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Purpose: To explore the application possibility of macrocephalic sperm from a patient with 100% macrocephalic sperm and AURKC gene variations., Methods: We diagnosed a case of macrozoospermia with 100% macrocephalic sperm and 39.5% multi-tailed spermatozoa by morphological analysis. Whole-exome sequencing (WES) was used for the patient and his wife. Sanger sequencing technique was used to verify the AURKC mutations in the patient's parents and his offspring. Sperm's ploidy was tested by flow cytometry. The couple asked for intra-couple ART therapy., Results: The patient presented novel compound heterozygous AURKC mutations (c.434C > T, c.497A > T) by WES. Sanger sequencing validation showed that variant of c.434C > T was observed in his father and c.497A > T was observed in his mother. Flow cytometry revealed that there existed a certain proportion of haploid sperm. Macrocephalic spermatozoa whose heads were smaller than the diameter of injection needle were selected for microinjection. A singleton pregnancy was achieved after embryo transfer. Prenatal diagnosis revealed that the fetus had normal chromosomal karyotype. Sanger sequencing technique showed that the fetus carried a c.434C > T mutation in one AURKC allele. A 3730 g healthy male fetus was delivered at term., Conclusion: Our study reported a successful live birth from a patient with definite AURKC gene variants and may provide insights for such patients to choose donor sperm or their own sperm., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Background: The complete absence of the azoospermia factor A (AZFa) region typically results in nonobstructive azoospermia. Partial deletions of the AZFa region are particularly noteworthy due to the limited and enigmatic reports of partial deletions in the AZFa region. Here, we present a rare case report of partial deletion of sY84 in the AZFa region but exhibiting normal sperm quality. The aim of this case report is to gain a deeper insight into the impact of AZFa region deletion on male fertility and to guide future clinical decisions and treatment strategies., Methods and Results: A 25-year-old man presented to the hospital with his 25-year-old wife due to recurrent spontaneous abortions. Routine semen analysis, sperm morphology analysis, acrosomal enzyme analysis, sperm DNA fragmentation indexed, and peripheral blood karyotype analysis revealed no abnormalities. Y chromosome microdeletion was detected by real-time fluorescence polymerase chain reaction, which showed that sY84 could not be amplified and sY86 was amplified nonspecifically. The man was diagnosed with partial deletions in the AZFa region. The wife underwent in vitro fertilization treatment for tubal infertility and recurrent spontaneous abortions. The couple successfully delivered a healthy daughter weighing 2.7 kg at 39 weeks of gestation, following 2 assisted reproductive pregnancies., Conclusion: Our findings contribute to expanding our knowledge of the AZFa region. A sY84 deficiency in the AZFa region may not lead to spermatogenesis failure and may potentially be one of the factors causing recurrent miscarriages, which needs to be confirmed by further data., Competing Interests: The authors have no conflicts of interest to disclose., (Copyright © 2024 the Author(s). Published by Wolters Kluwer Health, Inc.)

The establishment of left-right asymmetry is a fundamental process in animal development. Interference with this process leads to a range of disorders collectively known as laterality defects, which manifest as abnormal arrangements of visceral organs. Among patients with laterality defects, congenital heart diseases (CHD) are prevalent. Through multiple model organisms, extant research has established that myosin-Id (MYO1D) deficiency causes laterality defects. This study investigated over a hundred cases and identified a novel biallelic variant of MYO1D (NM_015194: c.1531G>A; p.D511N) in a consanguineous family with complex CHD and laterality defects. Further examination of the proband revealed asthenoteratozoospermia and shortened sperm. Afterward, the effects of the D511N variant and another known MYO1D variant (NM_015194: c.2293C>T; p.P765S) were assessed. The assessment showed that both enhance the interaction with β-actin and SPAG6. Overall, this study revealed the genetic heterogeneity of this rare disease and found that MYO1D variants are correlated with laterality defects and CHD in humans. Furthermore, this research established a connection between sperm defects and MYO1D variants. It offers guidance for exploring infertility and reproductive health concerns. The findings provide a critical basis for advancing personalized medicine and genetic counseling., (© 2024. Higher Education Press.)

Study Question: Could actin-related protein T1 (ACTRT1) deficiency be a potential pathogenic factor of human male infertility?, Summary Answer: A 110-kb microdeletion of the X chromosome, only including the ACTRT1 gene, was identified as responsible for infertility in two Chinese males with sperm showing acrosomal ultrastructural defects and fertilization failure., What Is Known Already: The actin-related proteins (e.g. ACTRT1, ACTRT2, ACTL7A, and ACTL9) interact with each other to form a multimeric complex in the subacrosomal region of spermatids, which is crucial for the acrosome-nucleus junction. Actrt1-knockout (KO) mice are severely subfertile owing to malformed sperm heads with detached acrosomes and partial fertilization failure. There are currently no reports on the association between ACTRT1 deletion and male infertility in humans., Study Design, Size, Duration: We recruited a cohort of 120 infertile males with sperm head deformations at a large tertiary hospital from August 2019 to August 2023. Genomic DNA extracted from the affected individuals underwent whole exome sequencing (WES), and in silico analyses were performed to identify genetic variants. Morphological analysis, functional assays, and ART were performed in 2022 and 2023., Participants/materials, Setting, Methods: The ACTRT1 deficiency was identified by WES and confirmed by whole genome sequencing, PCR, and quantitative PCR. Genomic DNA of all family members was collected to define the hereditary mode. Papanicolaou staining and electronic microscopy were performed to reveal sperm morphological changes. Western blotting and immunostaining were performed to explore the pathological mechanism of ACTRT1 deficiency. ICSI combined with artificial oocyte activation (AOA) was applied for one proband., Main Results and the Role of Chance: We identified a whole-gene deletion variant of ACTRT1 in two infertile males, which was inherited from their mothers, respectively. The probands exhibited sperm head deformations owing to acrosomal detachment, which is consistent with our previous observations on Actrt1-KO mice. Decreased expression and ectopic distribution of ACTL7A and phospholipase C zeta were observed in sperm samples from the probands. ICSI combined with AOA effectively solved the fertilization problem in Actrt1-KO mice and in one of the two probands., Limitations, Reasons for Caution: Additional cases are needed to further confirm the genetic contribution of ACTRT1 variants to male infertility., Wider Implications of the Findings: Our results reveal a gene-disease relation between the ACTRT1 deletion described here and human male infertility owing to acrosomal detachment and fertilization failure. This report also describes a good reproductive outcome of ART with ICSI-AOA for a proband., Study Funding/competing Interest(s): This work was supported by the Chongqing medical scientific research project (Joint project of Chongqing Health Commission and Science and Technology Bureau, 2023MSXM008 and 2023MSXM054). There are no competing interests to declare., Trial Registration Number: N/A., (© The Author(s) 2024. Published by Oxford University Press on behalf of European Society of Human Reproduction and Embryology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Research Question: Is the novel homozygous nonsense variant of AK7 associated with multiple morphological abnormalities of the sperm flagella (MMAF), a specific type of oligoasthenoteratozoospermia leading to male infertility?, Design: Whole-exome sequencing and Sanger sequencing were performed to identify potential gene variants. Immunoblotting and immunofluorescence were applied to confirm the relationship between mutated genes and disease phenotypes. The concentration of reactive oxygen species and the rate of apoptosis were measured to evaluate the mitochondrial function of spermatozoa. Transmission electron microscopy and scanning electron microscopy were employed to observe sperm ultrastructure., Results: A novel homozygous nonsense variant of AK7, c.1153A>T (p. Lys385*), was identified in two infertile siblings with asthenoteratozoospermia through whole-exome sequencing. Both immunoblotting and immunofluorescence assays showed practically complete absence of AK7 in the patient's spermatozoa. Additionally, the individual with the novel AK7 variant exhibited a phenotype characterized by severe oxidative stress and apoptosis caused by mitochondrial metabolic dysfunction of spermatozoa. Notably, remarkable flagellar defects with multiple axonemes in uniflagellate spermatozoa, accompanied by mitochondrial vacuolization, were observed; this has not been reported previously in patients with other AK7 variants., Conclusions: This study found that a novel identified homozygous nonsense variant of AK7 may be associated with MMAF-related asthenoteratozoospermia. The observed functional associations between mitochondria and sperm flagellar assembly provide evidence for potential mutual regulation between AK7 and flagella-associated proteins during spermatogenesis., (Copyright © 2023 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The study investigated midpiece defects in sperm from a 5-year-old Brangus bull with a high rate of semen batch rejection, due to morphologically abnormal sperm, with no reduction in sperm kinematics. A comprehensive evaluation was conducted over a 16-month period, involving 28 ejaculates. Notably, despite the high proportion of midpiece defects (average 37.73%, from 3% to 58%), the study revealed stable sperm production, with no discernible differences in the kinematic data before and after cryopreservation. Electron microscopy identified discontinuities in the mitochondrial sheath, characteristic of midpiece aplasia (MPA). The anomalies were attributed to be of genetic origin, as other predisposing factors were absent. Additionally, the electron microscopy unveiled plasma membrane defects, vacuoles and chromatin decondensation, consistent with previous findings linking acrosome abnormalities with midpiece defects. The findings underscored the necessity of conducting thorough laboratory evaluations before releasing cryopreserved semen for commercialization. Despite substantial morphological alterations, the initial semen evaluation data indicated acceptable levels of sperm kinematics, emphasizing the resilience of sperm production to severe morphological changes. This case report serves as a contribution to the understanding of midpiece defects in bull sperm, emphasizing the need for meticulous evaluation and quality control in semen processing and commercialization., (© 2024 Wiley‐VCH GmbH. Published by John Wiley & Sons Ltd.)

Variations in the dynein axonemal heavy chain gene, dynein axonemal heavy chain 6 ( DNAH6 ), lead to multiple morphological abnormalities of the flagella. Recent studies have reported that these deficiencies may result in sperm head deformation. However, whether DNAH6 is also involved in human acrosome biogenesis remains unknown. The purpose of this study was to investigate DNAH6 gene variants and their potential functions in the formation of defective sperm heads and flagella. Whole-exome sequencing was performed on a cohort of 375 patients with asthenoteratozoospermia from the First Affiliated Hospital of Anhui Medical University (Hefei, China). Hematoxylin and eosin staining, scanning electron microscopy, and transmission electron microscopy were performed to analyze the sperm morphology and ultrastructure. Immunofluorescence staining and Western blot analysis were conducted to examine the effects of genetic variants. We identified three novel deleterious variants in DNAH6 among three unrelated families. The absence of inner dynein arms and radial spokes was observed in the sperm of patients with DNAH6 variants. Additionally, deficiencies in the acrosome, abnormal chromatin compaction, and vacuole-containing sperm heads were observed in these patients with DNAH6 variants. The decreased levels of the component proteins in these defective structures were further confirmed in sperm from patients with DNAH6 variants using Western blot. After intracytoplasmic sperm injection (ICSI) treatment, the partner of one patient with a DNAH6 variant achieved successful pregnancy. Overall, novel variants in DNAH6 genes that contribute to defects in the sperm head and flagella were identified, and the findings indicated ICSI as an effective clinical treatment for such patients., (Copyright © 2023 Copyright: © The Author(s)(2023).)

Background: Analysis of sperm morphology defects (amorphous heads, abnormal acrosome, etc.) is useful for estimating the efficiency of spermiogenesis and sperm maturation. An advanced paternal age (more than 40 years) is associated with decreasing sperm count and reduced motility; however, there is little information on the effect of aging relating to sperm morphological defects. Moreover, searching for stable combinations of certain morphological defects in the same sperm can be useful for better understanding spermiogenesis. The aim of the study was to investigate age-related changes in sperm morphology and the prevalence of certain combinations of sperm morphological defects in men from the general population., Methods: Sperm morphology was assessed in 1266 volunteers from the Russian urban general population in different age groups (18-19, 20-24, 25-29, 30-34, 35-40, and over 40 years old). Two hundred sperm were evaluated from each semen sample (about 250 thousand spermatozoa in total). Sperm defects were classified according to the WHO laboratory manual (WHO, 2010). The total percentage of each sperm defect and the frequency of different combinations of sperm morphological anomalies for each age group were counted. Additionally, a similar analysis was performed for the groups of normospermia and pathozoospermia., Results: The frequency of coiled and short sperm tails increased in men over 40 years old compared to younger subjects; however, aging did not affect the percentage of morphologically normal sperm. It was shown that the combination of a misshaped head (amorphous, pyriform, and elongated) with a postacrosomal vacuole, acrosome defect, excess residual cytoplasm, or any anomaly of the midpiece or tail in the same spermatozoon were not random combinations of independent solitary defects. The increased frequency of combinations of coiled tails with amorphous, elongated, or vacuolated heads was observed in men older than 40 years. Sperm morphological defects, such as severely deformed heads (pyriform, elongated, and round) were more common in men with pathozoospermia compared to normospermic subjects., Conclusions: An age-related impairment in sperm morphology was found. Stable combinations of head defects with anomalies in the acrosome, midpiece or tail suggest that these defects may be the result of a general violation in the morphogenetic mechanism., Competing Interests: The authors declare no conflict of interest., (© 2023 The Author(s). Published by IMR Press.)

As a common metabolic disorder, hyperglycemia (HG) affects and disrupts the physiology of various systems in the body. Transplantation of mesenchymal stem cells (MSCs) has been used to control the complications of disease. Most of the therapeutic properties of MSCs are attributed to their secretome. This study aimed to investigate the effects of conditioned media extracted from sole or caffeine pre-treated bone-marrow-derived MSCs on hyperglycemia-induced detrimental impact on some aspects of reproduction. The HG was induced by intraperitoneally injection of streptozotocin (65 mg/kg) and nicotinamide (110 mg/kg). Twenty-four male Wistar rats (190 ± 20 g) were divided into control, HG, and the hyperglycemic groups receiving conditioned media of proliferated MSCs solely (CM) or MSCs pre-treated with caffeine (CCM). During the 49-day treatment, body weight and blood glucose were measured weekly. Finally, HbA1c, spermatogenesis development, sperm count, morphology, viability, motility, chromatin condensation, and DNA integrity were examined. Also, testicular total antioxidant capacity (TAC), malondialdehyde, sperm fertilization potential, and pre-implantation embryo development were evaluated. A one-way ANOVA and Tukey's post-hoc tests were used to analyze the quantitative data. The p < 0.05 was considered statistically significant. The CM and with a higher efficiency, the CCM remarkably (p < 0.05) improved body weight and HG-suppressed spermatogenesis, enhanced sperm parameters, chromatin condensation, DNA integrity, and TAC, reduced HbA1c, sperm abnormalities, and malondialdehyde, and significantly improved pre-implantation embryo development versus HG group. The conditioned media of MSCs solely (CM) and more effectively after pre-treatment of MSCs with caffeine (CCM) could improve spermatogenesis development, sperm quality, pre-implantation embryo development, and testicular global antioxidant potential during hyperglycemia., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2023 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

We present two cases of infertile males with teratozoospermia stemming from chromosome 17 translocation. The patients present karyotypes that have not been previously reported. Genes located on breakpoints (17p11.2, 9q31, and 11p15) were analysed to find the probable mechanism affecting sperm morphology. Our results suggest that ALKBH5, TOP3A, and LLGL1 interactions may be an underlying cause of abnormal sperm head morphology. Translocation of chromosome 17 occurred in conjunction with chromosome 9 and chromosome 11 translocation in the two cases, resulting in oligozoospermia and asthenozoospermia, respectively. These abnormal phenotypes may involve meiosis- and motility-related genes such as LDHC, DNHD1, UBQLN3, and NUP98. Translocation is thus a risk factor for sperm morphological abnormalities and motility deficiency. The interaction network of 22 genes on breakpoints suggests that they contribute to spermatogenesis as a group. In conclusion, this study highlighted the importance of investigating genes linked to sperm morphology, together with chromosome 17 translocation and reproductive risks. For patients interested in screening before a future pregnancy, we recommend preimplantation genetic diagnosis to reduce the risk of karyotypically unbalanced foetuses and birth defects., (© 2022 Wiley-VCH GmbH.)

Teratozoospermia is usually associated with defective spermiogenesis and is a disorder with considerable genetic heterogeneity. Although previous studies have identified several teratozoospermia-associated genes, the etiology remains unknown for a majority of affected men. Here, we identified a homozygous missense mutation and a compound heterozygous mutation of CCIN in patients suffering from teratozoospermia. CCIN encodes the cytoskeletal protein Calicin that is involved in the formation and maintenance of the highly regular organization of the calyx of mammalian spermatozoa, and has been proposed to play a role in sperm head structure remodeling during the process of spermiogenesis. Our morphological and ultrastructural analyses of the spermatozoa obtained from all three men harboring deleterious CCIN mutants reveal severe head malformation. Further immunofluorescence assays unveil markedly reduced levels of Calicin in spermatozoa. These patient phenotypes are successfully recapitulated in mouse models expressing the disease-associated variants, confirming the role of Calicin in male fertility. Notably, all mutant spermatozoa from mice and human patients fail to adhere to the zona mass, which likely is the major mechanistic reason for CCIN-mutant sperm-derived infertility. Finally, the use of intra-cytoplasmic sperm injections (ICSI) successfully makes mutated mice and two couples with CCIN variants have healthy offspring. Taken together, our findings identify the role of Calicin in sperm head shaping and male fertility, providing important guidance for genetic counseling and assisted reproduction treatments., Competing Interests: Conflict of interest The authors declare that they have no conflict of interest., (Copyright © 2022 Science China Press. Published by Elsevier B.V. All rights reserved.)

The Valley of Sacco River (VSR) (Latium, Italy) is an area with large-scale industrial chemical production that has led over time to significant contamination of soil and groundwater with various industrial pollutants, such as organic pesticides, dioxins, organic solvents, heavy metals, and particularly, volatile organic compounds (VOCs). In the present study, we investigated the potential impact of VOCs on the spermatozoa of healthy young males living in the VSR, given the prevalent presence of several VOCs in the semen of these individuals. To accomplish this, spermiograms were conducted followed by molecular analyses to assess the content of sperm nuclear basic proteins (SNBPs) in addition to the protamine-histone ratio and DNA binding of these proteins. We found drastic alterations in the spermatozoa of these young males living in the VSR. Alterations were seen in sperm morphology, sperm motility, sperm count, and protamine/histone ratios, and included significant reductions in SNBP-DNA binding capacity. Our results provide preliminary indications of a possible correlation between the observed alterations and the presence of specific VOCs.

Multiple morphological abnormalities of the sperm flagella (MMAF), characteristic with bent, short, coiled, absent, and abnormal caliber flagella, is an important basis of male infertility. Genetic factors account for a large proportion of patients with MMAF. The fibrous sheath interacting protein 2 (FSIP2) has a significant function in the spermatogenesis and flagellar motility. In our study, a novel compound heterozygous mutation (c.1494C > A, p.C498* and c.11020&#95;11024del, p.Tyr3675Cysfs*3) in FSIP2 gene was identified in an infertile male patient with MMAF. H&E staining presented typical MMAF phenotype and thick neck, midpiece in the patient's sperm cells. Transmission electron microscopy observation showed abnormal mitochondrial arrangement and disorganization and dysplastic of the fibrous sheath (FS), which were verified again under light microscopy. Immunofluorescence (IF) analysis of FISP2 expression showed that FSIP2 was absent in the flagellum of the patient's sperm cells. Our findings will be helpful to the precise diagnosis of MMAF and male infertility and enrich the mutational spectrum of FSIP2 gene., (© 2022. Society for Reproductive Investigation.)

Background: Multiple morphological abnormalities of the sperm flagella (MMAF) is a subtype of severe asthenoteratozoospermia with poorly understood genetic etiology. SPAG6 is a core axonemal component that plays a critical role in the formation of cilia and sperm flagella. Previous studies have reported that mutations in SPAG6 cause primary ciliary dyskinesia (PCD), but the association between SPAG6 gene variants and the MMAF phenotype has not yet been described., Methods: We performed whole-exome sequencing (WES) in two unrelated Han Chinese men with MMAF. Sanger sequencing was used to validate the candidate variants. Routine semen analysis was carried out according to the WHO guidelines (5 th Edition). Sperm morphology was assessed using modified Papanicolaou staining. Scanning and transmission electron microscopy (S/TEM) was performed to observe the ultrastructural defects of the sperm flagella. Western blot analysis and immunofluorescence (IF) of spermatozoa were performed to examine the expression of SPAG6 protein. Assisted fertilization with intracytoplasmic sperm injection (ICSI) was applied., Results: Two homozygous SPAG6 variants were identified by WES and Sanger validation in two patients with MMAF phenotype (F1 II-1: c.308C > A, p. A103D; F2 II-1: c. 585delA, p. K196Sfs*6). Semen analysis showed progressive rates of less than 1%, and most of the spermatozoa presented MMAF by Papanicolaou staining. TEM revealed that the overall axonemal ultrastructure was disrupted and primarily presented an abnormal "9 + 0" configuration. No other PCD-related symptoms were found on physical examination and medical consultations, as well as lung CT screening. The level of SPAG6 protein was significantly decreased in the spermatozoa, and IF analysis revealed that SPAG6 staining was extremely weak and discontinuous in the sperm flagella of the two patients. Notably, F1 II-1 and his wife conceived successfully after undergoing ICSI., Conclusions: Our research provides new evidence for a potential correlation between SPAG6 variants and the MMAF phenotype., (© 2022. The Author(s).)

Acrylamide (AA) has been shown to have neurological and reproductive toxicities, but little is known about transgenerational effects of AA. In this study, male C57BL/6 mice were exposed to AA (0.01, 1, 10 μg/mL) and its metabolite glycidamide (GA, 10 μg/mL) in drinking water, which were then mated with unexposed female mice to produce F1 and F2 generations. We found that both AA and GA at high concentrations decreased sperm motility in F0 mice and increased sperm malformation rates in mice from all the three generations. In addition, AA and GA increased sperm reactive oxygen species as well as decreased serum testosterone levels, and increased the escape latency time in exposed mice and their offspring. We further found that AA-induced mRNA expression changes in the hippocampus of F0 mice persist to the F2 generation. In the sperm of F0 mice, AA induced significant DNA methylation changes in genes involved in neural and reproduction; the mRNA expression levels of Dnmt3b, a DNA methyltransferase, were dramatically decreased in the testes of F0 and F1 mice. In conclusion, our study indicates that paternal AA exposure leads to DNA methylation-mediated transgenerational adverse effects on sperm parameters and leaning capability in mice., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Objective: To study the contagiousness of sperm and its influence on fertility after recovery from COVID-19 infection., Design: Prospective cohort study., Setting: University medical center., Patient(s): One hundred twenty Belgian men who had recovered from proven COVID-19 infection., Intervention(s): No intervention was performed., Main Outcome Measure(s): Semen quality was assessed using the World Health Organisation criteria. DNA damage to sperm cells was assessed by quantifying the DNA fragmentation index and the high density stainability. Finally antibodies against SARS-CoV2 spike-1 antigen, nuclear and S1-receptor binding domain were measured by Elisa and chemilumenscent microparticle immunoassays, respectively., Result(s): SARS-CoV-2 RNA was not detected in semen during the period shortly after infection nor at a later time. Mean progressive motility was reduced in 60% of men tested shortly (<1 month) after COVID-19 infection, 37% of men tested 1 to 2 months after COVID-19 infection, and 28% of men tested >2 months after COVID-19 infection. Mean sperm count was reduced in 37% of men tested shortly (<1 month) after COVID-19 infection, 29% of men tested 1 to 2 months after COVID-19 infection, and 6% of men tested >2 months after COVID-19 infection. The severity of COVID-19 infection and the presence of fever were not correlated with sperm characteristics, but there were strong correlations between sperm abnormalities and the titers of SARS-CoV-2 IgG antibody against spike 1 and the receptor- binding domain of spike 1, but not against nucleotide, in serum. High levels of antisperm antibodies developed in three men (2.5%)., Conclusion(s): Semen is not infectious with SARS-CoV-2 at 1 week or more after COVID-19 infection (mean, 53 days). However, couples with a desire for pregnancy should be warned that sperm quality after COVID-19 infection can be suboptimal. The estimated recovery time is 3 months, but further follow-up studies are under way to confirm this and to determine if permanent damage occurred in a minority of men., (Copyright © 2021 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Background: Potassium channels are important for the structure and function of the spermatozoa. As a potassium transporter, the mSlo3 is essential for male fertility as Slo3 knockout male mice were infertile with the series of functional defects in sperm cells. However, no pathogenic variant has been detected in human SLO3 to date. Here we reported a human case with homozygous SLO3 mutation. The function of SLO3 in human sperm and the corresponding assisted reproductive strategy are also investigated., Methods: We performed whole-exome sequencing analysis from a large cohort of 105 patients with asthenoteratozoospermia. The effects of the variant were investigated by quantitative RT-PCR, western blotting, and immunofluorescence assays using the patient spermatozoa. Sperm morphological and ultrastructural studies were conducted using haematoxylin and eosin staining, scanning and transmission electron microscopy., Results: We identified a homozygous missense variant (c.1237A > T: p.Ile413Phe) in the sperm-specific SLO3 in one Chinese patient with male infertility. This SLO3 variant was rare in human control populations and predicted to be deleterious by multiple bioinformatic tools. Sperm from the individual harbouring the homozygous SLO3 variant exhibited severe morphological abnormalities, such as acrosome hypoplasia, disruption of the mitochondrial sheath, coiled tails, and motility defects. The levels of SLO3 mRNA and protein in spermatozoa from the affected individual were reduced. Furthermore, the acrosome reaction, mitochondrial membrane potential, and membrane potential during capacitation were also afflicted. The levels of acrosome marker glycoproteins and PLCζ1 as well as the mitochondrial sheath protein HSP60 and SLO3 auxiliary subunit LRRC52, were significantly reduced in the spermatozoa from the affected individual. The affected man was sterile due to acrosome and mitochondrial dysfunction; however, intra-cytoplasmic sperm injection successfully rescued this infertile condition., Conclusions: SLO3 deficiency seriously impact acrosome formation, mitochondrial sheath assembly, and the function of K + channels. Our findings provided clinical implications for the genetic and reproductive counselling of affected families., (© 2021. The Author(s).)

Background: Allergy is associated with the loss of tolerance of environmental antigens, combined with a pathological immune response. There were no studies up to date that would show whether the quality of semen decreases in people with allergic diseases., Material and Methods: The research included men who reported to the Gynecological Outpatient Clinic due to reproductive difficulties, defined as the lack of pregnancy after one year of regular intercourse. Semen quality was assessed according to the World Health Organization (WHO) standard. All patients underwent skin prick tests with the most important inhalation allergens (such as hazel, silver birch, mugwort, rye, dog, cat, Dermatophagoides farinae, Dermatophagoides pteronyssinus , alder, Alternaria alternata, Cladosporium herbarum , and grass mix). The data was statistically analyzed., Results: Results of 52 patients aged 25-52 years (34.62 ± 4.96) were analyzed. The mean BMI (Body mass index) was 28.25 (+ -3.77). It was found that 38 men (73%) had increased body weight, and 14 men (26.9%) were obese (BMI > = 30). 13 patients were smokers (25%), and 24 patients (46%) had skin tests positive for at least one inhaled allergen. Sperm tail defects were statistically more significant in patients allergic to birch, rye, cat, alder, and grass. In patients allergic to Alternaria alternata , head defects were statistically more significant ( p < .05). No association was found between allergy to house dust mites, mugwort, hazel, and dogs and the deterioration of semen., Conclusion: Allergy due to inhalation allergens had an influence on the quality of male semen. Further research is necessary to establish the immunological bases of this phenomenon.

Research Question: Male infertility is a global issue worldwide and multiple morphological abnormalities of the sperm flagella (MMAF) is one of the most severe forms of the qualitative sperm defects with a heterogeneous genetic cause that has not been completely understood. Can whole-exome sequencing (WES) reveal novel genetic causes contributing to MMAF in a consanguineous Pakistani family, comprising three infertile brothers?, Design: WES and bioinformatic analysis were conducted to screen potential pathogenic variants. The identified variant was validated by Sanger sequencing in all available family members Transmission electron microscopy analyses was carried out to examine the flagella ultrastructure of spermatozoa from patient., Results: WES and Sanger sequencing identified a novel homozygous stop-gain mutation (ENST00000392644.4, c.182C>G, p.S61X) in ARMC2, which is expected to lead to loss of protein functions. Transmission electron microscopy analyses revealed that the flagellar ultrastructure of the patient's spermatozoa was disorganized along with a complete absence of central pair complex (CPC), suggesting that ARMC2 is involved in the assembly, stability of the axonemal complex, or both, particularly the CPC., Conclusion: We report that a familial stop-gain mutation in ARMC2 is associated with male infertility in humans caused by MMAF accompanied with loss of CPCs and axonemal disorganization. We provide genetic evidence that ARMC2 is essential for human spermatogenesis and its mutation may be pathogenic for MMAF. These findings will improve the knowledge about the genetic basis of MMAF and provide information for genetic counselling of this disease., (Copyright © 2021. Published by Elsevier Ltd.)

Research Question: Male infertility is a widespread symptom in patients with primary ciliary dyskinesia (PCD). PCD-related male infertility is often caused by asthenozoospermia, with barely normal sperm morphology. Multiple morphological abnormalities of the sperm flagella (MMAF) are a major cause of asthenozoospermia, characterized by various malformed morphologies of sperm flagella. To date, a limited number of genes have been suggested to be involved in the pathogenesis of both PCD and MMAF. What other genes associated with both PCD and MMAF are waiting to be discovered?, Design: Whole-exome sequencing (WES) was performed to identify the pathogenic mutation associated with MMAF in a PCD patient. Peripheral venous blood and semen samples were collected from the PCD patient. Transmission electron microscopy (TEM), immunofluorescence staining and western blotting were conducted to confirm the pathogenicity of the identified mutation., Results: A novel homozygous mutation in CCDC39, c.983 T>C (p. Leu328Pro), was identified in two PCD-affected siblings of a consanguineous family showing a typical PCD phenotype, while the proband was infertile, which is associated with characterized MMAF. Furthermore, TEM revealed the abnormal ultrastructure of the patient's sperm flagella. Moreover, immunofluorescence staining revealed that CCDC39 was almost undetectable in the spermatozoa, which was further confirmed by western blotting. The outcome of intracytoplasmic sperm injection (ICSI) in the patient with the CCDC39 mutation was also favourable., Conclusion: This study demonstrates that a novel loss-of-function mutation of CCDC39 is involved in the pathogenesis of PCD and MMAF and initially reported that ICSI treatment has a good outcome. Therefore, the novel variant of CCDC39 contributes to the genetic diagnosis, counselling and treatment of male infertility in PCD patients with MMAF phenotype., (Copyright © 2021. Published by Elsevier Ltd.)

The testicular deficiency associated with exposure to three widely used insecticides in Egyptian agriculture was evaluated. Animals were orally treated with sub-lethal dose (1/50 of the oral LD 50 ) of cypermethrin (CYP), imidacloprid (IMC), and chlorpyrifos (CPF) at 5, 9 and 1.9 mg/kg/day, respectively, five times a week for one month. The CYP, IMC, and CPF exposure resulted in a significant decline in animal body weight, sperm count, motility, normality, and viability with increased head and tail deformities. Significant reduction in serum testosterone, luteinizing hormone (LH), follicle-stimulating hormone (FSH), testis superoxide dismutase (SOD), and reduced glutathione (GSH) levels. In contrast, catalase (CAT), lipid peroxidation (LPO), and protein carbonyl content (PCC) levels were significantly stimulated. Jointly, obtained results were confirmed by microscopic examination of testis sections. The present data concluded that the CYP, IMC, and CPF have a public health impact and violently interferes with male rat reproductive system., (Copyright © 2021 Elsevier B.V. All rights reserved.)

Purpose: To identify the genetic causes for acephalic spermatozoa syndrome., Methods: Whole-exome sequencing was performed on the proband from a non-consanguineous to identify pathogenic mutations for acephalic spermatozoa syndrome. Quantitative real-time polymerase chain reaction and whole genome sequencing were subjected to detect deletion. The functional effect of the identified splicing mutation was investigated by minigene assay. Western blot and immunofluorescence were performed to detect the expression level and localization of mutant TSGA10 protein., Results: Here, we identified a novel heterozygous splicing mutation in TSGA10 (NM&#95;025244: c.1108-1G > T), while we confirmed that there was a de novo large deletion in the proband. The splicing mutation led to the skipping of the exon15 of TSGA10, which resulted in a truncated protein (p. A370Efs*293). Therefore, we speculated that the splicing mutation might affect transcription and translation without the dosage compensation of a normal allele, which possesses a large deletion including intact TSGA10. Western blot and immunofluorescence demonstrated that the very low expression level of truncated TSGA10 protein led the proband to present the acephalic spermatozoa phenotype., Conclusion: Our finding expands the spectrum of pathogenic TSGA10 mutations that are responsible for ASS and male infertility. It is also important to remind us of paying attention to the compound heterozygous deletion in patients from non-consanguineous families, so that we can provide more precise genetic counseling for patients., (© 2021. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The novel coronavirus disease (COVID-19) pandemic is emerging as a global health threat and shows a higher risk for men than women. Thus far, the studies on andrological consequences of COVID-19 are limited. To ascertain the consequences of COVID-19 on sperm parameters after recovery, we recruited 41 reproductive-aged male patients who had recovered from COVID-19, and analyzed their semen parameters and serum sex hormones at a median time of 56 days after hospital discharge. For longitudinal analysis, a second sampling was obtained from 22 of the 41 patients after a median time interval of 29 days from first sampling. Compared with controls who had not suffered from COVID-19, the total sperm count, sperm concentration, and percentages of motile and progressively motile spermatozoa in the patients were significantly lower at first sampling, while sperm vitality and morphology were not affected. The total sperm count, sperm concentration, and number of motile spermatozoa per ejaculate were significantly increased and the percentage of morphologically abnormal sperm was reduced at the second sampling compared with those at first in the 22 patients examined. Though there were higher prolactin and lower progesterone levels in patients at first sampling than those in controls, no significant alterations were detected for any sex hormones examined over time following COVID-19 recovery in the 22 patients. Although it should be interpreted carefully, these findings indicate an adverse but potentially reversible consequence of COVID-19 on sperm quality., Competing Interests: None

Research Question: Asthenoteratospermia is characterized by malformed spermatozoa with motility defects, which results in male infertility. Multiple morphological abnormalities of the sperm flagella (MMAF) is a hallmark of asthenoteratospermia. The genetic causes of MMAF, however, are unknown in about one-third of cases. Which other MMAF-associated genes are waiting to be discovered?, Design: Whole-exome sequencing was conducted to identify causative genes in a man with MMAF. Immunofluorescence staining and western blot were applied to assess the pathogenicity of the identified variant. Intracytoplasmic sperm injection (ICSI) was used to assist fertilization for the patient with MMAF., Result: Sanger sequencing of the family demonstrated that the infertile man carried a homozygous DNAH17 variant (c. 4810C>T [p.R1604C]). The obviously decreased DNAH17 expression was observed in HEK293T cells transfected with MUT-DNAH17 plasmid compared with cells with WT-DNAH17 plasmid. Immunofluorescence analysis showed that this mutation induced significant decrease in DNAH17 expression, which negatively affected the DNAH8 expression in the patient's spermatozoa. Moreover, the outcome of ICSI in the patient was unsuccessful., Conclusion: Our study revealed a novel homozygous missense mutation in DNAH17 involved in MMAF phenotype. The finding of the novel mutation in DNAH17 enriches the gene variant spectrum of MMAF, further contributing to diagnosis, genetic counselling and prognosis for male infertility., (Copyright © 2021 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.)

The global prevalence of diabetes mellitus is rapidly increasing. This disease is associated with many complications including male reproductive dysfunctions and infertility. Seahorse ( Hippocampus kuda) is a marine teleost fish well known for its beneficial effects on the reproductive system in traditional Chinese medicine books. Recently, several studies have been shown that the enzymatic hydrolysate of seahorse has multiple pharmacological activities. This study aimed to investigate the seahorse peptide hydrolysate (SH) ameliorative effects on the diabetic-induced male reproductive dysfunction in rat models. The in vivo studies were carried out with three different doses of SH (4, 8, and 20 mg/kg) and the diabetes condition was induced by administrating with streptozotocin (35 mg/kg) and fed a 40% high-fat diet. Seahorse hydrolysate (20 mg/kg) inhibited lipid peroxidation, increased antioxidant enzyme activity, and restored seminiferous tubules morphology in testis. Moreover, it improved reproductive dysfunction by increasing the level of testosterone, follicle-stimulating hormone, luteinizing hormone, sperm count, and motility. According to these results, we suggested that SH exhibited amelioration effects on the reproductive dysfunction., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Sperm morphology was once believed as one of the most predictive indicators of pregnancy outcome in assisted reproductive technology (ART). However, the impact of teratozoospermia on in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) outcomes and its offspring remains inconclusive. In order to evaluate the influence of teratozoospermia on pregnancy outcome and newborn status after IVF and ICSI, a retrospective study was conducted. This was a matched case-control study that included 2202 IVF cycles and 2574 ICSI cycles and was conducted at the Reproductive and Genetic Hospital of CITIC-Xiangya in Changsha, China, from June 2013 to June 2018. Patients were divided into two groups based on sperm morphology: teratozoospermia and normal sperm group. The pregnancy outcome and newborn outcome were analyzed. The results indicated that couples with teratozoospermia had a significantly lower optimal embryo rate compared to those with normal sperm morphology in IVF (P = 0.007), while there were no statistically significant differences between the two groups in terms of the fertilization rate, cleavage rate, implantation rate, and pregnancy rate (all P > 0.05). Additionally, teratozoospermia was associated with lower infant birth weight in multiple births after IVF. With regard to ICSI, there was no significant difference in both pregnancy outcome and newborn outcome between the teratozoospermia and normal groups (both P > 0.05). Furthermore, no increase in the risk of birth defects occurred in the teratozoospermia group after IVF/ICSI. Consequently, we believe that teratozoospermia has limited predictive value for pregnancy outcomes in IVF/ICSI, and has little impact on the resulting offspring if multiple pregnancy is avoided., Competing Interests: None

Purpose: The present study sought to investigate the common abnormalities and mtDNA mutations in the sperm of Ghanaian men attending the fertility Clinic at the Korle-Bu Teaching Hospital (KBTH). The study therefore provides a baseline data mtDNA mutations in a cross-section of Ghanaian men on referral to the fertility clinic at the KBTH., Materials and Methods: The semen of 55 men attending the fertility clinic were collected from the Urology and the Obstetrics and Gynaecology Departments of the KBTH. Demographic and clinical data were also collected using questionnaires. Semen analyses were performed and were followed by amplification and purification of mtDNA from total DNA extracted from the semen. Sequencing of the mtDNA amplicons was performed using the next generation sequencer (Illumina-MiSeq)., Results: Asthenozoospermia, oligospermia and oligoasthenoteratozoospermia were observed in 1.79%, 5.36% and 28.57%, respectively, of the study participants. There was no association between drinking and/or smoking and history of gonorrhea infection on sperm status/morphology. A total of 785 point mutations were detected in the non-coding control regions, rRNA genes, tRNA genes and the coding regions of the mtDNA samples from the participants. Amongst these mutations, 16 transition mutations were predominantly detected in the mtDNA samples. Missense mutations that were present in only specific sperm abnormalities were identified and they may contribute to infertility in the study population., Conclusion: The present study has identified various abnormal sperm phenotypes that are prevalent in the study population and provided a baseline data on mtDNA mutations in the spermatozoa of the patients. A wide range of sperm abnormalities were detected in the study population with no association with life style or history of gonorrhea infection. The mtDNA point mutations detected in the selected genes that were analysed were mostly transition mutations. These transition mutations might be critical for the development of abnormal sperm phenotypes underlying male infertility in the Ghanaian population., Competing Interests: The authors have declared that no competing interests exist.

Purpose: To study the potential paternal contribution to aneuploidies in the man of a couple who obtained trisomic embryos with natural and assisted fertilization., Methods: Semen analysis, immunofluorescence for localization of tubulin and centrin 1, transmission electron microscopy (TEM), and fluorescence in situ hybridization (FISH) analysis for chromosomes 18 and 9 were performed. Sperm of fertile men were used as controls., Results: The percentages of sperm motility and normal forms were decreased. The percentages of sperm with tail reduced in dimension, headless tails, coiled tails, and altered head-tail junction were significantly higher (P < 0.01) in the patient than in controls, whereas the percentage of sperm with a normal centrin 1 localization (two spots in the centriolar area) was significantly reduced (P < 0.01) in the patient. Immunofluorescence with anti-tubulin antibody showed that in most of the patient's sperm connecting pieces (83.00 ± 1.78%), two spots were present, indicating prominent proximal centriole/centriolar adjunct and evident distal centriole, whereas controls' sperm displayed a single spot, indicating the proximal centriole. The percentage of sperm with two spots was significantly higher (P < 0.01) in the patient than in controls. TEM analysis showed that centriolar adjuncts of the patient's sperm were significantly longer (721.80 ± 122.26 nm) than in controls' sperm (310.00 ± 64.11 nm; P < 0.001). The aneuploidy frequencies of the patient's sperm, detected by FISH analysis, were increased with respect to controls., Conclusion: A paternal contribution to sperm aneuploidies cannot be excluded since the patient's sperm showed altered morphology, immature centriolar adjunct, presence of evident distal centriole, scarce presence of centrin 1, and high aneuploidy frequency.

Purpose: To evaluate the correlation between the DNA Fragmentation Index (DFI) and sperm morphology in patients undergoing ICSI, as a predictive parameter in reproductive outcomes., Methods: A retrospective study was conducted on 125 infertile patients enrolled in a fertility clinic. Seminal characteristics were measured following the WHO guidelines (2010) for the examination of the seminal fluid. After collecting motile sperm population by pellet swim up, DFI was calculated and simultaneously associated with sperm morphology using in situ TUNEL assay and an image analyzer software in at least 250 spermatozoa for each patient., Results: All subjects were divided into two groups according to a cutoff established, by choice, of the sperm DFI (15%): group A (< 15%) consisting of 65 patients and group B (≥ 15%) of 60 patients. Data were analyzed using non-parametric statistical methods. The results demonstrate that there is no statistical difference between the two groups in seminal characteristics. The collective data show a high significant correlation, suggesting that spermatozoa with abnormal morphology are the best candidates to contain DNA damage (p < 0.001). Also, when group A is compared with group B, an increased percentage of morphologically normal spermatozoa with fragmented DNA was observed in patients, with DFI values ≥ 15% (p < 0.001)., Conclusion: These results are aimed at providing an exact value of DFI in morphologically normal spermatozoa, which will be helpful to the embryologist in evaluating the risk of transferring, during the ICSI procedure, a spermatozoon whit normal morphology but fragmented DNA.

Purpose: Acephalic spermatozoa syndrome (ASS) is known as a severe type of teratozoospermia, defined as semen composed of mostly headless spermatozoa that affect male fertility. In this regard, this systematic review aimed to discuss gene variants associated with acephalic spermatozoa phenotype as well as the clinical outcomes of intracytoplasmic sperm injection (ICSI) treatment for the acephalic spermatozoa-associated male infertility., Methods: A systematic search was performed on PubMed, Embase, Scopus, and Ovid databases until May 17, 2020. This systematic scoping review was reported in terms of the Preferred Reporting Items for Systematic reviews and Meta-Analyses extension for Scoping Reviews (PRISMA-ScR) statement., Results: Twenty articles were included in this systematic review. Whole-exome and Sanger sequencing have helped in the identification of variants in SUN5, PMFBP1, BRDT, TSGA10, DNAH6, HOOK1, and CEP112 genes as possible causes of this phenotype in humans. The results of the ICSI are conflicting due to both positive and negative reports of ICSI outcomes., Conclusion: ASS has a genetic origin, and several genetic alterations related to the pathogenesis of this anomaly have been recently identified. Notably, only SUN5 and PMFBP1 mutations are well-known to be implicated in ASS. Accordingly, more functional studies are needed to confirm the pathogenicity of other variants. ICSI could provide a promising treatment for acephalic spermatozoa-associated male infertility. Besides the importance of sperm head-tail junction integrity, some other factors, whether within the sperm cell or female factors, may be involved in the ICSI outcome.

Nonylphenol (NP) is an environmental contaminant, which adversely affects the male fertility due to endocrine disruption and generation of oxidative stress. The current research was planned to assess the effects of nobiletin (NOB), a polymethoxyflavone, on NP-induced testicular damages. Twenty-four male rats were divided into 4 groups: control (0.1% DMSO), NP group (50 mg/kg), NP+NOB group (50 mg/kg + 25 mg/kg), and NOB group (25 mg/kg). Our results revealed that NP brought down the activities of catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GPx) and glutathione reductase (GSR), while elevated the level of thiobarbituric acid reactive substances (TBARS). Additionally, NP decreased the level of follicle-stimulating hormone (FSH), luteinizing hormone (LH), plasma testosterone, daily sperm production (DSP), epididymal sperm count, viability, motility, gene expression of testicular steroidogenic enzymes (StAR, 3β-HSD and 17β-HSD) and anti-apoptotic protein (Bcl-2), as well as number of spermatogenic cells belonging to various stages. Whereas, sperm (head, mid-piece/neck and tail) abnormalities, expression of apoptotic proteins (Bax and caspase-3), and histopathological damages were increased. However, NOB remarkably reversed all the damages caused by NP. Therefore, it is deduced that NOB could be used as a potential therapeutic to counter the NP-prompted oxidative stress and apoptotic damages in testes.

Multiple morphological abnormalities of the sperm flagella (MMAF) is a severe type of teratozoospermia. In this study, whole-exome sequencing was conducted on 55 patients with MMAF, and biallelic mutations of CFAP58 were identified in two patients. The variants are rare and pathogenic, and CFAP58 was absent in the CFAP58-mutated sperm. The F037/II:1 couple benefited from intracytoplasmic sperm injection (ICSI). This study further indicated that CFAP58 is a pathogenic gene associated with MMAF and ICSI is an effective treatment., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Although it is reported that olanzapine (OLZ), which is an atypical antipsychotic drug, causes sexual dysfunction in men, it is noteworthy that there is not any study evaluating the toxic effects of OLZ on the male reproductive system. In the scope of this research, it was aimed to assess the reproductive toxic effects of OLZ by oral administration of 2.5, 5, or 10 mg/kg of it to male rats for 28 days. For this purpose, sperm concentration, motility and morphology, and DNA damage were determined, and histopathological examination of testis tissue was carried out in rats. Also, the levels of serum follicle-stimulating hormone (FSH), luteinizing hormone (LH), and testosterone, which play roles in the regulation of reproductive functions, and the levels of glutathione (GSH), catalase (CAT), superoxide dismutase (SOD) and malondialdehyde (MDA) which play roles in reproductive pathologies as oxidative stress biomarkers, were determined. According to the results, normal sperm morphology was decreased in 5 ve 10 mg/kg OLZ-administered groups, and pathological findings were evident in the testicular structure of the OLZ-administered group when compared with the control group. It was determined that serum LH, FSH, and testosterone levels were decreased in the OLZ-administered group. Also, decreases of GSH levels in testis tissue were determined and evaluated as the markers of the oxidative stress induced by OLZ in the testis. In conclusion, it was determined that reproductive toxic effects were induced in rats by OLZ administration. This pathology was accompanied by alterations of the hormone levels and testicular oxidative stress.

Cisplatin is an effective anticancer drug used against a variety of cancers. The full therapeutic potential of cisplatin is often hampered due to concurrent development of various side effects in the hosts. Rutin, a naturally occurring bioflavonoid shows several pharmacological activities. It has been earlier reported by us that rutin and cisplatin in combination show better antitumor activity against murine ascites Dalton's lymphoma. As cisplatin is given to cancer-bearing hosts only, the present study was undertaken to explore the histoprotective effect of rutin against some toxicities induced by cisplatin in tumor-bearing mice. Cisplatin treatment caused severe damages in tissue architecture such as degenerated hepatocytes with nuclear condensation and sinusoidal dilatation in the liver, glomerular deterioration, infiltration of cells, and tubular congestion in the kidney, and vacuolization of Sertoli cells or dense granules in the cytoplasm and damaged seminiferous tubules in the testes. In the rutin plus cisplatin combination-treated mice, all the abnormal tissue architectural features were decreased. Further, as compared to cisplatin treatment, combination treatment did not show any significant elevation in the liver functional biomarkers (serum aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase) and renal functional biomarkers (serum urea and creatinine levels). The combination treatment reduced the sperm abnormalities also as compared to the cisplatin alone treatment. The in vitro hemolysis assay of red blood cells and scanning electron microscopy revealed that combination treatment lessened the cisplatin-induced hemolysis and abnormalities in RBCs. Thus, the present findings demonstrate that rutin has histoprotective ability against cisplatin-induced toxicities in tumor-bearing mice.

The declining rate of male fertility is a growing concern. Tributyltin (TBT) is a well-known endocrine disruptor (ED), that induces imposex in female gastropods and is widely used in various industrial applications. The aim of this study was to evaluate the toxic effects of TBT on the testes of adult albino rats and the possible role of omega-3. Forty two adult male albino rats were divided into five groups; control group (Group I) and four experimental groups: omega-3 treated group, TBT treated group, TBT & omega-3 treated group and follow up group. At the end of the study, the rats were subjected to biochemical, histological, immunohistochemical staining for Ki-67 and seminal examinations. Our results clarfied that TBT induced a significant decrease in testosterone, FSH, LH and serum glutathione peroxidase levels and a significant increase in the serum Malondialdehyde as compared to the control group. Tributyltin induced disorganization and shrinkage of seminiferous tubules, apoptosis, cellular damage and marked reduction in the germinal epithelium. A significant decrease in the cell proliferation and arrested spermatogenesis were also detected. Seminal analysis of TBT group showed a significant affection of all parameters as compared to other groups. Omega-3 ameliorated all of these hazardous effects. Follow up group still showed toxic effects. In conclusion, TBT has a toxic effect on the testis. Increased testicular oxidative stress, cellular damage and arrest of spermatogenesis with attenuation in antioxidant defenses are all contributing factors. Omega-3 can protect against TBT induced reproductive toxicity.

Heat stress reduces the reproductive capacity of bulls raised in tropical climate. However, the reestablishment of scrotal thermoregulation and the dynamics of sperm defects emergence after stress are not completely known in buffaloes. Thus, the study aimed to evaluate the effect of short-term heat stress over scrotal thermoregulation and sperm attributes, relating them to spermatogenesis stages. Five buffalo bulls went through scrotal insulation during 48 h (from day 0 to day 2). Semen samples were collected every 7 days (from day -7 to day 49) and analyzed about the progressive motility, viability, and sperm morphology. Heat stress significantly destabilized scrotal thermoregulation (P < 0.001). Scrotal temperature was from 4.2 to 6.3 °C lower than the core body temperature, except on insulation days (P < 0.001), and returned to the basal condition five days after the removal of the stressing stimulus. More significant deleterious effects were observed in sperm morphology than in cell concentration, motility, and viability. The chronology of morphologic defects expression demonstrated tail defects (days 7-14), cytoplasmic droplets (days 14-28), and head defects (day 28), returning to pre-insulation condition 35 days after the thermal challenge. Thus, hyperthermia harmed more intensely spermatozoa in epididymal transit, elongated spermatids, and secondary spermatocytes. It is concluded that water buffalo bulls present a peculiar manifestation of sperm morphology after short-term stress, indicating an important difference related to the bovine species. Therefore, during the andrological evaluation of buffalo bulls, it is necessary to avoid the allometric extrapolation between these species., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Globozoospermia is a rare phenotype of primary male infertility inducing the production of round-headed spermatozoa without acrosome. Anomalies of DPY19L2 account for 50-70% of all cases and the entire deletion of the gene is by far the most frequent defect identified. Here, we present a large cohort of 69 patients with 20-100% of globozoospermia. Genetic analyses including multiplex ligation-dependent probe amplification, Sanger sequencing and whole-exome sequencing identified 25 subjects with a homozygous DPY19L2 deletion (36%) and 14 carrying other DPY19L2 defects (20%). Overall, 11 deleterious single-nucleotide variants were identified including eight novel and three already published mutations. Patients with a higher rate of round-headed spermatozoa were more often diagnosed and had a higher proportion of loss of function anomalies, highlighting a good genotype phenotype correlation. No gene defects were identified in patients carrying < 50% of globozoospermia while diagnosis efficiency rose to 77% for patients with > 50% of globozoospermia. In addition, results from whole-exome sequencing were scrutinized for 23 patients with a DPY19L2 negative diagnosis, searching for deleterious variants in the nine other genes described to be associated with globozoospermia in human (C2CD6, C7orf61, CCDC62, CCIN, DNAH17, GGN, PICK1, SPATA16, and ZPBP1). Only one homozygous novel truncating variant was identified in the GGN gene in one patient, confirming the association of GGN with globozoospermia. In view of these results, we propose a novel diagnostic strategy focusing on patients with at least 50% of globozoospermia and based on a classical qualitative PCR to detect DPY19L2 homozygous deletions. In the absence of the latter, we recommend to perform whole-exome sequencing to search for defects in DPY19L2 as well as in the other previously described candidate genes.

Background: Spermatogenesis is a complex biological process highlighted by synthesis and activation of proteins that regulate meiosis and cellular differentiation occur during spermatogenesis. 14-3-3 proteins are adaptor proteins that play critical roles in kinase signaling, especially for regulation of cell cycle and apoptosis in eukaryotic cells. There are seven isoforms of the 14-3-3 family proteins encoded by seven genes (β, ε, γ, η, θ/τ, ζ and σ). 14-3-3 isoforms have been shown to have many interacting partners in several tissues including testis., Objective: While it is known that 14-3-3 proteins are expressed in the functions of testis and spermatozoon, the role for each of the seven isoforms is not known. In this study, we investigated the roles of 14-3-3η and 14-3-3ε isoforms in spermatogenesis., Materials and Methods: To study the in vivo function of 14-3-3η and 14-3-3ε in spermatogenesis, we generated testis-specific and global knockout mice for each of 14-3-3η and 14-3-3ε isoforms (CKO and GKO, respectively). Computer-assisted semen analysis was used to assess sperm motility, while immunohistochemical studies were conducted to check spermatogenesis., Results: Although both 14-3-3η and 14-3-3ε isoforms were present in mouse testis, only the expression of 14-3-3ε, but not 14-3-3η, was detected in spermatozoa. Mice lacking 14-3-3η were normal and fertile while 14-3-3ε CKO and GKO males showed infertility. Low sperm count with higher abnormal spermatozoa was seen in 14-3-3ε CKO mice. The motility of 14-3-3ε knockout spermatozoa was lower than that of the control. A reduction in the phosphorylation of both glycogen synthase kinase 3 and PP1γ2 was also seen in spermatozoa from 14-3-3ε CKO mice, suggesting a specific role of 14-3-3ε in spermatogenesis, sperm motility, and fertility., Discussion and Conclusion: This is the first demonstration that of the seven 14-3-3 isoforms, 14-3-3ε is essential for normal sperm function and male fertility., (© 2020 American Society of Andrology and European Academy of Andrology.)

Spermatozoa contain highly specialized structural features reflecting unique functions required for fertilization. Among them, the flagellum is a sperm-specific organelle required to generate the motility, which is essential to reach the egg. The flagellum integrity is, therefore, critical for normal sperm function and flagellum defects consistently lead to male infertility due to reduced or absent sperm motility defined as asthenozoospermia. Multiple morphological abnormalities of the flagella (MMAF), also called short tails, is among the most severe forms of sperm flagellum defects responsible for male infertility and is characterized by the presence in the ejaculate of spermatozoa being short, coiled, absent and of irregular caliber. Recent studies have demonstrated that MMAF is genetically heterogeneous which is consistent with the large number of proteins (over one thousand) localized in the human sperm flagella. In the past 5 years, genomic investigation of the MMAF phenotype allowed the identification of 18 genes whose mutations induce MMAF and infertility. Here we will review information about those genes including their expression pattern, the features of the encoded proteins together with their localization within the different flagellar protein complexes (axonemal or peri-axonemal) and their potential functions. We will categorize the identified MMAF genes following the protein complexes, functions or biological processes they may be associated with, based on the current knowledge in the field.

Identifying the genes causing male infertility is important to increase our biological understanding as well as the diagnostic yield and clinical relevance of genetic testing in this disorder. While significant progress has been made in some areas, mainly in our knowledge of the genes underlying rare qualitative sperm defects, the same cannot be said for the genetics of quantitative sperm defects. Technological advances and approaches in genomics are critical for the process of disease gene identification. In this review we highlight the impact of various technological developments on male infertility gene discovery as well as functional validation, going from the past to the present and the future. In particular, we draw attention to the use of unbiased genomics approaches, the development of increasingly relevant functional assays and the importance of large-scale international collaboration to advance disease gene identification in male infertility.

Importin α proteins play a central role in the transport of cargo from the cytoplasm to the nucleus. In this study, we observed that male knock-out mice for importin α4, which is encoded by the Kpna4 gene (Kpna4 -/- ), were subfertile and yielded smaller litter sizes than those of wild-type (WT) males. In contrast, mice lacking the closely related importin α3 (Kpna3 -/- ) were fertile. In vitro fertilization and sperm motility assays demonstrated that sperm from Kpna4 -/- mice had significantly reduced quality and motility. In addition, acrosome reaction was also impaired in Kpna4 -/- mice. Transmission electron microscopy revealed striking defects, including abnormal head morphology and multiple axoneme structures in the flagella of Kpna4 -/- mice. A five-fold increase in the frequency of abnormalities in Kpna4 -/- mice compared to WT mice indicates the functional importance of importin α4 in normal sperm development. Moreover, Nesprin-2, which is a component of the linker of nucleus and cytoskeleton complex, was expressed at lower levels in sperm from Kpna4 -/- mice and was localized with abnormal axonemes, suggesting incorrect formation of the nuclear membrane-cytoskeleton structure during spermiogenesis. Proteomics analysis of Kpna4 -/- testis showed significantly altered expression of proteins related to sperm formation, which provided evidence that genetic loss of importin α4 perturbed chromatin status. Collectively, these findings indicate that importin α4 is critical for establishing normal sperm morphology in mice, providing new insights into male germ cell development by highlighting the requirement of importin α4 for normal fertility., (© 2020 Federation of American Societies for Experimental Biology.)

Non-obstructive azoospermia is a distinct diagnosis within male infertility in which no sperm is found in the ejaculate as a result of spermatogenesis failure. Because of the increased prevalence of genetic abnormalities in men with non-obstructive azoospermia, male infertility guidelines recommend screening for karyotype abnormalities and Y chromosome microdeletions in this population. Numerous karyotype abnormalities may be present resulting in impaired spermatogenesis, including: Klinefelter syndrome, translocations, and deletions. Y chromosome microdeletions of the AZFa, AZFb, AZFc subregions all can also result in non-obstructive azoospermia with the possibility of sperm being present if only the AZFc subregion is deleted. While these are the two genetic tests recommended by the guidelines, nearly 50%-80% of non-obstructive azoospermia has no identifiable cause and is deemed idiopathic. Several other genetic defects can lead to non-obstructive azoospermia including Kallmann syndrome, mild androgen insensitivity syndrome, and TEX11. While many additional candidate genes have been proposed, many have yet to be verified or are so infrequent in the population that screening is cost-ineffective. Much research is still required in the genetics of non-obstructive azoospermia and will require multi-institutional initiatives to better understand the genetics of condition., Competing Interests: Declaration of competing interest None., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Exposure to arsenic, which occurs via various routes, can cause reproductive toxicity. However, the mechanism for arsenic-induced reproductive disorders in male mice has not been extensively investigated. Here, 6-week-old male mice were dosed to 0, 5, 10, or 20 ppm sodium arsenite (NaAsO 2 ), an active form of arsenic, in drinking water for six months. For male mice exposed to arsenite, fertility was lower compared to control mice. Moreover, for exposed mice, there were lower sperm counts, lower sperm motility, and higher sperm malformation ratios. Further, the mRNA and protein levels of the gonadotropin-regulated testicular RNA helicase (DDX25) and chromosome region maintenance-1 protein (CRM1), along with proteins associated with high mobility group box 2 (HMGB2), phosphoglycerate kinase 2 (PGK2), and testicular angiotensin-converting enzyme (tACE) were lower. Furthermore, chronic exposure to arsenite led to lower H2A ubiquitination (ubH2A); histone H3 acetylation K18 (H3AcK18); and histone H4 acetylations K5, K8, K12, and K16 (H4tetraAck) in haploid spermatids from testicular tissues. These alterations disrupted deposition of protamine 1 (Prm1) in testes. Overall, the present results indicate that the ubiquitination and acetylation of histones is involved in the spermiogenesis disorders caused by chronic exposure to arsenite, which points to a previously unknown connection between the modification of histones and arsenite-induced male reproductive toxicity., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Klinefelter syndrome (KS) is defined as the presence of one or more extra "X" chromosome in a male patient. It affects approximately 1 in 600 newborn males and the most common chromosomal abnormality, leading to male hypogonadism and infertility. There is a lack of data supporting best practices for KS patients' care. In this paper we review controversial issues in KS research ranging from mechanisms of variation in KS phenotype to abnormalities resulting in reduced sperm production to successful sperm retrieval disparities after testicular sperm extraction (TESE). Translation to live birth and offspring health is also examined. Finally, medical therapies used to optimize the hormonal status and chances of fertility in KS patients are reviewed. We will also discuss the experimental spermatogonial stem cell (SSC) treatments, which are considered the future for TESE negative patients., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Vitamin E (VE) plays numerous important roles in mammals because of its antioxidant activity. As a result, VE deficiency (VED) leads to the dysfunction of central nervous, reproductive, and immune systems. However, few studies have reported the effects of VED on the male reproductive system. In this study, we investigated the effects of VED on male reproductive function and examined its relationship to involution in the male reproductive system with aging. We fed a VED or control diet to 4-week-old mice for 12 or 24 weeks. Following the histopathological analysis of reproductive organs, we found seminiferous tubules with exfoliation in the VED groups, and its frequency was significantly increased compared with the controls. Additionally, in the epididymis, a decrease in spermatozoa and an increase in apoptotic germ cells were observed in the VED groups compared with the controls. By Papanicolaou staining, we also found an increase in the proportion of sperm with abnormal morphology in the VED groups compared with the controls. These reproductive effects induced by VED were highly similar to one aspect of those observed in aged mice. Our findings demonstrate that the aging of the male reproductive system may be accelerated because of the impaired in vivo antioxidant capacity induced by VED., (Copyright © 2020 Elsevier Inc. All rights reserved.)

Sperm genetic testing has been proposed for clinical diagnosis of possible causes of male infertility. We reviewed the most remarkable publications of sperm DNA integrity and sperm aneuploidy as they relate to clinical outcomes, and the relationship between both genetic defects, and its association to embryo aneuploidy and recurrent pregnancy loss., (Copyright © 2020. Published by Elsevier Ltd.)

Routine semen analysis is a poor predictor of pregnancy rates after intra-cytoplasmic sperm injection (ICSI). There is an assumption that embryos' quality and fertilization rates could be influenced by epigenetic factors. We aimed at comparing global sperm DNA methylation level (GSDML) between normal and abnormal semen, investigating its relationship with sperm parameters and assessing its effect on ICSI outcomes; fertilization, good embryo and pregnancy rates. Ejaculates were obtained from 104 infertile and 60 fertile males undergoing ICSI at Faculty of Medicine, Alexandria, Egypt. Data was analyzed using IBM SPSS software package, 20. Inter-group differences in semen parameters were assessed by t-test. GSDML, measured by ELISA, showed significant positive correlation with sperm count, concentration and motility. It correlated positively but insignificantly with morphology and fertilization rate. High levels were significantly associated with embryos having good quality and positive pregnancy rates. GSDML could predict good embryo rate and pregnancy occurrence after ICSI.

Spermatozoa are polarized cells with a head and a flagellum joined by the connecting piece. Head integrity is critical for normal sperm function, and head defects consistently lead to male infertility. Abnormalities of the sperm head are among the most severe and characteristic sperm defects. Patients presenting with a monomorphic head sperm defects such as globozoospermia or marcrozoospermia were analyzed permitting to identify several key genes for spermatogenesis such as AURKC and DPY19L2. The study of patients with other specific sperm head defects such as acephalic spermatozoa have also enabled the identification of new infertility genes such as SUN5. Here, we review the genetic causes leading to morphological defects of sperm head. Advances in the genetics of male infertility are necessary to improve the management of infertility and will pave the road towards future strategies of treatments, especially for patients with the most severe phenotype as sperm head defects., (Copyright © 2020 Elsevier Ltd. All rights reserved.)