day-old embryos showed that four (57%) of them reached the blastocyst stage. One embryo was transferred, and the remaining embryos were cryopreserved (vitrified). The onset of pregnancy was detected on the 14th day after embryo transfer, and one healthy girl (3300 g) was born at term.

Sox9 downregulation in non-obstructive azoospermia by UTF1 and mediator role of POU5F1.

BACKGROUND: Spermatogenesis is the process of producing mature sperm from Spermatogonial stem cells (SSCs) and this process requires a complex cooperation of different types of somatic and germ cells. Undifferentiated spermatogonia initiate the spermatogenesis and Sertoli cells as the only somatic cells inside of the seminiferous tubule play a key role in providing chemical and physical requirements for normal spermatogenesis, here, we investigated the dysfunction of these cells in non-obstructive azoospermia. MATERIAL AND METHOD: In this study, we analyzed the expression of sox9 and UTF1 in the non-obstructive human testis by immunohistochemistry. Moreover, we used the KEGG pathway and bioinformatics analysis to reveal the connection between our object genes and protein. RESULTS: The immunohistochemistry analysis of the non-obstructive human seminiferous tubule showed low expression of Sox9 and UTF1 that was detected out of the main location of the responsible cells for these expressions. Our bioinformatics analysis clearly and strongly indicated the relation between UTF1 in undifferentiated spermatogonia and Sox9 in Sertoli cells mediated by POU5F1. CONCLUSION: Generally, this study showed the negative effect of POU5F1 as a mediator between Sertoli cells as the somatic cells within seminiferous tubules and undifferentiated spermatogonia as the spermatogenesis initiator germ cells in non-obstructive conditions.

Micromapping testicular sperm extraction: A new technique for microscopic testicular sperm extraction in nonobstructive azoospermia.

Purpose: In microscopic testicular sperm extraction (mTESE) for nonobstructive azoospermia (NOA), sperm can be recovered relatively easily in some cases, and mTESE may be retrospectively considered excessive. However, mTESE is routinely performed in the majority of NOA patients because of the difficulty in predicting tissue status. A minimally invasive and comprehensive sperm retrieval method that allows on-the-spot tissue assessment is needed. We have developed and evaluated a novel sperm retrieval technique for NOA called micromapping testicular sperm extraction (MMTSE). Methods: MMTSE involves dividing the testis into four sections and making multiple small needle holes in the tunica albuginea to extract seminiferous tubules and retrieve sperm. The sperm-positive group by MMTSE (Group I) underwent additional tissue collection (ATC) via a small incision, whereas the sperm-negative group by MMTSE (Group 0) underwent mTESE. Results: In total, 40 NOA participants underwent MMTSE. Group I included 15 patients and Group 0 included 25 patients. In Group 1, sperm were recovered from all patients by ATC. In Group 0, sperm were recovered in 4 of 25 cases using mTESE. Conclusions: MMTSE shows promise as a simple method that comprehensively searches testicular tissue and retrieves sperm using an appropriate method while minimizing patient burden.