Role In Male Reproduction Research Articles

EP300‐interacting inhibitor of differentiation 3 is required for spermatogenesis in mice

AbstractBackgroundMammalian spermatogenesis is a highly complex process of cell proliferation, meiosis, and differentiation. A series of genes are expressed in an orderly and precise manner to ensure spermatogenesis, with chromatin undergoing intricate changes throughout. EP300‐interacting inhibitor of differentiation 3 (Eid3) is a testis‐enriched gene, but its role in male reproduction remains unclear.ObjectiveTo investigate the role of EID3 in male spermatogenesis and explore the potential underlying mechanism.Materials and MethodsWe generated Eid3 knockout mouse model using the CRISPR‐Cas9 system. We measured the expression of EID3 in mouse tissues and testicular cell populations by qRT‐PCR and western blot. Histological analysis, including hematoxylin and eosin (H&E) and periodic acid‐Schiff (PAS) staining, together with computer‐assisted sperm analysis (CASA), were performed to evaluate the effect of EID3 on spermatogenesis in mice. Light and ultrastructural microscopy were used to evaluate the morphology and structure of the Eid3−/− spermatozoa. We used western blot and immunofluorescence to further analyze the function of EID3 in spermiogenesis.ResultsEid3−/− mouse showed a significant decrease in sperm count, motility, and morphology. Loss of EID3 impaired the normal meiotic process and induced apoptosis of abnormally developing spermatocytes, ultimately resulting in the decrease of sperm cell number. Additionally, EID3 deficiency led to a decrease in histone acetylation levels in spermatids, impaired histone‐to‐protamine transition and chromatin condensation process, and ultimately resulted in abnormal sperm morphology.Discussion and ConclusionsThis study confirms for the first time that EID3 is crucial for meiosis and chromatin condensation during spermatogenesis, and EID3 deficiency leads to a significant decrease in sperm parameters. Given the high expression paradigm of Eid3 in human testis, EID3 likely plays a role in human reproduction. Future research could provide a new target for the clinical diagnosis and treatment of male infertility.

Roles of thyroid and leptin hormones and their crosstalk in male reproductive functions: an updated review.

This review aims to provide updated information regarding the role of thyroid and leptin hormones and their crosstalk in affecting the male reproductive function in hypothyroid and obesity conditions. A wide literature search was made using online search engines on published articles using keywords including thyroid hormone, hypothyroidism, leptin hormone, hyperleptinemia, obesity, the relationship between thyroid and leptin hormones and male reproduction, and hypothyroidism, obesity, and male reproduction. All information pertaining thyroid and leptin hormone effects on male reproduction, hypothyroidism, hyperleptinemia, and obesity effect on male fertility as well as the related molecular mechanisms are obtained. Thyroid and leptin hormones individually play a significant role in male reproduction. Alterations of these hormones' levels could adversely affect the male reproductive functions. PI3K/AKT signaling was found to be the major signaling pathway involved in mediating the effect of both hormones on male reproduction. Impaired crosstalk between the two hormones may occur in hypothyroidism with obesity which would contribute towards male reproductive dysfunction.

Novel Genes of the Male Reproductive System: Potential Roles in Male Reproduction and as Non-hormonal Male Contraceptive Targets.

The development of novel non-hormonal male contraceptives represents a pivotal frontier in reproductive health, driven by the need for safe, effective, and reversible contraceptive methods. This comprehensive review explores the genetic underpinnings of male fertility, emphasizing the crucial roles of specific genes and structural variants (SVs) identified through advanced sequencing technologies such as long-read sequencing (LRS). LRS has revolutionized the detection of structural variants and complex genomic regions, offering unprecedented precision and resolution over traditional next-generation sequencing (NGS). Key genetic targets, including those implicated in spermatogenesis and sperm motility, are highlighted, showcasing their potential as non-hormonal contraceptive targets. The review delves into the systematic identification and validation of male reproductive tract-specific genes, utilizing advanced transcriptomics and genomics studies with validation using novel knockout mouse models. We discuss the innovative application of small molecule inhibitors, developed through platforms like DNA-encoded chemistry technology (DEC-Tec), which have shown significant promise in preclinical models. Notable examples include inhibitors targeting serine/threonine kinase 33 (STK33), soluble adenylyl cyclase (sAC), cyclin-dependent kinase 2 (CDK2), and bromodomain testis associated (BRDT), each demonstrating nanomolar affinity and potential for reversible and specific inhibition of male fertility. This review also honors the contributions of Dr. David L. Garbers whose foundational work has paved the way for these advancements. The integration of genomic, proteomic, and chemical biology approaches, supported by interdisciplinary collaboration, is poised to transform male contraceptive development. Future perspectives emphasize the need for continued innovation and rigorous testing to bring these novel contraceptives from the laboratory to clinical application, promising a new era of male reproductive health management.

The interactome of cystic fibrosis transmembrane conductance regulator and its role in male fertility: A critical review.

The cystic fibrosis transmembrane conductance regulator (CFTR) is a cyclic adenosine monophosphate (cAMP)-regulated chloride and bicarbonate ion channel found in many human cells. Its unique biochemical characteristics and role as a member of the adenosine triphosphate (ATP)-binding cassette transporters superfamily are pivotal for the transport of several substrates across cellular membranes. CFTR is known to interact, physically and functionally, with several other cellular proteins. Hence, its properties are essential for moving various substances across cell membranes and ensuring correct cell functioning. Genetic mutations or environmental factors may disrupt CFTR's function resulting in different possible phenotypes due to gene variations that affect not only CFTR's function, localization, and processing within cells, but also those of its interactors. This has been reported as an underlying cause of various diseases, including cystic fibrosis. The severe clinical implications of cystic fibrosis have driven intense research into the role of CFTR in lung function but its significance to fertility, particularly in men, has been comparatively understudied. However, ongoing and more recent research into CFTR and its interacting proteins in the testis or specific testicular cells is beginning to shed light on this field. Herein, we provide a comprehensive and up-to-date overview of the CFTR, its interactome, and its crucial role in male reproduction, highlighting recent discoveries and advancements in understanding the molecular mechanisms involved. The comprehension of these complex interactions may pave the way for potential therapeutic approaches to improve fertility of men suffering from alterations in the function of CFTR.

Eighteen genes primarily expressed in the testis are not required for male fertility in mice†.

There are approximately 20000 protein-coding genes in humans and mice. More than 1000 of these genes are predominantly expressed in the testis or are testis-specific and thought to play an important role in male reproduction. Through the production of gene knockout mouse models and phenotypic evaluations, many genes essential for spermatogenesis, sperm maturation, and fertilization have been discovered, greatly contributing to the elucidation of their molecular mechanisms. On the other hand, there are many cases in which single-gene knockout models do not affect fertility, indicating that tissue-specific genes are not always critical. Here, we selected 18 genes whose mRNA expression is restricted to the testis or higher than in other tissues, but whose function in male reproduction is unknown. We then created single-gene KO mouse models using the CRISPR/Cas9 system. The established KO males were subjected to mating tests and screened for effects on fecundity, revealing that these genes were not essential for spermatogenesis and male fertility. This knowledge will contribute to understanding the functions of genes characteristic of the testis and identify the cause of male infertility.

TPCA-1 compound, inhibiting testis-specific serine/threonine protein kinase 3 for potential male sterile in Bombyx mori.

Protein kinases are a type of transferase enzyme that catalyze the phosphorylation of protein substrates, including receptor proteins. Testis-specific serine/threonine kinases (TSSKs) are a highly conserved group of protein kinases found in various organisms. They play an essential role in male reproduction by influencing sperm development and function. In this study, we report on the characterization of BmTSSK3, a TSSK from the silkworm, Bombyx mori. We found that BmTSSK3 is specifically expressed in the testis and localized to the sperm flagella, particularly in the sperm tail cyst. Furthermore, we developed BmTSSK3 inhibitors through molecular docking and binding assays. Small molecules 5-(4-Fluorophenyl)-2-ureidothiophene-3-carboxamide (TPCA-1)and Imidurea were identified to bind to BmTSSK3. Using site-specific mutation technology, we identified amino acid residues R134 and S184 as crucial binding sites for small molecules. RNA interference assay and Western blot analysis showed that knockdown of BmTSSK3 significantly decreased histone 3 phosphorylation. To confirm the inhibitory effect of these small molecules, we treated silkworm testes with TPCA-1 and observed a strong inhibitory effect. TPCA-1 is an inhibitor of BmTSSK3, which raises its potential as a future candidate for male sterility of the silkworm. Thus, this study may offer a novel strategy for sterile silkworms as well as insects. © 2024 Society of Chemical Industry.

P-094 Genetic variations of antioxidant genes may coordinate with lifestyle to define male fertility: A cross-sectional study with control group

Abstract Study question Are polymorphisms of the oxidative stress-related genes different between infertile men and reproductively healthy men? Does lifestyle contribute to fertility outcomes? Summary answer A strong association between the polymorphisms of two antioxidant genes and sperm quality was identified. The infertile men had higher BMI and alcohol usage. What is known already Antioxidant genes including catalase (CAT), superoxide dismutase (SOD), and nitric oxide synthase (NOS) play a critical role in spermatogenesis and sperm functions, especially in stress-response cell signaling pathways. Polymorphisms of antioxidant genes have been reported to be associated with gene function, hence sperm quality, ultimately affecting male fertility in various cohorts. Body mass index (BMI) is universally considered a marker of health reflecting whole health, nutrition, and lifestyle. Smoking and alcohol consumption expose the body to harmful chemicals and oxidative stress. Here, we examined the polymorphisms of antioxidant genes in Vietnamese men and the link with lifestyle to regulate fertility. Study design, size, duration This cross-sectional study with control group recruited from Hanoi Medical University Hospital from 2018-2020. The participants were divided into two groups regarding semen analysis based on WHO 2010 criteria. The participants gave consent, then were clinically examined, interviewed for lifestyle, and genetically analyzed for antioxidant gene variations. An extra inclusion criterion for the control men was having at least one healthy baby. The exclusion criteria were chromosome aberrations and Y chromosome microdeletion. Participants/materials, setting, methods The control group (N = 85) and infertile group (N = 107) were recruited. Semen analysis was carried out by the CASA instrument based on WHO 2010 criteria. Lifestyle characteristics including exercise (per week), smoking (yes/no), alcohol consumption (the threshold for “yes” was five standard drinks/occasion, at least one occasion/month) were recorded. Four variants SOD1:7958G>A, SOD2:c.47T>C, CAT:-262C>T, and NOS3:-786C>T were analyzed by Sanger sequencing. Pearson’s, Chi-squared (χ2), Wilcoxon rank-sum, ANOVA test, and Bonferroni correction were applied where specifically required. Main results and the role of chance The average age of the two groups was not different (31.96 ± 4.87 vs 31.93 ± 6.3 years old, p = 0.92). The body mass index (BMI) of infertile men was significantly higher than the control (24.84 ± 2.31 vs 23.53 ± 2.55 kg/m2, p = 0.00026). No significant difference was identified regarding exercise and smoking. Notably, 96.26% of infertile men consumed alcohol in comparison with 88.23% of the control (p = 0,034). Genetically, SOD1:7958GA genotype and NOS3:-786CT genotype in the infertility group were significantly higher than in the control, OR = 2.191 (95% CI: 1.226–3.915, p = 0.004) and OR = 3.135 (95% CI: 1.591–6.180, p < 0.001), respectively. The frequency of the SOD2:c.47TC genotype was significantly higher in the male infertility group than in the control (OR = 1.941, 95% CI: 1.063–3.595, p = 0.029). Individuals carrying the SOD1:GA+AA, SOD2:TC+CC, and CAT:CT/TT genotype pattern had an increased risk of male infertility (OR = 7.614, p = 0.007), suggesting that gene-gene interaction plays an important role in male reproduction. To our knowledge, this is the first report worldwide on the association between the SOD1:7958G>A polymorphism and male infertility. Together, the data suggest that antioxidant gene polymorphisms and lifestyle, especially alcohol usage, coordinate to regulate male fertility. Limitations, reasons for caution Direct measurement of oxidative stress in the semen of men having different gene variations would be required to ascertain the link between the polymorphisms of antioxidant genes and fertility. The sample size was relatively small, a larger sample size would be required to accurately represent the Vietnamese population. Wider implications of the findings The findings can assist in infertility diagnosis focusing on the group of higher infertile risk genetic variations. Interventions such as antioxidant supplements would be developed and applied, lifestyle advice would be given specifically for a group having high-risk gene variations to accurately target fertility issues. Trial registration number Not applicable

VD3/VDR attenuates bisphenol A-induced impairment in mouse Leydig cells via regulation of autophagy.

Bisphenol A (BPA) is an endocrine disruptor with reproductive toxicity. Further, 1,25-dihydroxyvitamin D3 (VD3) plays an important role in male reproduction by binding vitamin D receptor (VDR) and mediating the pleiotropic biological actions that include spermatogenesis. However, whether VD3/VDR regulates the effect of BPA on Leydig cells (LCs) injury remains unknown. This study aimed to explore the effects of VD on BPA-induced cytotoxicity in mouse LCs. Hereby, LCs treated with BPA, VD3, or both were subjected to the assays of cell apoptosis, proliferation, autophagy, and levels of target proteins. This study unveiled that cell viability was dose-dependently reduced after exposure to BPA. BPA treatment significantly inhibited LC proliferation, induced apoptosis, and also downregulated VDR expression. By jointly analyzing transcriptome data and Comparative Toxicogenomics Database (CTD) data, autophagy signaling pathways related to testicular development and male reproduction were screened out. Therefore, the autophagy phenomenon of cells was further detected. The results showed that BPA treatment could activate cell autophagy, Vdr-/- inhibits cell autophagy, and active VD3 does not have a significant effect on the autophagy of normal LCs. After VD3 and BPA were used in combination, the autophagy of cells was further enhanced, and VD3 could alleviate BPA-induced damage of LCs. In conclusion, this study found that supplementing VD3 could eliminate the inhibition of BPA on VDR expression, further enhance LCs autophagy effect, and alleviate the inhibition of LCs proliferation and induction of apoptosis by BPA, playing a protective role in cells. The research results will provide valuable strategies to alleviate BPA-induced reproductive toxicity.

Bacterial Zoonotic Diseases and Male Reproduction

Bacterial zoonotic diseases hold significant public health importance due to their substantial contribution to human morbidity and mortality. These infections have been implicated in reducing the fertility rate among couples of reproductive age. Despite the increasing prevalence of infertility and sub-fertility in men, there has been limited investigation into the possible effects of bacterial zoonotic infections on the male reproductive system. The purpose of this review is to describe common bacterial zoonotic diseases and their effects on human reproduction in order to unveil the hidden roles these infections could play in male factor infertility. While there is a dearth of information on this subject from human studies, available evidence from experimental animals suggests that bacterial zoonotic diseases impair male reproductive functions and structures primarily through the activation of the inflammatory response and distortion of the antioxidant system, resulting in the generation of oxidative species. In light of the limited research on bacterial zoonotic diseases and their role in male reproduction, efforts must be directed towards the subject to unravel the underlying pathological mechanisms and reduce the incidence among the human populace, either through preventive or curative measures.

Transcriptome Analysis Reveals Spermatogenesis-Related CircRNAs and LncRNAs in Goat Spermatozoa.

Mammalian spermatozoa comprises both coding and non-coding RNAs, which are traditionally believed to be a residual of spermatogenesis. The differential expression level of spermatozoal RNAs is also observed between fertile and infertile human, thereby anticipated as potential molecular marker of male fertility. This study investigated the transcriptome profile of goat (Capra hircus) spermatozoa. The sperm transcriptome was analyzed by three different methods viz. RLM-RACE, long-read RNA sequencing (RNAseq) in Nanopore™ platform, and short-read RNAseq in Illumina™ platform. The Illumina™ sequencing discovered 16,604 transcripts with 357 mRNAs having FPKM (fragments per kilobase per million mapped reads) of more than five. The spermatozoal RNA suite included mRNA (94%), rRNA (3%), miscRNA (1%), circRNA (1%), miRNA (1%), etc. This study also predicted circRNAs (127), lncRNAs (655), and imprinted genes (160) that have potential role in male reproduction. The gene ontology analysis revealed the involvement of spermatozoal RNA in regulating male meiosis (TET3, STAT5B), capacitation (ACRBP, CATSPER4), sperm motility (GAS8, TEKT2), spermatogenesis (ADAMTS2, CREB3L4), etc. The spermatozoal RNA were also associated with different biological pathways viz. Wnt signaling pathway, cAMP signaling pathway, AMPK signaling pathway, and MAPK signaling pathways having potential role in spermatogenesis. Overall, this study enlightened the suite of spRNA transcripts in goat and their relevance in male fertility for diagnostic approach.

Leptin Receptor Gln223Arg Polymorphism of Human Spermatozoa Associated with Male Infertility in a Chinese Population.

Leptin (LEP) is believed to play a crucial role in male reproduction, while the molecular mechanisms through which LEP affects the male reproductive system are unclear. LEP acts by binding to a leptin receptor (LEPR) which mediates its physiological action, but there are only limited studies on the function of LEPR in human sperm. This study aimed to determine the Gln223Arg polymorphisms of the LEPR gene in human spermatozoa and evaluate their possible relationship with semen variables. The study was performed on Chinese men: 115 healthy subjects and 108 patients with primary and 98 with secondary infertility. Semen samples were obtained from all patients, and semen variables were analyzed. The genotypic and allelic frequencies of Gln223Arg polymorphism in spermatozoa were determined by PCR and restriction fragment length polymorphism (RFLP) analyses. Statistical analyses were performed using the chi-square test, the Kruskal-Wallis test, and the Mann-Whitney test. There were no significant differences in genotypic or allelic frequency distributions of Gln223Arg polymorphism among men with primary infertility, secondary infertility, and controls. Similarly, semen volume and sperm concentration did not differ with the different genotypes in all groups of men. The percentages of motile sperm for AA + AG genotypes in men with primary infertility (31.98%) were significantly lower than those in secondary infertility, and control men with GG genotypes were 34.41% and 59.36%, respectively. At the same time, the percentages of normal morphology sperm for AA + AG genotypes in men with primary infertility (2.93%) were significantly lower than those in secondary infertility and control men with GG genotypes 3.71% and 6.54%, respectively. This study reveals a possible association between the Gln223Arg polymorphism of the LEPR gene in spermatozoa affecting spermatozoal membrane integrity and having a direct role in sperm motility.

WNK1 is required during male pachynema to sustain fertility

WNK1 is an important regulator in many physiological functions, yet its role in male reproduction is unexplored. In the male germline, WNK1 is upregulated in preleptotene spermatocytes indicating possible function(s) in spermatogenic meiosis. Indeed, deletion of Wnk1 in mid-pachytene spermatocytes using the Wnt7a-Cre mouse led to male sterility which resembled non-obstructive azoospermia in humans, where germ cells failed to complete spermatogenesis and produced no sperm. Mechanistically, we found elevated MTOR expression and signaling in the Wnk1-depleted spermatocytes. As MTOR is a central mediator of translation, we speculated that translation may be accelerated in these spermatocytes. Supporting this, we found the acrosome protein, ACRBP to be prematurely expressed in the spermatocytes with Wnk1 deletion. Our study uncovered an MTOR-regulating factor in the male germline with potential implications in translation, and future studies will aim to understand how WNK1 regulates MTOR activity and impact translation on a broader spectrum.

P-106 The inhibition of sperm SIRT1 enhances fertilization and modifies subsequent embryonic epigenetic quality

Abstract Study question Is SIRT1 present in sperm and thus involved in fertilization and/or sperm epigenetic code? Do the SIRT1-driven epigenetic marks be transmitted to the progeny? Summary answer SIRT1 localized at the sperm head and influences fertilization and sperm epigenetic code via its deacetylation activity. SIRT1-modulated epigenome is transmitted to zygotes and blastocysts. What is known already The addition or removal of acetyl group from lysine residua (i.e. acetylation) post-translationally modulates proteins through the action of acetylases and deacetylases, respectively. SIRT1 (also Sirtuin 1) is capable to deacetylate a number of proteins, including histones, with roles in male reproduction. Testicular SIRT1 deficit leads to subfertility and increases number of abnormal and immotile spermatozoa that fail to fertilize. However, neither sperm SIRT1 presence nor its function has been investigated despite the significance of acetylation in the events around fertilization. Moreover, the epigenetic mode of action of sperm SIRT1 highlights its relevance for the success of the pre-implantation embryo. Study design, size, duration Mouse sperm of 14-18-weeks-old males of CD-1 x C57Bl6 hybrid strain were isolated from the epididymis. Sperm were incubated under capacitating conditions in presence of sirtinol (5 µM), a selective SIRT1 inhibitor, or the vehicle (0.1% v/v DMSO) for 1 h in 37 °C and 5% CO2. Participants/materials, setting, methods Motility and acrosome reaction was evaluated as previously stated (Ritagliati et al., 2018). Moreover, sperm proteins were extracted into RIPA-buffer and acetylated proteins were depicted by western blot. Finally, in vitro fertilization and embryo culture were performed. Above parameters defining IVF outcomes (i.e., fertilization, cleavage, and blastocyst rate), immunocytochemistry of acetylated histone 4 in the 16th residua of lysine (H4K16) in zygotes and blastocyst was quantified. Main results and the role of chance SIRT1 was localized in the sperm head, apparently at the perinuclear theca and perforatorium. Inhibition of SIRT1 leads to the increase of lysine acetylation (2.28 vs 1, fold change values) and hyperactivation (16.55 ± 0.79 vs 10.84 ± 0.79 %sperm), the enhanced motility that facilitates sperm to penetrate the oocyte, as well as the acrosome reaction (28.69 ± 0.41 vs 17.98 ± 0.41 %sperm). Subsequently, these spermatozoa were able to better fertilize oocytes in vitro (90.5 ± 7.21 vs 66.25 ± 7.21, %). Interestingly, zygotes derived from sirtinol-pre-treated spermatozoa developed to 2-cell embryos at a higher rate than those fertilized by untreated spermatozoa (79.33 ± 7.21 vs 67.00 ± 7.21, %). Moreover, we observed significant hyperacetylation of H4K16 in the male pronuclei of zygotes and embryos derived from sirtinol-treated sperm (8.19 ± 1.02 vs 6.10 ± 2.09 and 45.55 ± 13.96 vs 27.62 ± 13.96, integrated density of the signals, respectively). Limitations, reasons for caution Although acetylation of H4K16 was accumulated in male pronuclei, we do not know which subset of genes was affected. Therefore, we only can assure that SIRT1-driven epigenetic marks are transmitted to the offspring, but their consequences should be further elucidated. For knowledge transfer to humans, a verification study is required. Wider implications of the findings the reduction in testicular SIRT1 expression is an age-related phenomenon. Therefore, our results support SIRT1 as the basis for new interventions against age-related male infertility. Moreover, due to the high reactivity of SIRT1 to its inactivation, SIRT1 becomes a target to pharmacologically enhance sperm fertilization ability in assisted reproductive techniques. Trial registration number Animal procedures were conducted in accordance with Act No. 246/1992 Coll. on the Protection of Animals Against Cruelty under the supervision of the Animal Welfare Committee of the Ministry of Education, Youth and Sports of the Czech Republic, approval ID MSMT-11925/2016-3.

Expression profiles of meiotic genes in male vs. female gonads and gametes: Insights into fertility issues.

Gametes are specialized cells that, at fertilization, give rise to a totipotent zygote capable of generating an entire organism. Female and male germ cells undergo meiosis to produce mature gametes; however, sex-specific events of oogenesis and spermatogenesis contribute to specific roles of gametes in reproductive issues. We investigate the differential gene expression (DGE) of meiosis-related genes in human female and male gonads and gametes in normal and pathological conditions. The transcriptome data for the DGE analysis was obtained through the Gene Expression Omnibus repository, comprising human ovary and testicle samples of the prenatal period and adulthood, additionally to male (non-obstructive azoospermia (NOA) and teratozoospermia), and female (polycystic ovary syndrome (PCOS) and advanced maternal age) reproductive conditions. Gene ontology terms related to meiosis were associated with 678 genes, of which 17 genes in common were differentially expressed between the testicle and ovary during the prenatal period and adulthood. Except for SERPINA5 and SOX9, the 17 meiosis-related genes were downregulated in the testicle during the prenatal period and upregulated in adulthood compared to the ovary. No differences were observed in the oocytes of PCOS patients; however, meiosis-related genes were differentially expressed according to the patient's age and maturity of the oocyte. In NOA and teratozoospermia, 145 meiosis-related genes were differentially expressed in comparison to the control, including OOEP; despite no recognized role in male reproduction, OOEP was co-expressed with genes related to male fertility. Taking together, these results shed light on potential genes that might be relevant to comprehend human fertility disorders.

Identification of biallelic variations of CEP70 in patients with male infertility.

Male infertility is a severe health issue caused by complex and multifactorial pathological conditions. Genetic factors are a major cause of male infertility. CEP70, a centrosomal protein, has been reported to play an important role in male reproduction in mice. However, the role of CEP70 in human male infertility is limited. Whole exome sequencing and Sanger sequencing were used to identify the genetic cause of the infertile patients. Papanicolaou staining, scanning electron microscopy and transmission electron microscopy were further conducted to explore morphological and ultrastructural defects in spermatozoa from the patient. Immunofluorescence staining was used to detect the pathogenicity of the identified variants and the particular expression of CEP70 in testis. In this study, we identified biallelic mutations of CEP70 in two unrelated infertile male individuals with oligoasthenoteratozoospermia that followed a recessive inheritance pattern. Papanicolaou staining, scanning electron microscopy and transmission electron microscopy showed that morphological and ultrastructural defects in the acrosome and flagellum of sperm from the patient in a pattern strikingly similar to that in Cep70-/- male mice. The results of immunofluorescence staining suggested that CEP70 was normally expressed in the acrosome and flagellum of control sperm but was hardly detected in the sperm of patient carrying CEP70 variation. We also explored the particular expression pattern of CEP70 during spermatogenesis in humans and mice. Biallelic mutations of CEP70 might be a novel genetic cause of human male infertility, which could potentially serve as a basis for genetic counseling and diagnosis of male infertility.

Impact of endocrine disrupting chemicals and pharmaceuticals on Sertoli cell development and functions.

Sertoli cells play essential roles in male reproduction, from supporting fetal testis development to nurturing male germ cells from fetal life to adulthood. Dysregulating Sertoli cell functions can have lifelong adverse effects by jeopardizing early processes such as testis organogenesis, and long-lasting processes such as spermatogenesis. Exposure to endocrine disrupting chemicals (EDCs) is recognized as contributing to the rising incidence of male reproductive disorders and decreasing sperm counts and quality in humans. Some drugs also act as endocrine disruptors by exerting off-target effects on endocrine tissues. However, the mechanisms of toxicity of these compounds on male reproduction at doses compatible with human exposure are still not fully resolved, especially in the case of mixtures, which remain understudied. This review presents first an overview of the mechanisms regulating Sertoli cell development, maintenance, and functions, and then surveys what is known on the impact of EDCs and drugs on immature Sertoli cells, including individual compounds and mixtures, and pinpointing at knowledge gaps. Performing more studies on the impact of mixtures of EDCs and drugs at all ages is crucial to fully understand the adverse outcomes these chemicals may induce on the reproductive system.

Effect of Organic Zinc Supplementation on Semen Production and Quality in Kankrej Bulls

Background: Zinc is found in very high concentration in semen and male reproductive tract and acts as a hormone receptor modulator for testosterone production in Leydig cells. Thus, it plays a very important role in male reproduction. Moreover, Indian soils are very deficient in zinc as per earlier reports and recent studies have shown that supplementation of zinc has improved qualitative as well as the quantitative aspects of semen quality in bulls. So, a study has been planned to study the effect of organic zinc supplementation on semen production and quality in kankrej bulls. Methods: A total of 8 healthy Kankrej bulls under semen collection were selected for the study. Basal diet (concentrate mixture, green and dry fodder) was fed during pre-supplementation and zinc propionate @ 100 ppm/bull was supplemented to the basal diet during post-supplementation for a period of 60 days to complete one spermatogenic cycle. A total of 128 ejaculates (64 pre-supplementation and 64 post-supplementation) were collected. Result: Mean semen volume was non-significant (P greater than 0.05) between the treatment groups. Mean sperm concentration was 1325.68±80.08 and 2162.36±112.0 million/ml pre- and post-supplementation, respectively. Feeding of zinc propionate significantly (P less than 0.01) increased semen sperm concentration. The average mass motility of sperms was significantly (P less than 0.01) higher during post-supplementation (3.92%) than the pre-supplementation (3.78%). The overall live sperms were significantly (P less than 0.01) higher during post-supplementation (73.33±0.91%) than the pre-supplementation (63.53±1.03%). It may be concluded that supplementation of 100 ppm of zinc propionate in the diet of Kankrej bulls improved semen quality in terms of quantitative and qualitative characteristics of semen.

Identification of some calcium binding proteins and neural cell markers in rat testis and epididymis during postnatal development.

Calcium-binding proteins (CaBPs) have an essential role in male reproduction by modulating calcium ion metabolism. Although the brain and testis are structurally and functionally different, they show a high degree of transcriptomic and proteomic similarities. The purpose of the present study was to explore some CaBPs (Iba-1, Calbindin, Calretinin and Parvalbumin) and neural cell markers (CNPase, NG2 and Drebrin) expression in rat testis and epididymis during postnatal periods by using immunohistochemistry. Iba-1, calbindin, calretinin, parvalbumin, CNPase, NG2 and drebrin were expressed in the epididymal epithelium in each postnatal period. Iba-1 and calbindin expression showed stage-dependent expression in spermatids. CaBPs and neural cell markers showed a positive reaction in Leydig cells in the postpubertal and mature periods. Sertoli cells, gonocytes, spermatogonium, and peritubular myoid cells showed heterogeneity in the expression of CaBPs and nerve markers throughout postnatal development. Interestingly, CNPase, NG2 and drebrin were positive in spermatocytes, spermatids, and sperm. Expression dynamics of calcium-binding proteins and nerve markers showed differences in germ cells and somatic cells during postnatal development. The expression of these proteins in the testis and epididymis supports that they may have important roles in reproductive physiology.

Identification and Characterization of Piwi-Interacting RNAs for Early Testicular Development in Yak

Normal testicular development plays a crucial role in male reproduction and is the precondition for spermatogenesis. PIWI-interacting RNAs (piRNAs) are novel noncoding RNAs expressed in animal germ cells that form complexes with PIWI family proteins and are involved in germ cell development, differentiation, and spermatogenesis. However, changes in piRNA expression profiles during early testicular development in yak have not been investigated. In this study, we used small RNA sequencing to evaluate the differences and potential functions of piRNA expression profiles in 6-, 18-, and 30-month-old yak testis tissues. Differential expression analysis found 109, 293, and 336 differentially expressed piRNAs in M30 vs. M18, M18 vs. M6, and M30 vs. M6, respectively, and found 30 common differentially expressed piRNAs in the three groups of M6, M18, and M30. In addition, the functional enrichment analysis of differentially expressed piRNAs target genes indicated that they were related to testicular development and spermatogenesis. Finally, we detected the expression of the PIWI protein family in the yak testis at different developmental stages and found that PIWIL1, PIWIL2, PIWIL3, and PIWIL4 were highly expressed in 18- and 30-month-old yak testis and almost not expressed in 6-month-old yak testis. In conclusion, this study summarizes the changes of piRNA expression patterns during the early development of yak testis and provides new clues for the regulatory role of piRNA in yak testis.

Vitamin D and Male Reproduction: Updated Evidence Based on Literature Review.

The purpose of this study was to clarify the vitamin D (VD) effect on male infertility. Our research was conducted using the following electronic databases: MEDLINE, Embase, Web of Science, Scopus, ClinicalTrials.gov, and Cochrane Library. Selection criteria included all published randomized controlled trials and non-randomized studies, focusing on vitamin D and male reproductive function. The results showed that the effects of VD on male reproduction has been investigated in three different topics: the molecular mechanism underlying VD effects on semen quality (SQ), the relationship between VD levels and SQ, and the effect of VD supplementation on SQ. Results supported the hypothesis of a relevant interrelation between concentrations of male VD and semen parameters, with particular reference to sperm motility; on the contrary, evidence on the effect of VD on male sex steroid hormone levels was inconclusive. The results of this review hold up the thesis that VD plays a role in male reproduction. Most of the data highlighted a positive effect on semen quality, particularly in sperm motility, both in fertile and infertile men. Additional dedicated studies are required to elucidate the still controversial aspects of this topic.