Search Results (108)

Artificial gynogenesis is an essential technique for aquaculture breeding. Fertile offspring of the WuLi carp (Cyprinus carpio var. Quanzhounensis, 2n = 100, WLC) were successfully produced via gynogenesis using ultraviolet-irradiated sperm from the blunt snout bream (Megalobrama amblycephala, 2n = 48, BSB). As anticipated, gonadal section examination confirmed that all gynogenetic WuLi carp (2n = 100, GWB) were female. To investigate whether paternal DNA fragments from BSB were integrated into the GWB genome, comparative analyses of morphological traits, DNA content, chromosomal numbers, 5S rDNA sequences, microsatellite DNA markers, fluorescence in situ hybridization (FISH), growth performance and nutritional composition were systematically conducted between GWB and maternal WLC. The results revealed pronounced maternal inheritance patterns across morphological characteristics, DNA quantification, chromosomal configurations, 5S rDNA sequences and FISH signals, while microsatellite detection unequivocally confirmed paternal BSB DNA fragment integration into the GWB genome. Remarkably, GWB demonstrated significantly superior growth performance and elevated unsaturated fatty acid content relative to the maternal line. This approach not only addressed germplasm degradation in WLC but also provided valuable theoretical foundations for breeding programs in this commercially significant species. Full article

Background/Objectives: Preimplantation genetic screening improves embryo selection in intracytoplasmic sperm injection cycles, especially for women of advanced maternal age. As chromosomal normality declines with age, high-dose gonadotropins are commonly used to enhance follicular response. This study compares minimal and high-dose stimulation protocols in terms of euploidy, pregnancy, and live birth rates following single embryo transfer. Methods: In this prospective study, 198 women aged 38–45 years were enrolled and divided into two groups: minimal stimulation (100 mg clomiphene citrate and 75 IU human menopausal gonadotropin) and high stimulation (300–450 IU gonadotropins). Women with severe male factor infertility, endometriosis, or absolute tubal factor were excluded. Clinical outcomes were compared using a t-test or Mann–Whitney U test. Results: Baseline characteristics were similar between groups. The high-dose group had a significantly higher number of retrieved oocytes (p = 0.009) and metaphase II oocytes (p = 0.003). However, there were no significant differences in euploid embryo rates (35.4% vs. 37.4%, p = 0.768), clinical pregnancy rates (67.6% vs. 69.4%, p > 0.999), gestational sac rates (58.8% vs. 58.3%, p > 0.999), or live birth rates (47.1% vs. 50.0%, p = 0.995). Conclusions: This is the first prospective study to compare euploid embryo rates, pregnancy rates, and live birth rates between minimal stimulation protocol and high stimulation protocol in AMA patients. Although there has been no difference in euploid and pregnancy rates, minimal stimulation protocol has advantages in cost and comfort. Full article

In vitro maturation (IVM) of oocytes retrieved from ovum pick-up (OPU) or ovarian tissue (OT) is a standard approach for patients with specific conditions where prior hormonal stimulation is contraindicated. However, the developmental competence of oocytes matured in vitro is still inferior to that of oocytes matured in vivo. Capacitation IVM (CAPA-IVM) includes an extra step of pre-maturation culture (PMC) with c-type natriuretic peptide (CNP) as a meiotic arrestor to better synchronize cytoplasmic and nuclear maturity in oocytes by allowing the cytoplasm additional time to acquire essential components critical for optimal competency. This study aims to evaluate the effect of CAPA-IVM on equine oocyte quality and developmental competence. Immature cumulus–oocyte complexes (COCs) were retrieved from slaughterhouse ovaries and matured in vitro either in CAPA-IVM (short 6 h, long 24 h pre-maturation) or standard IVM. Mature oocytes from each group were analyzed for calcium-releasing potential (n = 52) and single-oocyte proteomics (n = 44), and embryo development (n = 229) was assessed after fertilization with piezo-drilled intracytoplasmic sperm injection (ICSI). Genetic analysis of developed blastocysts (n = 41) was performed to detect chromosomal aberrations. Our findings demonstrate that CAPA-IVM of equine COCs yields significantly higher maturation rates than controls. Moreover, short CAPA-IVM with six hours pre-maturation culture showed substantially higher embryo development potential than the control group (20/69 vs. 9/63, respectively). Genetic analysis revealed a high euploidy rate in equine blastocysts regardless of the maturation conditions. Live calcium imaging of the fertilized oocytes demonstrated that the majority of oocytes displayed non-continuous calcium oscillation patterns, irrespective of maturation conditions. Single-oocyte proteomics reveals a comparable proteomic landscape between mature oocytes subjected to short CAPA-IVM and standard IVM. However, we identified four enriched gene sets with positive enrichment scores after short CAPA-IVM, related to cytoskeleton regulation, ribosomal function, and cytosolic components. Our findings indicate that CAPA-IVM holds the potential to improve oocyte quality and competence in horses. However, further fine-tuning of culture conditions would benefit the effective use of these IVM systems. Moreover, given that the mare serves as an excellent model for human reproduction, the molecular trends identified in this study could provide valuable insights for advancing human artificial reproductive technologies. Full article

Artificial gynogenesis is an effective means of producing pure lines and is widely used for genetic analysis of fish and for sex control. In this study, inactivated sperm from heterogenous blunt snout bream (Megalobrama amblycephala, MA) were used to activate Kohaku koi (Cyprinus carpio var. koi, CK) and produce high-quality female offspring. To determine whether the offspring were gynogenetic fish, the karyotype and DNA content of the CK, MA and the induced offspring (IO) were first compared and it was found that the IO were diploid with 100 chromosomes and their karyotype was 22m + 34sm + 22st + 22t. The DNA content of the IO was not significantly different from that of the CK. Subsequently, the amplified band profiles of CK, MA and IO were analyzed with species-specific microsatellite markers. The results showed that there were no amplified MA microsatellite bands in IO. The size of the amplified bands and the sequence of the 5S rDNA in CK, MA and IO were also analyzed. It was found that the amplified 5S rDNA gene fragments in IO contained two fragments that were both the same size as those of CK and matched more than 90% with those of CK. Finally, the sex of IO was verified using gonadal tissue sections. The result showed that IO was not an all-female population; males were also present (36.7%). In summary, a series of validation methods confirmed that the induced offspring were gynogenetic fish, which is the basis for the subsequent genetic improvement of pure lines of high-quality koi. Full article

DNA methylation is crucial in gene expression regulation and tissue differentiation in livestock. However, genome-wide methylation patterns among tissues remain underexplored in cattle, one of the world’s most important farm animals. This study investigates sex- and tissue-specific DNA methylation in cattle using CpG site methylation data generated by an Infinium DNA Methylation array (HorvathMammalMethyl-Chip40) across seven tissues. Our analysis revealed significant tissue-specific methylation differences, with reproductive tissues/cells, such as the sperm, exhibiting distinct profiles compared to somatic tissues like hair and blood. Principal component analysis (PCA) highlighted tissue differentiation as the primary driver of methylation variability. We also identified 222 CpG sites with significant sex-based methylation differences, particularly on the X chromosome, suggesting the potential epigenetic regulation of sex-specific traits. The Gene Ontology (GO) enrichment analysis indicated that these methylation patterns may influence biological processes such as epithelial cell proliferation and blood vessel remodeling. Overall, this study provides important insights into sex- and tissue-specific epigenetic regulation in cattle, with implications for improving livestock breeding strategies through integrating epigenetic data. Full article

Although yaks and cattle belong to the same Bovinae subfamily and have the same number of chromosomes, hybrid males are sterile because of the inactivation or abnormality of gene expression related to the production of healthy normal sperm. Recently, the analysis of gene expression not only in the testis but also in the epididymis has offered hints about the mechanism of infertility, because the epididymis supports the maturation of sperm in acquiring the capacity of fertilisation. Sperm maturation processes have been thought to be androgen-dependent, and the androgen receptor (AR) can be activated by dihydrotestosterone converted from plasma testosterone by the 5α-reductase isoform 2 (SRD5A2) in epididymal cells. In the present study, we investigated the immuno-expression levels of the AR and SRD5A2 in the epithelial cells of the hybrid cattle–yak epididymal caput in comparison with yak samples using image analysis. Epididymal tissues from yaks (1–3 years of age) and hybrid cattle–yaks (2 years of age) were used in this study. In yaks, AR signal intensity did not show any changes in epididymal epithelial cells during maturation. However, in 2-year-old hybrid cattle–yaks, AR signal intensity was significantly higher in the principal cells of the epididymis compared to that of yaks of the same age, indicating that hybrid sterility is not likely related to AR deficiency in the epididymal epithelium. On the other hand, SRD5A2 signal intensity was stable during maturation in the epithelial cells of the yak epididymis. However, the epididymal SRD5A2 signal intensity in the epithelial cells of the hybrid cattle–yak was lower than that of the yak. This suggests that a deficiency in SRD5A2 production in the epididymis may result in hybrid infertility, as it can subsequently cause incomplete AR signal transduction and altered spermatozoa physiology. Full article

Chinese tongue sole (Cynoglossus semilaevis) is an important mariculture fish in China, and female individuals present a growth advantage. However, genetic females (ZW) can sex reverse to phenotypic males, designated pseudomales. The pseudomale shows abnormal spermatogenesis and produces only Z sperm. Histone is pivotal in spermatogenesis, and post-translational modification could regulate its function. A comparison of testis phosphorylated and ubiquitinated proteins revealed 8 and 12 differentially phosphorylated and ubiquitinated histones in the testes of male and pseudomale Chinese tongue soles, respectively, but there was no difference in the translation level of these proteins. We selected four histone genes, h1.1-like, h1.2-like, h3, and h3.3-like, for further analysis. The expression levels of the h1.1-like, h3, and h3.3-like genes reached their highest levels at 2 years post-hatching (yph), and the expression level of h1.2-like reached its highest level at 1.5 years post-hatching (1.5 yph), indicating that its role began during the late stage of gonadal development. Promoter activity verification revealed that the promoters of the h1.1-like, h1.2-like, h3, and h3.3-like genes were located approximately upstream 2000 bp and six histone-related transcription factor sites were predicted. YY1A, YY1B, C-JUN, and JUNB may have negative regulatory effects on h1.1-like, h1.2-like, h3, and h3.3-like; AR and ETS-2 may have positive regulatory effects on h3 and h3.3-like. The ISH results revealed that h1.1-like, h1.2-like, h3, and h3.3-like mRNAs were located mainly in the sperm cells in the testes and the oocytes at various stages in the ovaries. After siRNA knockdown, the expression of dmrt1 in testis cell lines and the expression of tesk1 and neurl3 in males was downregulated, suggesting that the h1.1-like, h1.2-like, h3, and h3.3-like genes may have a negative regulatory role in spermatogenesis. The regulatory role in female fish remains to be explored. Mass spectrometry analysis revealed that histones have an important role in chromosome remodeling. These results provide a genetic basis for the molecular mechanism of gonadal development and spermatogenesis in Chinese tongue sole. Full article

Background: With the widespread use of static magnetic fields (SMFs) in applications such as magnetic resonance imaging (MRI) and electric vehicles, concerns have arisen regarding their potential effects on reproductive health. Despite increasing research, the impact of SMFs on reproductive function remains a subject of debate, requiring further exploration. Methods: This review synthesizes animal and clinical studies on the effects of SMF on reproductive function. It examines various SMF intensities and exposure durations, focusing on mitochondrial function, chromosomal division, and embryonic development. Results: The review reveals that low-intensity SMF exposure adversely affects mitochondrial function in sperm and eggs, reducing their activity. It also impacts follicular cells, delaying chromosomal division. Medium- and high-intensity SMF exposure shows mixed results, with both potential benefits and risks, requiring further research. High-intensity SMFs may pose teratogenic risks to embryos and delay the development of fertilized eggs. The position of SMF exposure also matters, likely due to field non-uniformity. Conclusions: This review provides a foundation for further investigation into the effects of SMFs on reproductive function, highlighting the need for more comprehensive studies to assess safety and applications. Special caution is advised for pregnant women regarding SMF exposure, given its potential risks. Full article

Current sperm sexing methods are costly and largely restricted to cattle, while immunological techniques targeting sex-specific membrane proteins may offer more economical alternatives. To advance these methods, understanding the proteomic differences between the cell membranes of X- and Y-chromosome-bearing spermatozoa is essential. This study aimed to characterize the cell surface proteome of bovine sperm and identify potential targets for sperm sexing through LC-MS/MS analysis. Cell surface protein lysates were extracted from unsexed, X-sperm (BX), and Y-sperm (BY) samples via biotinylation. Promising targets were identified through functional annotation (UniProt, eggNOG-mapper v.2.1.7) and topology prediction (DeepTMHMM v.1.0.13). Additionally, statistical overrepresentation (PANTHER 18.0) and orthology analyses were performed. Excluding contaminants, 130 proteins were detected, of which 64 proteins were detected in the BX samples and not in the BY samples. Of these, five transmembrane proteins stood out as potential X-sperm targets (ADAM2, ATP11C, DG1, MCT1, and PMCA4). They were identified as potential cell surface targets, based on GO terms and topology predictions, detected in at least two replicates of the BX samples, and shown to share orthology with other livestock species. These findings enhance our understanding of bovine sperm proteomics; however, further validation is required to confirm the utility of these five proteins in sperm sexing technologies. Full article

Telomeres protect chromosome ends from damage, but they shorten with each cell division due to the limitations of DNA replication and are further affected by oxidative stress. This shortening is a key feature of aging, and telomerase, an enzyme that extends telomeres, helps mitigate this process. Aging is also associated with mitochondrial dysfunction, leading to increased reactive oxygen species (ROS) that exacerbate cellular damage and promote apoptosis. Elevated ROS levels can damage telomeres by oxidizing guanine and disrupting their regulation. Conversely, telomere damage impacts mitochondrial function, and activation of telomerase has been shown to reverse this decline. A critical link between telomere shortening and mitochondrial dysfunction is the DNA damage response, which activates the tumor suppressor protein p53, resulting in reduced mitochondrial biogenesis and metabolic disruptions. This highlights the bidirectional relationship between telomere maintenance and mitochondrial function. This review explores the complex interactions between telomeres and mitochondria across various cell types, from fibroblasts to sperm cells, shedding light on the interconnected mechanisms underlying aging and cellular function. Full article

Infertility is a significant global health issue, affecting 8–12% of couples of reproductive age, with male factor infertility contributing to 30–50% of cases. Despite advances in assisted reproductive technologies, particularly intra-cytoplasmic sperm injection, male infertility remains understudied compared to female infertility. This review aims to explore the genetic underpinnings of male factor infertility, including identified genetic mutations, chromosomal abnormalities, and epigenetic factors, and to investigate the broader health implications for affected men. The emerging data suggest that male infertility is not only a reproductive issue but also a potential predictor of chronic diseases, including autoimmune disorders, cancer, and premature death. Additionally, the inheritance of male factor infertility and its potential effects on offspring health remains indeterminate. Studies have shown conflicting results regarding the impact of parental infertility and fertility treatments on the semen quality and reproductive health of offspring. This review summarizes the current understanding of the genetic causes of male infertility, highlights the impact of chromosomal disorders, reviews the spectrum of sperm quality and hormonal profiles, and discourses on the need for further research to clarify the relationships between parental subfertility, male infertility, and offspring health. By investigating these complex interrelationships, future research can help shape more effective diagnostic and treatment strategies for male infertility and its broader implications for men’s health and future generations. Full article

N6-methyladenosine (m6A) modification is a key methylation modification involved in reproductive processes. Myostatin gene editing (MT) in cattle is known to enhance muscle mass and productivity. However, the changes in m6A modification in MT bull sperm remain poorly understood. In the MT and wild-type (WT) groups, we identified 25,542 and 22,253 m6A peaks, respectively, mainly concentrated in the coding sequence (CDS) and 3′ untranslated region (UTR) of genes. The MT group showed an increase in gene transcription, but there was no significant difference in the overall m6A peaks pattern. There was also no significant difference in m6A motif and chromosome distribution between MT and WT groups. Most genes had less m6A modification sites. A total of 1120 m6A peaks were significantly different, corresponding to 1053 differentially m6A-methylated genes (DMMGs). These DMMGs are mainly associated with G protein-coupled receptor signaling pathways and the overall composition of the cell membrane. Furthermore, an MCL clustering analysis of 111 differentially m6A-methylated and expressed genes identified seven key genes (RHOA, DAAM1, EXOC4, GNA12, PRICKLE1, SCN1A, and STXBP5L), with the cytoskeleton and migration-related gene, RHOA, being the most important gene located at the center of the gene network. However, the analysis of sperm morphology and motility indicated no significant changes in semen volume, sperm count, sperm viability, plasma membrane integrity, acrosome membrane integrity, or mitochondrial membrane integrity. This study provides a map of m6A methylation in spermatozoa from MT and WT bulls, identifies key differential m6A genes that are affected by the myostatin gene but do not affect sperm morphology and viability in MT bulls, and provides a theoretical basis for the breeding quality of MT bulls. Full article

Background and Objectives: Recurrent pregnancy loss (RPL) is a multifactorial condition, encompassing genetic, anatomical, immunological, endocrine, as well as infectious and environmental factors; however, the etiology remains elusive in a substantial number of cases. Genetic factors linked to RPL include parental karyotype abnormalities (e.g., translocations, inversions, copy number variants), an increase in sperm aneuploidy, fetal microchimerism, severe skewing of X chromosome inactivation, and various gene polymorphisms. Our study aims to explore the value of routine conventional parental karyotyping in couples with RPL. Materials and Methods: A total of 213 couples (426 individuals) with a history of RPL were enrolled in this retrospective study. The peripheral blood samples included in this study were referred to the Human Genomics Laboratory of the University of Medicine and Pharmacy in Craiova, Romania, for conventional cytogenetic analysis between January 2013 and December 2023, by the Outpatient Medical Genetics Clinic of the Emergency Clinical County Hospital of Craiova. Chromosome analysis was performed using standard protocols and karyotypes were reported according to ISCN. Results: Out of 426 patients provided with conventional G-banded chromosome analysis, 410 had a normal karyotype (96.2%) and 16 had chromosome abnormalities (3.8%). The most common chromosomal abnormalities were reciprocal and Robertsonian translocations, with chromosomes 8, 11, 14, and 21 being most frequently involved. A single numerical anomaly was detected (47,XYY). One or multiple chromosomal polymorphisms were identified in 104 subjects (24.4%). In addition, we conducted a stratified analysis of the unselected group and detected chromosome abnormalities in only four cases (0.94%). Conclusions: Our results are consistent with recommendations for paternal karyotyping after an individual risk assessment in instances such as a previous live birth with congenital anomalies and/or the detection of unbalanced chromosomes or a translocation in product of conception or chorionic villi/amniotic fluid samples. In the absence of a positive history, blindly karyotyping couples may prove too expensive and labor intensive, while providing no information on fertility status or live birth rates. Full article

Pre-implantation genetic testing (PGT) is a crucial process for selecting embryos created through assisted reproductive technology (ART). Couples with chromosomal rearrangements, infertility, recurrent miscarriages, advanced maternal age, known single-gene disorders, a family history of genetic conditions, previously affected pregnancies, poor embryo quality, or congenital anomalies may be candidates for PGT. Preimplantation genetic testing for aneuploidies (PGT-A) enables the selection and transfer of euploid embryos, significantly enhancing implantation rates in assisted reproduction. Fluorescence in situ hybridization (FISH) is the preferred method for analyzing biopsied cells to identify these abnormalities. While FISH is a well-established method for identifying sperm aneuploidy, NGS offers a more comprehensive assessment of genetic material, potentially enhancing our understanding of male infertility. Chromosomal abnormalities, arising during meiosis, can lead to aneuploid sperm, which may hinder embryo implantation and increase miscarriage rates. This review provides a comparative analysis of fluorescence in situ hybridization (FISH) and next-generation sequencing (NGS) in sperm evaluations, focusing on their implications for preimplantation genetic testing. This analysis explores the strengths and limitations of FISH and NGS, aiming to elucidate their roles in improving ART outcomes and reducing the risk of genetic disorders in offspring. Ultimately, the findings will inform best practices in sperm evaluations and preimplantation genetic testing strategies. Full article

ProAKAP4 is a sperm structural protein that regulates motility through the PKA-dependent cAMP signaling pathway, which is synthesized as an X chromosome-linked member of the gene family. This study aims to determine the optimal level of proAKAP4 for evaluating sexed semen through investigating its relationship with the longevity of sperm quality in sexed Holstein bull sperm. A total of 30 sexed sperm samples (bearing X chromosomes) from 30 distinct Holstein bulls (n = 30) were analyzed. The frozen bull sperm samples were assessed for their proAKAP4 levels, mitochondrial membrane potential, plasma membrane and acrosome integrity (PMAI), and spermatozoa movement parameters at hours 0 and 3 after thawing. The proAKAP4 levels in the sexed sperm samples ranged from 16.35 to 72.10 ng/10 M spz, with an average of 37.18 ± 15.1 ng/10 M spz. A strong positive correlation was observed between proAKAP4 levels and total motility, progressive motility, PMAI, high mitochondrial membrane potential, VAP, and VCL values after 3 h of incubation, when compared to post-thaw analyses. The results also reveal that spermatozoa with proAKAP4 levels of ≥40 ng/10 M spz exhibit higher quality. In conclusion, the level of proAKAP4 in sexed sperm aligns with previous studies and shows potential as a biomarker for assessing the longevity of sexed sperm quality. Full article