Sperm epigenetics and male infertility: unraveling the molecular puzzle.

BACKGROUND: The prevalence of infertility among couples is estimated to range from 8 to 12%. A paradigm shift has occurred in understanding of infertility, challenging the notion that it predominantly affects women. It is now acknowledged that a significant proportion, if not the majority, of infertility cases can be attributed to male-related factors. Various elements contribute to male reproductive impairments, including aberrant sperm production caused by pituitary malfunction, testicular malignancies, aplastic germ cells, varicocele, and environmental factors. MAIN BODY: The epigenetic profile of mammalian sperm is distinctive and specialized. Various epigenetic factors regulate genes across different levels in sperm, thereby affecting its function. Changes in sperm epigenetics, potentially influenced by factors such as environmental exposures, could contribute to the development of male infertility. CONCLUSION: In conclusion, this review investigates the latest studies pertaining to the mechanisms of epigenetic changes that occur in sperm cells and their association with male reproductive issues.

Tetrabromobisphenol A reduces male rats reproductive organ coefficients and disrupting sexual hormone by causing oxidative stress.

Tetrabromobisphenol A (TBBPA) has become a topic of public attention due to its pervasive detection in the environment and organisms in recent decades. However, limited information is available regarding the toxicity of TBBPA on reproductive ability of male mammals. Herein, the reproductive toxicity of TBBPA was investigated in male rats to fill the knowledge gap. In this study, male rats were exposed to TBBPA (0, 10, 100, and 1000 mg/kg) for 6 weeks. Subsequently, body and organ indexes, histopathological evaluation of testis and epididymis, ultrastructural observation of sperm, testosterone and progesterone levels, and oxidative stress indicators were conducted to reveal corresponding mechanisms. Results obtained showed that compare to the control group, the body weight, testes weight, epididymis weight, seminal vesicle and coagulation glands weight of rats in the 1000 mg/kg group lost 8.30%, 16.84%, 20.16%, 19.72% and 26.42%, respectively. Intriguingly, exposure to TBBPA (10, 100, 100 mg/kg) resulted in substantial pathological damage in testis, epididymis and sperm. TBBPA exposure also increased malondialdehyde (MDA) and hydrogen peroxide (H2O2) contents, as well as superoxide dismutase (T-SOD) and catalase (CAT) activities in testicular tissue. What's more, the testosterone and progesterone levels in male rat serum were significantly decreased after exposure to TBBPA for 6 weeks. Meanwhile, results of molecular docking showed that TBBPA has a strong affinity with estrogen receptors (ERs). These findings demonstrated that TBBPA exposure negatively impacts the reproductive ability of male rats, thus providing new insights for risk assessment for reproductive health under TBBPA exposure.

Effects of quercetin on gentamicin-induced experimental testicular injury in rats.

The purpose of this study was to investigate the effects of gentamicin (GEN) on the testis and whether quercetin (QUE) has any protective effect. Twenty-four adult male Sprague-Dawley rats were divided into equal four groups: control (0.9% saline solution), GEN (80 mg∕kg GEN), QUE (50 mg∕kg QUE) and GEN+QUE (80 mg∕kg GEN + 50 mg∕kg QUE). Histopathological (HP) evaluation of testis was performed, epididymal sperm parameters were analyzed and oxidative status was evaluated. The use of QUE improved the HP findings, such as decrease in the germinal epithelial thickness in the testicular tissue of the GEN group, decrease in the Johnsen's tubular biopsy score (JTBS), increase in the rate of immature cell shedding tubules, and the apoptotic index (AI). In the GEN group, sperm count, and abnormal morphology increased compared to the control group; the viability and motility decreased according to the sperm analysis results. In the GEN+QUE group, QUE was found to improve sperm viability and morphology. In the GEN group, tissue malondialdehyde (MDA) levels increased while superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) levels decreased. Compared with the GEN+QUE group, it was found that the tissue MDA level decreased, while the levels of SOD, CAT and GPx increased. The results demonstrate that GEN impairs testicular structure and function, and QUE treatment can prevent this adverse effect.

Evaluation of human bone marrow mesenchymal stem cells in the treatment of non obstructive azoospermia.

BACKGROUND: Non-obstructive azoospermia (NOA) represents an infertility problem that is usually difficult to treat. Such patients usually have testicular biopsy of germ cell aplasia or spermatogenic arrest. In recent decades, mesenchymal stem cells (MSCs) had been studied thoroughly and proved safe and effective regarding their capability for trans-differentiation into different cell types. The aim of this study was to evaluate the effect of MSCs local intratesticular injection in induction of spermatogenesis. PATIENTS AND METHOD: The current study included 87 infertile non-obstructive azoospermic patients. Clinical assessment and repeated semen analysis with centrifugation were done to confirm azoospermia. Karyotyping and AZF study were done. Some of the patients had previous testicular biopsy proving a lack of sperm in the testes. Single intratesticular injection of purified MSCs suspension was done. RESULTS: 20.7% of patients showed sperm in their semen after variable period of time. Hormonal profile among treated patients showed significant improvement regardless success of treatment. Also most of the treated patients appreciated the improvement of their sexual function and libido. CONCLUSIONS: Bone marrow derived MSCs could be a new hope and therapeutic modality for treatment of refractory cases of NOA.

Identification of two hidden clinical subgroups among men with idiopathic cryptozoospermia.

STUDY QUESTION: Are there subgroups among patients with cryptozoospermia pointing to distinct etiologies? SUMMARY ANSWER: We reveal two distinct subgroups of cryptozoospermic (Crypto) patients based on testicular tissue composition, testicular volume, and FSH levels. WHAT IS KNOWN ALREADY: Cryptozoospermic patients present with a sperm concentration below 0.1 million/ml. While the etiology of the severely impaired spermatogenesis remains largely unknown, alterations of the spermatogonial compartment have been reported including a reduction of the reserve stem cells in these patients. STUDY DESIGN, SIZE, DURATION: To assess whether there are distinct subgroups among cryptozoospermic patients, we applied the statistical method of cluster analysis. For this, we retrospectively selected 132 cryptozoospermic patients from a clinical database who underwent a testicular biopsy in the frame of fertility treatment at a university hospital. As controls (Control), we selected 160 patients with obstructive azoospermia and full spermatogenesis. All 292 patients underwent routine evaluation for endocrine, semen, and histological parameters (i.e. the percentage of tubules with elongated spermatids). Moreover, outcome of medically assisted reproduction (MAR) was assessed for cryptozoospermic (n = 73) and Control patients (n = 87), respectively. For in-depth immunohistochemical and histomorphometrical analyses, representative tissue samples from cryptozoospermic (n = 27) and Control patients (n = 12) were selected based on cluster analysis results and histological parameters. PARTICIPANTS/MATERIALS, SETTING, METHODS: This study included two parts: firstly using clinical parameters of the entire cohort of 292 patients, we performed principal component analysis (PCA) followed by hierarchical clustering on principal components (i.e. considering hormonal values, ejaculate parameters, and histological information). Secondly, for histological analyses seminiferous tubules were categorized according to the most advanced germ cell type present in sections stained with Periodic acid Schif. On the selected cohort of 39 patients (12 Control, 27 cryptozoospermic), we performed immunohistochemistry for spermatogonial markers melanoma-associated antigen 4 (MAGEA4) and piwi like RNA-mediated gene silencing 4 (PIWIL4) followed by quantitative analyses. Moreover, the morphologically defined Adark spermatogonia, which are considered to be the reserve stem cells, were quantified. MAIN RESULTS AND THE ROLE OF CHANCE: The PCA and hierarchical clustering revealed three different clusters, one of them containing all Control samples. The main factors driving the sorting of patients to the clusters were the percentage of tubules with elongated spermatids (Cluster 1, all Control patients and two cryptozoospermic patients), the percentage of tubules with spermatocytes (Cluster 2, cryptozoospermic patients), and tubules showing a Sertoli cells only phenotype (Cluster 3, cryptozoospermic patients). Importantly, the percentage of tubules containing elongated spermatids was comparable between Clusters 2 and 3. Additional differences were higher FSH levels (P < 0.001) and lower testicular volumes (P < 0.001) in Cluster 3 compared to Cluster 2. In the spermatogonial compartment of both cryptozoospermic Clusters, we found lower numbers of MAGEA4+ and Adark spermatogonia but higher proportions of PIWIL4+ spermatogonia, which were significantly correlated with a lower percentage of tubules containing elongated spermatids. In line with this common alteration, the outcome of MAR was comparable between Controls as well as both cryptozoospermic Clusters. LIMITATIONS, REASONS FOR CAUTION: While we have uncovered the existence of subgroups within the cohort of cryptozoospermic patients, comprehensive genetic analyses remain to be performed to unravel potentially distinct etiologies. WIDER IMPLICATIONS OF THE FINDINGS: The novel insight that cryptozoospermic patients can be divided into two subgroups will facilitate the strategic search for underlying genetic etiologies. Moreover, the shared alterations of the spermatogonial stem cell compartment between the two cryptozoospermic subgroups could represent a general response mechanism to the reduced output of sperm, which may be associated with a progressive phenotype. This study therefore offers novel approaches towards the understanding of the etiology underlying the reduced sperm formation in cryptozoospermic patients. STUDY FUNDING/COMPETING INTEREST(S): German research foundation CRU 326 (grants to: SDP, NN). Moreover, we thank the Faculty of Medicine of the University of Münster for the financial support of Lena Charlotte Schülke through the MedK-program. We acknowledge support from the Open Access Publication Fund of the University of Münster. The authors have no potential conflicts of interest. TRIAL REGISTRATION NUMBER: N/A.

Semen parameters in testicular tumor patients before orchiectomy: What is the impact of testicular tumor stage and histology?

PURPOSE: To evaluate the influence of testicular cancer histology and stage on sperm parameters in cryopreserved samples collected prior to orchiectomy. MATERIALS AND METHODS: We conducted a retrospective analysis of tumor histology, stage and sperm parameters of patients who underwent pre-orchiectomy sperm cryopreservation for testicular cancer between March 2010 and March 2023. The World Health Organization (WHO) 2010 sperm reference values were used to identify patients with subnormal semen parameters and to further categorize patients by sperm alteration. Localized disease was classified as Stage I, while metastatic disease encompassed Stages II and III. Continuous variables were compared using t-test or Mann Whitney U test, and categorical variables using Chi-square and Fishers exact test. RESULTS: A total of 64 patients was identified, 48 (75%) classified as stage I and 16 (25%) classified as stage II/III. No difference was found in semen parameters between patients with seminoma and patients with non-seminoma germ cell tumor (NSGCT). Patients with stage II/III disease had significantly lower percentages of progressive motility (36% vs 53%, p=0.021) and total motility (60% vs 69%, p=0.015) than stage I patients. When categorizing by sperm alterations according to WHO 2010 reference values, patients with stage II/III disease had significantly higher proportions of asthenozoospermia (38% vs 15%, p=0.048) and teratozoospermia (63% vs 31%, p=0.027) than stage I patients. Elevated tumor markers were not associated with sperm abnormalities. CONCLUSIONS: Patients with metastatic testicular cancer present with worse sperm quality than patients with localized disease. Sperm cryopreservation should be offered to all patients with testicular cancer, and especially emphasized in patients with metastatic disease.

Case report: remedial microdissection testicular sperm extraction after onco-microdissection testicular sperm extraction failure.

BACKGROUND: Testicular cancer (TC) mostly occurs in men aged 14 to 44. Studies have shown that TC seriously damages male fertility, and 6% to 24% of patients with TC were even found to suffer from azoospermia when they are diagnosed. At present, some studies have pointed out that onco-microdissection testicular sperm extraction (mTESE) can extract sperm from tumor testicles. However, there are almost no reports on remedial measures after onco-mTESE failure. Given the valuable opportunity for fertility preservation in patients with TC and azoospermia, it is necessary to provide effective remedial methods for patients with failed onco-mTESE. METHODS: Two young men, who were diagnosed with TC and also found to have azoospermia, tried onco-mTESE while undergoing radical orchiectomy for fertility preservation. However, sperm extraction failed in both patients. Subsequently, the isolated testicular tissue of the patient in case 1 suffered from TC again, and the patient in case 2 was scheduled to receive multiple cycles of gonadotoxic chemotherapy. Because both had a plan to have a birth in the future, we performed remedial mTESE. RESULTS: Sperm was successfully extracted from both patients. The patient recovered well, without complications. The patient couple in case 1 underwent 1 intracytoplasmic sperm injection (ICSI) cycle but did not achieve clinical pregnancy. CONCLUSIONS: There is still an opportunity to extract sperm successfully using onco-mTESE, despite the difficulty of fertility preservation in TC patients with azoospermia. If sperm extraction from the tumor testis fails, implementing remedial mTESE as early as possible would likely preserve the last chance of fertility for these patients.

Timing of Testicular Biopsy in Relation to Oocyte Retrieval and the Outcomes of Intracytoplasmic Sperm Injection.

PURPOSE: We evaluate microscopic (micro) testicular sperm extraction (TESE) timing relative to oocyte retrieval on intracytoplasmic sperm injection outcome. MATERIALS AND METHODS: Couples with nonobstructive azoospermia who underwent intracytoplasmic sperm injection with freshly retrieved spermatozoa were analyzed based on whether micro-TESE was performed at least 1 day prior to oocyte retrieval (TESE-day-before group) or on the day of oocyte retrieval (TESE-day-of group). Embryology and clinical outcomes were compared. RESULTS: The percentage of patients who underwent a successful testicular sperm retrieval was significantly lower in the TESE-day-before cohort (62%) than in the TESE-day-of cohort (69%; odds ratio [OR] 1.4, 95% CI [1.1, 1.7], P < .001). The fertilization rate was also found to be significantly lower in the TESE-day-before group (45%) than in the TESE-day-of group (53%; OR 1.4, 95% CI [1.2, 1.7], P = .01). Although the association between the cleavage rate and TESE timing was not statistically significant, the implantation rate was found to be significantly higher in the day-before cohort (28%) than in the day-of cohort (22%; OR 0.7, 95% CI [0.6, 0.9], P = .01). Nevertheless, it was found that the clinical pregnancy and delivery rates were not statistically significantly associated with the TESE timing. CONCLUSIONS: Although sperm retrieval and fertilization rates were lower in the TESE-day-before cohort, the 2 cohorts showed comparable embryologic and clinical outcomes. Micro-TESE can be performed before oocyte harvesting to provide physicians ample time to decide between cancelling oocyte retrieval or retrieving oocytes for cryopreservation.

Genetic profiling of azoospermic men to identify the etiology and predict reproductive potential.

PURPOSE: To identify germline mutations related to azoospermia etiology and reproductive potential of surgically retrieved spermatozoa, and to investigate the feasibility of predicting seminiferous tubule function of nonobstructive azoospermic men by transcriptomic profiling of ejaculates. MATERIALS AND METHODS: Sperm specimens were obtained from 30 men (38.4 ± 6 years) undergoing epididymal sperm aspiration for obstructive azoospermia (OA, n = 19) acquired by vasectomy, or testicular biopsy for nonobstructive azoospermia (NOA, n = 11). To evaluate for a correlation with azoospermia etiology, DNAseq was performed on surgically retrieved spermatozoa, and cell-free RNAseq on seminal fluid (n = 23) was performed to predict spermatogenesis in the seminiferous tubule. RESULTS: Overall, surgically retrieved sperm aneuploidy rates were 1.7% and 1.8% among OA and NOA cohorts, respectively. OA men carried housekeeping-related gene mutations, while NOA men displayed mutations on genes involved in crucial spermiogenic functions (AP1S2, AP1G2, APOE). We categorized couples within each cohort according to ICSI clinical outcomes to investigate genetic causes that may affect reproductive potential. All OA-fertile men (n = 9) carried mutations in ZNF749 (sperm production), whereas OA-infertile men (n = 10) harbored mutations in PRB1, which is essential for DNA replication. NOA-fertile men (n = 8) carried mutations in MPIG6B (stem cell lineage differentiation), whereas NOA-infertile individuals (n = 3) harbored mutations in genes involved in spermato/spermio-genesis (ADAM29, SPATA31E1, MAK, POLG, IFT43, ATG9B) and early embryonic development (MBD5, CCAR1, PMEPA1, POLK, REC8, REPIN1, MAPRE3, ARL4C). Transcriptomic assessment of cell-free RNAs in seminal fluid from NOA men allowed the prediction of residual spermatogenic foci. CONCLUSIONS: Sperm genome profiling provides invaluable information on azoospermia etiology and identifies gene-related mechanistic links to reproductive performance. Moreover, RNAseq assessment of seminal fluid from NOA men can help predict sperm retrieval during testicular biopsies.

Predictors of successful salvage microdissection testicular sperm extraction (mTESE) after failed initial TESE in patients with non-obstructive azoospermia: A systematic review and meta-analysis.

BACKGROUND: There has been no systematic review and meta-analysis to analyze and summarize the predictive factors of successful sperm extraction in salvage microdissection testicular sperm extraction. OBJECTIVES: We aimed to investigate the factors predicting the result of salvage microdissection testicular sperm extraction in patients with non-obstructive azoospermia who failed the initial microdissection testicular sperm extraction or conventional testicular sperm extraction. MATERIALS AND METHODS: We conducted a systematic literature search in PubMed, Web of Science, EMBASE, and the Cochrane Library for literature that described the characteristics of patients with non-obstructive azoospermia who underwent salvage microdissection testicular sperm extraction after failing the initial microdissection testicular sperm extraction or conventional testicular sperm extraction published prior to June 2022. RESULTS: This meta-analysis included four retrospective studies with 332 patients with non-obstructive azoospermia who underwent a failed initial microdissection testicular sperm extraction and three retrospective studies with 177 non-obstructive azoospermia patients who underwent a failed conventional testicular sperm extraction. The results were as follows: among non-obstructive azoospermia patients whose first surgery was microdissection testicular sperm extraction, younger patients (standard mean difference: -0.28, 95% confidence interval [CI]: -0.55 to -0.01) and those with smaller bilateral testicular volume (standard mean difference: -0.55, 95% CI: -0.95 to -0.15), lower levels of follicle-stimulating hormone (standard mean difference: -0.86, 95% CI: -1.18 to -0.54) and luteinizing hormone (standard mean difference: -0.68, 95% CI: -1.16 to -0.19), and whose testicular histological type was hypospermatogenesis (odds ratio: 3.52, 95% CI: 1.30-9.53) were more likely to retrieve spermatozoa successfully, while patients with Sertoli-cell-only syndrome (odds ratio: 0.41, 95% CI: 0.24-0.73) were more likely to fail again in salvage microdissection testicular sperm extraction. Additionally, in patients who underwent salvage microdissection testicular sperm extraction after a failed initial conventional testicular sperm extraction, those with testicular histological type of hypospermatogenesis (odds ratio: 30.35, 95% CI: 8.27-111.34) were more likely to be successful, while those with maturation arrest (odds ratio: 0.39, 95% CI: 0.18-0.83) rarely benefited. CONCLUSION: We found that age, testicular volume, follicle-stimulating hormone, luteinizing hormone, hypospermatogenesis, Sertoli-cell-only syndrome, and maturation arrest were valuable predictors of salvage microdissection testicular sperm extraction, which will assist andrologists in clinical decision-making and minimize unnecessary injury to patients.

Improved phenotypic classification of male infertility to promote discovery of genetic causes.

An increasing number of genes are being described in the context of non-syndromic male infertility. Linking the underlying genetic causes of non-syndromic male infertility with clinical data from patients is important to establish new genotype-phenotype correlations. This process can be facilitated by using universal nomenclature, but no standardized vocabulary is available in the field of non-syndromic male infertility. The International Male Infertility Genomics Consortium aimed at filling this gap, providing a standardized vocabulary containing nomenclature based on the Human Phenotype Ontology (HPO). The "HPO tree" was substantially revised compared with the previous version and is based on the clinical work-up of infertile men, including physical examination and hormonal assessment. Some causes of male infertility can already be suspected based on the patient's clinical history, whereas in other instances, a testicular biopsy is needed for diagnosis. We assembled 49 HPO terms that are linked in a logical hierarchy and showed examples of morphological features of spermatozoa and testicular histology of infertile men with identified genetic diagnoses to describe the phenotypes. This work will help to record patients' phenotypes systematically and facilitate communication between geneticists and andrologists. Collaboration across institutions will improve the identification of patients with the same phenotypes, which will promote the discovery of novel genetic causes for non-syndromic male infertility.

A novel NPHP4 homozygous missense variant identified in infertile brothers with multiple morphological abnormalities of the sperm flagella.

PURPOSE: Asthenozoospermia is an important cause of male infertility, and the most serious type is characterized by multiple morphological abnormalities of the sperm flagella (MMAF). However, the precise etiology of MMAF remains unknown. In the current study, we recruited a consanguineous Pakistani family with two infertile brothers suffering from primary infertility due to MMAF without obvious signs of PCD. METHODS: We performed whole-exome sequencing on DNAs of the patients, their parents, and a fertile brother and identified the homozygous missense variant (c.1490C > G (p.P497R) in NPHP4 as the candidate mutation for male infertility in this family. RESULTS: Sanger sequencing confirmed that this mutation recessively co-segregated with the MMAF in this family. In silico analysis revealed that the mutation site is conserved across different species, and the identified mutation also causes abnormalities in the structure and hydrophobic interactions of the NPHP4 protein. Different bioinformatics tools predict that NPHP4p.P497R mutation is pathogenic. Furthermore, Papanicolaou staining and scanning electron microscopy of sperm revealed that affected individuals displayed typical MMAF phenotype with a high percentage of coiled, bent, short, absent, and/or irregular flagella. Transmission electron microscopy images of the patient's spermatozoa revealed significant anomalies in the sperm flagella with the absence of a central pair of microtubules (9 + 0) in every section scored. CONCLUSIONS: Taken together, these results show that the homozygous missense mutation in NPHP4 is associated with MMAF.

Microdissection testicular sperm extraction outcomes in azoospermic patients with bilateral orchidopexy.

BACKGROUND: Cryptorchidism is considered to be one of the most common causes of non-obstructive azoospermia. There are several surgical techniques to retrieve sperm in these patients. Microdissection testicular sperm extraction (m-TESE) is a recent sperm retrieval technique which is considered to be a safe, non-blind, and feasible method. OBJECTIVES: This study aimed to investigate sperm retrieval rate (SRR) by the mTESE method in patients who have undergone orchidopexy due to bilateral cryptorchidism. MATERIALS AND METHODS: In this retrospective study, 56 ex-cryptorchid patients, who underwent mTESE due to post orchidopexy azoospermia, were included. Patients with hypogonadotropic hypogonadism, Klinefelter syndrome, azoospermia factors (AZF) microdeletion, or chromosomal translocation were excluded from the study. Data were obtained from medical files. RESULTS: SRR in this study was 46%. Patients were divided into two groups of negative (n = 30) and positive (n = 26) based on the sperm extraction outcomes. There was no statistically significant difference between two groups regarding the mean age at mTESE, mean age at orchidopexy, testicular size, and serum testosterone concentration. However, testicular location, histological patterns, FSH, and LH level showed to have statistically significant relation with sperm retrieval results. But, according to our logistic regression, none of the included variable in the model including FSH, LH, histopathology, and testis location have a significant effect on the presence of the sperm. DISCUSSION: In the present study, SRR was significantly higher in patients with scrotal testis and low level of FSH and LH. CONCLUSIONS: Performing mTESE could be recommended in ex-cryptorchid patients with post orchidopexy NOA. Preoperative testicular biopsy seems to be unnecessary while clinical criteria can perfectly define NOA.

Effects of 4G mobile phone radiation exposure on reproductive, hepatic, renal, and hematological parameters of male Wistar rat.

BACKGROUND AND OBJECTIVE: Mobile phones have become a vital part of human life. Due to drastic increase in the number of mobile phone subscribers, exposure to radiofrequency radiation (RFR) emitted from these phones has increased dramatically. Hence, the effect of RFR on humans is an area of concern. This study was performed to determine the impact of 4G mobile phone radiation on the male reproductive system, liver, kidney, and hematological parameters. METHODS: Seventy-day-old Wistar rats were exposed to 4G radiation (2350 MHz for 2 h/day for 56 days). Sperm parameters such as sperm count, viability, sperm head morphology, mitochondrial activity, total antioxidant activity, and lipid peroxidation of sperm were evaluated. Histopathology of the testis, prostate, epididymis, seminal vesicle, liver, and kidney was carried out. Complete blood count, liver and kidney function tests, and testosterone hormone analysis were done. RESULTS: At the end of the experiment, results showed a significant (p < 0.05) decrease in sperm viability with alterations in the histology of the liver, kidney, testis, and other reproductive organs in the exposed group of rats. A reduced level of testosterone, total antioxidant capacity, and decreased sperm mitochondrial function were also observed in the exposed rats. Moreover, the exposed rats showed an increase in sperm lipid peroxidation and sperm abnormality. Hematological parameters like hemoglobin, red blood cells (RBC), and packed cell volume (PCV) showed a significant (p < 0.05) increase in the exposed rats. CONCLUSION: The results indicate that chronic exposure to 4G radiation may affect the male reproductive system, hematological system, liver, and kidney of rats.

Results from the first autologous grafting of adult human testis tissue: a case report.

Fertility restoration using autologous testicular tissue transplantation is relevant for infertile men surviving from childhood cancer and, possibly, in men with absent or incomplete spermatogenesis resulting in the lack of spermatozoa in the ejaculate (non-obstructive azoospermia, NOA). Currently, testicular tissue from pre-pubertal boys extracted before treatment with gonadotoxic cancer therapy can be cryopreserved with good survival of spermatogonial stem cells. However, strategies for fertility restoration, after successful cancer treatment, are still experimental and no clinical methods have yet been developed. Similarly, no clinically available treatments can help men with NOA to become biological fathers after failed attempts of testicular surgical sperm retrieval. We present a case of a 31-year-old man with NOA who had three pieces of testis tissue (each ∼2 × 4 × 2 mm3) extracted and cryopreserved in relation to performing microdissection testicular sperm extraction (mTESE). Approximately 2 years after mTESE, the thawed tissue pieces were engrafted in surgically created pockets bilaterally under the scrotal skin. Follow-up was performed after 2, 4, and 6 months with assessment of reproductive hormones and ultrasound of the scrotum. After 6 months, all engrafted tissue was extracted and microscopically analyzed for the presence of spermatozoa. Furthermore, parts of the extracted tissue were analyzed histologically and by immunohistochemical analysis. Active blood flow in the engrafted tissue was demonstrated by doppler ultrasound after 6 months. No spermatozoa were found in the extracted tissue. Histological and immunohistochemical analysis demonstrated graft survival with intact clear tubules and normal cell organization. Sertoli cells and spermatocytes with normal morphology were located near the basement membrane. MAGE-A and VASA positive spermatogonia/spermatocytes were detected together with SOX9 positive Sertoli cells. Spermatocytes and/or Sertoli cells positive for γH2AX was also detected. In summary, following autologous grafting of frozen-thawed testis tissue under the scrotal skin in a man with NOA, we demonstrated graft survival after 6 months. No mature spermatozoa were detected; however, this is likely due to the pre-existing spermatogenic failure.

Impact of testicular cancer on sperm small non-coding RNA signature: a pilot study.

Testicular germ cell tumours (TGCTs) are the most common tumours in young adults of European ancestry. The high heritability and the constantly increased incidence, which has doubled over the last 20 years, strongly suggest that both genetic and environmental factors are likely to shape the TGCT susceptibility. While genome-wide association studies have identified loci associated with TGCT susceptibility, the role played by environmental molecular vectors in TGCT susceptibility remains unclear. Evidence shows that sperm non-coding RNAs provide a good vision of the environmental stresses experienced by men. Here, to determine whether TGCT impacts the abundance of specific non-coding RNAs in sperm, small RNA deep sequencing analysis of sperm of 25 men aged between 19 and 42 years, diagnosed with (n = 16) or without (n = 9) TGCT was performed. The primary analysis showed no statistical significance in the sncRNA population between the TGCT and non-TGCT groups. However, when sperm physiological parameters were considered to look for differentially expressed sncRNA, we evidenced 11 differentially expressed sncRNA between patients and control which allow a clear discrimination between control and TGCT samples after Hierarchical Clustering analysis. Together, these findings indicate that sperm small non-coding RNAs abundance may have the potential for diagnosing men with TGCT. However, specific care should be taken regarding sperm physiological parameters of the TGCT patients. Hence, larger studies are needed to confirm our findings and to determine whether such a signature associates with the risks to develop TGCT.

The Relationship of Testicular Sonoelastography with Gonadotropin Hormone Levels and Sperm Parameters.

Background: Elastography is a non-invasive medical imaging technique that helps determine the stiffness of organs and other structures in our body. In this study, we investigated the effectiveness of elastography in the diagnosis of infertility. Aim: In this study, we aimed to examine the relationship between testicular elastography and hormonal parameters and sperm parameters. Patients and Methods: The study included 136 patients, 272 testicles were examined, and the mean age of the study participants was 30.1 years. Testicular tissue stiffness was measured by scrotal ultrasonographic shear wave elastography. Gonadotropin and testosterone hormones were measured from blood samples. Spermiogram parameters were studied manually. Results: The control group included 66 patients, and the varicocele group consisted of 70 patients. Testicular stiffness degrees of the control group were measured as 4.29 kPa for the right testis and 4.23 kPa for the left testis. The varicocele group was divided into grades 1, 2, and 3 according to physical examination. In group 1 (grade 1), the right testis was 4.07 ± 1.24 kPa and the left testis was 3.77 ± 0.98 kPa. In group 2 (grade 2), the right testis was 4.31 ± 1.40 kPa and the left testis was 3.98 ± 0.93 kPa. In group 3 (grade 3), the right testis was 4.73 ± 1.50 kPa and the left testis was 3.99 ± 1.68 kPa. Hormone and sperm parameters were not statistically significant when comparing the control and varicocele groups. There was no statistical significance between the testicular tissue stiffness degrees of the control and varicocele groups. Hormone and spermiogram findings were also similar in groups. Conclusion: It is known that varicocele leads to histological tissue changes in the testes. These changes result in tissue softness and loss while affecting sperm parameters and testosterone levels in a negative way. Before varicocele surgery, there is a need for new imaging methods with more sensitivity that can detect tissue changes in the testes.

Sperm protein antigen 17 and Sperm flagellar 1 cancer testis antigens are expressed in a rare case of ciliated foregut cyst of the common hepatic duct.

INTRODUCTION: Ciliated foregut cysts (CFCs) are frequently described in liver, pancreas and gallbladder and generally considered benign although one case of squamous cell metaplasia and five cases of squamous cell carcinoma arising from a ciliated hepatic foregut cyst have been reported. Here we explore two cancer-testis antigens (CTAs), Sperm protein antigen 17 (SPA17) and Sperm flagellar 1 (SPEF1) expression in a rare case of CFC of the common hepatic duct MATERIALS AND METHODS: 3 µm-thick CFC sections were immunohistochemically treated with antibodies raised against human SPA17 or SPEF1. In silico Protein-Protein Interaction (PPI) network and differential protein expression were also investigated RESULTS: Immunohistochemistry revealed SPA17 and SPEF1 in the cytoplasm of ciliated epithelium. SPA17, but not SPEF1, was also detected in cilia. The PPI networks demonstrated that other CTAs are significantly predicted functional partners with SPA17 and SPEF1. The differential protein expression demonstrated that SPA17 was higher in breast cancer, cholangiocarcinoma, liver hepatocellular carcinoma, uterine corpus endometrial carcinoma, gastric adenocarcinoma, cervical squamous cell carcinoma, bladder urothelial carcinoma. SPEF1 expression was higher in breast cancer, cholangiocarcinoma, uterine corpus endometrial carcinoma and kidney renal papillary cell carcinoma CONCLUSIONS: Our study suggests that further characterization of SPA17 and SPEF1 in patients with CFCs might provide significant insights to understand the mechanisms underlying their potential to malignant transformation.

Touch print smear of testicular tissue with thionine stain for intraoperative diagnosis in azoospermic men.

PURPOSE: To compare the diagnostic value of testicular tissue touch print smear (TPS) conducted on azoospermic patients with results obtained from histopathology and in vitro fertility (IVF) lab findings. METHODS: Microdissection testicular sperm extraction was performed on a group of 148 azoospermic patients and testicular samples obtained intraoperatively. Using TPS, the samples were smeared onto a sterile slide, followed with staining using thionine. The testis tissue bulk samples were also transferred to the IVF lab, and determinations of sperm presence or absence obtained from IVF lab tests were compared with the TPS sample results. Needle testis biopsy was separately performed on a group of 360 azoospermic patients, and results of pathohistology review on the biopsies were further compared with determinations of spermatogenesis stage obtained from TPS for those patients. RESULTS: When compared with IVF lab results, TPS was found to have 100% (126/126) positive predictive value and 95.5% (25/26) negative predictive value for predicting sperm presence or absence, respectively. Furthermore, TPS was further found to have a 93.6% correlation (337 of 360 biopsies) with results of histological diagnoses performed by needle biopsy. Results from histology and TPS for the detection of sperm presence were concordant in 96.1% (346/360) of biopsies. Diagnosis of SCO by TPS shows the highest correlation with histopathology (98.6%), followed by complete spermatogenesis (97.5%), early maturation arrest (78.9%), and late maturation arrest (27.3%). CONCLUSIONS: The results support the continued use of TPS in testicular tissue analysis for more rapid assessment of spermatogenesis and for detection of spermatozoa in azoospermic subjects.