An Analysis of the Structural Relationship between Thyroid Hormone-Signaling Disruption and Polybrominated Diphenyl Ethers: Potential Implications for Male Infertility.

Polybrominated diphenyl ethers (PBDEs) are a common class of anthropogenic organobromine chemicals with fire-retardant properties and are extensively used in consumer products, such as electrical and electronic equipment, furniture, textiles, and foams. Due to their extensive use, PBDEs have wide eco-chemical dissemination and tend to bioaccumulate in wildlife and humans with many potential adverse health effects in humans, such as neurodevelopmental deficits, cancer, thyroid hormone disruption, dysfunction of reproductive system, and infertility. Many PBDEs have been listed as chemicals of international concern under the Stockholm Convention on Persistent Organic Pollutants. In this study, the aim was to investigate the structural interactions of PBDEs against thyroid hormone receptor (TRα) with potential implications in reproductive function. Structural binding of four PBDEs, i.e., BDE-28, BDE-100, BDE-153 and BDE-154 was investigated against the ligand binding pocket of TRα using Schrodinger's induced fit docking, followed by molecular interaction analysis and the binding energy estimation. The results indicated the stable and tight binding of all four PDBE ligands and similarity in the binding interaction pattern to that of TRα native ligand, triiodothyronine (T3). The estimated binding energy value for BDE-153 was the highest among four PBDEs and was more than that of T3. This was followed by BDE-154, which is approximately the same as that of TRα native ligand, T3. Furthermore, the value estimated for BDE-28 was the lowest; however, the binding energy value for BDE-100 was more than BDE-28 and close to that of TRα native ligand, T3. In conclusion, the results of our study suggested the thyroid signaling disruption potential of indicated ligands according to their binding energy order, which can possibly lead to disruption of reproductive function and infertility.

Sperm proteins ODF2 and PAWP as markers of fertility in breeding bulls.

Low fertility is the single most important factor limiting livestock reproductive performance, adversely affecting the cattle industry and causing millions of dollars of economic loss. In the livestock industry, male fertility is of crucial importance for the reproductive performance of livestock. However, there is a lack of reliable biomarkers to predict bull fertility in artificial insemination service. The objective of this study was to identify sperm proteins as biomarkers for bull fertility. To discover candidate sperm quality biomarkers, sperm proteome profiling was conducted in extreme high- and extreme low-fertile bulls selected from a pool of 1000 AI sires with varied fertility. Thirty-two differentially expressed proteins were identified. Among them, high levels of sperm outer dense fiber of sperm tails 2 (ODF2) and post-acrosomal assembly of sperm head protein (PAWP/WBP2NL) represented the most extreme differences in quantity between high- and low-fertility bulls. Protein immunodetection and flow cytometry used to validate these putative fertility markers in a combined cohort of 154 AI sires. Both ODF2 and PAWP correlated significantly with fertility. In conclusion, ODF2 and PAWP can be used to assess semen quality and predict sire fertility.

Contributions of seminal plasma proteins to fertilizing ability of bull sperm: A meta-analytical review.

Seminal plasma is a dynamic, intricate combination of fluids from the testicles, epididymides, seminal vesicles, bulbourethral glands, and prostate, containing molecules that modulate sperm functions, post-fertilization events, and the female reproductive tract physiology. Significant variations in sperm parameters and fertility status of bulls relate to differences in the seminal plasma proteome. In this framework, a meta-analytical study was conducted examining 29 studies (published between 1990 and 2021) to ascertain the effects of seminal fluid proteins on parameters associated with bull fertility and the influence of distinct methodologies on such effects. Our results revealed that seminal proteins ameliorate sperm parameters, such as motility, integrity, capacitation, and fertilizing ability, and favours sperm protection. Seminal binder of sperm proteins and beta-defensin 126 highly favoured sperm protection when cells were collected from the epididymis by retrograde flux and analysed under room temperature conditions. Furthermore, seminal proteins improved the motility and quality of Bos taurus sperm collected by artificial vagina, mainly in the presence of heparin-binding proteins. The key limitations faced by this meta-analysis were the paucity of studies evaluating the effects of whole seminal fluid proteins and the limited number of studies conducted in vivo. In conclusion, the present meta-analytical study confirms that seminal proteins improve fertility-related parameters in the bovine species. However, methodological strategies used by authors are diverse, with distinct endpoints and methods. Thus, the translational aspects of seminal plasma research should be taken into consideration to precisely define how seminal proteins can be harnessed to advance reproductive biotechnology.

Functional attributes of seminal proteins in bull fertility: a systematic review.

Proteomic approaches have been widely used in reproductive studies to uncover protein biomarkers of bull fertility. Seminal plasma is one of the most relevant sources of these proteins that may influence sperm physiology. Nonetheless, there are still gaps in existing knowledge in the functional attributes of seminal proteins. Thus, we reviewed the relationships between seminal plasma proteins and bull fertility by conducting a systematic review with data obtained from 71 studies. This review showed that the associations related to fertility improvement with the use of total seminal plasma proteins are still controversial. None of the studies explored the sperm fertilizing ability following these interactions. By contrast, the exposure to a single protein, such as osteopontin, binder of sperm proteins, and heparin binding proteins, can increment sperm motility, capacitation, and fertilizing ability by modulating intracellular calcium concentrations, removing lipids from sperm membranes, and regulating the acrosome reaction. Variations in protein analyses and the protein contents and their abundances between animals contributed to the difficulty of establishing protein biomarkers of fertilizing potential of the bull sperm. Indeed, the heterogenicity of methodologies was a limitation of this review. Standardized methods of seminal protein analyses, as well as sperm endpoints, may minimize such discrepancies. In conclusion, potential biomarkers of sperm parameters are still to be established. Future studies should evaluate protein isoforms and how they interact with sperm to ascertain their biological functions.

EThcD and 213 nm UVPD for Top-Down Analysis of Bovine Seminal Plasma Proteoforms on Electrophoretic and Chromatographic Time Frames.

Seminal plasma is a critical and complex fluid that carries sperm to eggs to initiate the fertilization process. Here, we present a top-down mass spectrometry (TDMS) strategy for identifying proteins and posttranslational modifications (PTMs) in bovine seminal plasma. In this study, proteins were separated using sheathless capillary zone electrophoresis (CZE)-MS and reversed-phase liquid chromatography (LC)-MS, and then fragmented using electron-transfer/higher-energy collisional dissociation (EThcD) and 213 nm ultraviolet photodissociation (213 nm UVPD) to provide more comprehensive information about the proteomic landscape of this biological fluid. Four hundred and seventeen proteoforms were identified by sheathless CZE-MS, and one hundred and seventy-two species were unique to this method. LC-MS identified 3090 proteoforms, including 1707 unique species. All identifications were within ±10 ppm (mass error) and with a P-Score ≤1 × 10-04. Pooling results (triplicate measurements) from sheathless CZE-MS and LC-MS resulted in the identification of 1433 proteoforms (EThcD) and 2151 proteoforms (213 nm UVPD) with 612 species unique for EThcD and 1021 for 213 nm UVPD. The average sequence coverage was found to be higher for EThcD (28%) than for 213 nm UVPD (23%). The use of sheathless CZE-MS and LC-MS with EThcD and 213 nm UVPD provided complementary protein profiling and proteoform data that were more comprehensive than those of either method alone.

Expression profile of Toll-like receptor 4 in rat testis and epididymis throughout postnatal development.

Toll-like receptors (TLRs) belonging to pattern recognition receptors are involved in maintaining testicular and epididymal immune homeostasis. The purpose of the current study was to investigate TLR4 expression in rat testis and epididymis throughout postnatal development. Weak staining was detected in peritubular myoid cells and immature Sertoli cells while no staining was observed in gonocytes during prepubertal period. However, TLR4 expression began to appear in spermatocytes in pubertal period and gradually increased in spermatids. An intense staining was observed in steps 5-19 spermatids in post pubertal and mature periods. Similarly, TLR4 expression in the testes steadily increased from pubertal period to mature period. Puberty also caused a significant increase in TLR4 expression in epididymis. TLR4 expression in cauda epididymis was lower as compared to those of other epididymal segments. The majority of epididymal epithelial cells exhibited apical TLR4 expression, whereas basal cells showed intense intracytoplasmic immunoreaction. We detected an intense staining in epididymal smooth muscle cells. The expression levels of TLR4 showed dynamic changes in both spermatogenic cells, and entire testicular and epididymal tissues during postnatal development. These results suggest that TLR4 expression contributes not only to inflammation but also to the development of spermatogenic cells.

Sperm miR-15a and miR-29b are associated with bull fertility.

MicroRNAs modulate male fertility by regulating gene expression. In this study, dynamics of sperm miR-15a, miR-29b and miR-34a from high fertility (HF) and low fertility (LF) bulls using RT-qPCR were evaluated. Bioinformatic tools were employed to ascertain genes of interest of the sperm miRNAs. The expression levels of p53, BCL2, BAX and DNMT1 in bull spermatozoa were determined by immunoblotting. MicroRNA levels of miR-15a and miR-29 were higher in LF sires when compared with those present in HF bulls. Expression levels of miR-34a did not differ between the two groups. We found an inverse correlation between miR-15a and bull fertility. MiR29-b was also negatively associated with fertility scores. BCL2 and DNMT1 were higher in HF bulls while BAX was higher in the LF group. Our data showed a positive correlation between BCL2 and bull fertility. In addition, DNMT1 was positively associated with bull fertility. Furthermore, levels of BAX were negatively linked with bull fertility scores. Identification of miRNAs found in the spermatozoa of sires with different in vivo fertility helps understand the alterations in the fertilising capacity from cattle and other mammals. These potential biomarkers can be used in reproductive biotechnology as fertility markers to assess semen quality and predict male fertility.

Expression dynamics of Integrin Subunit Beta 5 in bovine gametes and embryos imply functions in male fertility and early embryonic development.

Integrins have been shown to act as signalling receptors, and they primarily recognise extracellular matrix ligands on the oocyte surface. However, their possible roles in oocyte activation and embryo development are not clearly understood. The objectives of this study were to evaluate expression of Integrin Subunit Beta 5 (ITGß5) in bovine sperm, oocytes, and early embryos and to ascertain the evolutionary conservation of ITGß5. To accomplish these objectives, we used western blotting to study expression levels of ITGß5 protein in sperm and RT-qPCR to determine expression levels of ITGß5 transcripts in oocytes and embryos. We have also used bioinformatic analysis to determine the evolutionary conservation of the ITGß5 protein among various species. Western blotting showed that ITGß5 protein was detectable in bull sperm. Moreover, results of RT-qPCR showed that levels of ITGß5 were significantly higher in the two-cell embryos, followed by the 8-16-cell embryos. However, no significant difference in expression levels were noted for the morula and blastocyst stages as compared to MII oocytes. Bioinformatic analysis revealed that ITGß5 is conserved among various species. We conclude that expression of ITGß5 in bovine gametes and embryos implies an important role in fertilisation and embryogenesis.

Testis specific histone 2B is associated with sperm chromatin dynamics and bull fertility-a pilot study.

BACKGROUND: Bull fertility is the degree of sperm's ability to fertilize and activate the egg and support embryo development, and this is critical for herd reproductive performance. We used the bull as a unique model organism for the study of male fertility because cattle genetics and physiology is similar to those of other mammals including humans. Moreover, reliable fertility data along with well-established in vitro systems are available for bovine. The objective of this original study was to ascertain evolutionary diversification and expression dynamics of Testis Specific Histone 2B (TH2B) in sperm from Holstein bulls with different fertility scores. METHODS: The intensity of TH2B was determined by using flow cytometry in sperm from 13 high and 13 low fertility bulls. Expression levels of TH2B were measured using immunofluorescence and Western blotting in sperm from five high and five low fertility bulls. Sequence identity, evolutionary distance and interactome of TH2B were evaluated by dotmatcher, STRING and Cytoscape. Data were analyzed using linear mixed effects model and regression plots were drawn. RESULTS: The intensity of TH2B as measured by flow cytometry was significantly affected by an interaction between fertility group and fertility score (P = 0.0182). The intensity of TH2B in sperm from the high fertility group decreased (P = 0.0055) as fertility increased. TH2B was constantly detectable in sperm and expression levels of TH2B decreased in relation to fertility in sperm from the high fertility group (P = 0.018). TH2B biological functions include male gamete generation, chromosome organization, DNA packaging, DNA conformation change, chromatin organization, nucleosome organization, chromatin disassembly, spermatid nucleus elongation, spermatid nucleus differentiation, sperm motility, chromatin organization, chromatin condensation, chromatin silencing, nucleus organization, and chromatin remodeling (P < 0.05). CONCLUSIONS: We elucidated the cellular localization and molecular physiology of TH2B using both computational and cell biology approaches. In addition to advancing the fundamental science of mammalian male gamete, the present findings can be potentially used to evaluate semen quality and predict male fertility in the future. TRIAL REGISTRATION: This study did not involve any live animals. We did not perform any anesthesia, euthanasia, or any kind of animal sacrifice. The cryopreserved semen samples were obtained from Alta Genetics, Inc., Watertown, WI, USA. All samples were preserved in liquid nitrogen.

Sperm protamine-status correlates to the fertility of breeding bulls.

During fertilization, spermatozoa make essential contributions to embryo development by providing oocyte activating factors, centrosomal components, and paternal chromosomes. Protamines are essential for proper packaging of sperm DNA; however, in contrast to the studies of oocyte-related female infertility, the influence of sperm chromatin structure on male infertility has not been evaluated extensively. The objective of this study was to determine the sperm chromatin content of bull spermatozoa by evaluating DNA fragmentation, chromatin maturity/protamination, PRM1 protein status, and nuclear shape in spermatozoa from bulls with different fertility. Relationships between protamine 1 (PRM1) and the chromatin integrity were ascertained in spermatozoa from Holstein bulls with varied (high vs. low) but acceptable fertility. Sperm DNA fragmentation and chromatin maturity (protamination) were tested using Halomax assay and toluidine blue staining, respectively. The PRM1 content was assayed using Western blotting and in-gel densitometry, flow cytometry, and immunocytochemistry. Fragmentation of DNA was increased and chromatin maturity significantly reduced in spermatozoa from low-fertility bulls compared to those from high-fertility bulls. Field fertility scores of the bulls were negatively correlated with the percentage of spermatozoa displaying reduced protamination and fragmented DNA using toluidine blue and Halomax, respectively. Bull fertility was also positively correlated with PRM1 content by Western blotting and flow cytometry. However, detection of PRM1 content by Western blotting alone was not predictive of bull fertility. In immunocytochemistry, abnormal spermatozoa showed either a lack of PRM1 or scattered localization in the apical/acrosomal region of the nuclei. The nuclear shape was distorted in spermatozoa from low-fertility bulls. In conclusion, we showed that inadequate amount and localization of PRM1 were associated with defects in sperm chromatin structure, coinciding with reduced fertility in bulls. These findings are highly significant because they reveal molecular and morphological phenotypes of mammalian spermatozoa that influence fertility.

Sperm superoxide dismutase is associated with bull fertility.

Decreasing mammalian fertility and sperm quality have created an urgent need to find effective methods to distinguish non-viable from viable fertilising spermatozoa. The aims of the present study were to evaluate expression levels of ?-tubulin 2C (TUBB2C), heat shock protein 10 (HSP10), hexokinase 1 (HXK1) and superoxide dismutase 1 (SOD1) in spermatozoa from Holstein bulls with varying fertility using western blotting and to analyse the biological networks of these key sperm proteins using a bioinformatics software (Metacore; Thomson-Reuters, Philadelphia, PA, USA). The rationales behind this study were that the sperm proteins play crucial roles in fertilisation and early embryonic development in mammals and ascertaining the biological networks of the proteins helps us better understand sperm physiology and early mammalian development. The results showed that expression of SOD1 was higher in spermatozoa from high fertility bulls (PPin vivo bull fertility. The findings are important because they illuminate molecular and cellular determinants of sperm viability and the identified protein markers can be used to determine bull fertility.

Environmental stressors influencing hormones and systems physiology in cattle.

Environmental stressors undoubtedly influence organismal biology, specifically the endocrine system that, in turn, impact cattle at the systems physiology level. Despite the significant advances in understanding the genetic determinants of the ideal dairy or beef cow, there is a grave lack of understanding of the systems physiology and effects of the environmental stressors that interfere with the endocrine system. This is a major problem because the lack of such knowledge is preventing advances in understanding gene-environment interactions and developing science-based solutions to these challenges. In this review, we synthesize the current knowledge on the nature of the major environmental stressors, such as climate (heat, cold, wind, and humidity), nutrition (feeds, feeding systems, and endocrine disruptors) and management (housing density and conditions, transportation, weaning practices). We summarize the impact of each one of these factors on cattle at the systems level, and provide solutions for the challenges.

Sperm long non-coding RNAs as markers for ram fertility.

It is critical in sheep farming to accurately estimate ram fertility for maintaining reproductive effectiveness and for production profitability. However, there is currently a lack of reliable biomarkers to estimate semen quality and ram fertility, which is hindering advances in animal science and technology. The objective of this study was to uncover long non-coding RNAs (lncRNAs) in sperm from rams with distinct fertility phenotypes. Mature rams were allocated into two groups: high and low fertility (HF; n = 31; 94.5 ± 2.8%, LF; n = 25; 83.1 ± 5.73%; P = 0.028) according to the pregnancy rates sired by the rams (average pregnancy rate; 89.4 ± 7.2%). Total RNAs were isolated from sperm of the highest- and lowest-fertility rams (n = 4, pregnancy rate; 99.2 ± 1.6%, and 73.6 ± 4.4%, respectively) followed by next-generation sequencing of the transcripts. We uncovered 11,209 lncRNAs from the sperm of rams with HF and LF. In comparison to each other, there were 93 differentially expressed (DE) lncRNAs in sperm from the two distinct fertility phenotypes. Of these, 141 mRNAs were upregulated and 134 were downregulated between HF and LF, respectively. Genes commonly enriched for 9 + 2 motile cilium and sperm flagellum were ABHD2, AK1, CABS1, ROPN1, SEPTIN2, SLIRP, and TEKT3. Moreover, CABS1, CCDC39, CFAP97D1, ROPN1, SLIRP, TEKT3, and TTC12 were commonly enriched in flagellated sperm motility and sperm motility. Differentially expressed mRNAs were enriched in the top 16 KEGG pathways. Targets of the differentially expressed lncRNAs elucidate functions in cis and trans manner using the genetic context of the lncRNA locus, and lncRNA sequences revealed 471 mRNAs targets of 10 lncRNAs. This study illustrates the existence of potential lncRNA biomarkers that can be implemented in analyzing the quality of ram sperm and determining the sperm fertility and is used in breeding soundness exams for precision livestock farming to ensure food security on a global scale.

Risk Factor Analysis and Genetic Parameter Estimation for Pre-Weaning Mortality Traits in Boer, Spanish, and Crossbred Goat Kids.

The objectives of this study were to evaluate fixed risk factors associated with PWM and to estimate genetic parameters for PWM. A total of 927 birth records from a mixed population of purebred and crossbred Boer and Spanish goats born between 2016 and 2023 at the International Goat Research Center (IGRC) were used for this study. Four binary traits were studied: D0-3 (death within 3 days after birth), D4-60 (death between 4 and 60 days), D61-90 (death between 61 and 90 days), and D0-90 (death within 90 days). Logistic regression models were used to evaluate the risk factors associated with PWM traits. Bayesian threshold models and Gibbs sampling were used to estimate the genetic parameters. Birth weight, season, litter size, sex, dam age, breed, and heterosis were found to be significantly associated with at least one of the PWM traits. Heritability estimates were 0.263, 0.124, 0.080, and 0.207, for D0-3, D4-60, D61-90, and D0-90, respectively. The genetic correlations between the studied traits ranged from 0.892 (D0-3 and D0-90) to 0.999 (D0-3 and D61-90). These results suggest that PWM in goats is influenced by both non-genetic and genetic factors and can be reduced by management, genetic selection, and crossbreeding approaches.

Molecular morphology and function of bull spermatozoa linked to histones and associated with fertility.

Sub-par fertility in bulls is influenced by alterations in sperm chromatin, and it might not be solved with increased sperm concentration in artificial insemination. Appropriate histone retention during sperm chromatin condensation plays critical roles in male fertility. The objective of this study was to determine failures of sperm chromatin condensation associated with abnormal persistence or accessibility of histones by aniline blue (ANBL) test, expression levels, and cellular localizations of one variant and two core histones (H3.3, H2B, and H4 respectively) in the spermatozoa of low-fertility (LF) vs high-fertility (HF) bulls. The expression levels and cellular localizations of histones in spermatozoa were studied using immunoblotting, immunocytochemistry, and staining methods. The bioinformatics focused on the sequence identity and evolutionary distance of these proteins among three mammalian species: bovine, mouse, and human. We demonstrated that ANBL staining was different within the LF (1.73 (0.55, 0.19)) and HF (0.67 (0.17, 0.06)) groups (P<0.0001), which was also negatively correlated with in vivo bull fertility (r=-0.90, P<0.0001). Although these histones were consistently detectable and specifically localized in bull sperm cells, they were not different between the two groups. Except H2B variants, H3.3 and H4 showed 100% identity and were evolutionarily conserved in bulls, mice and humans. The H2B variants were more conserved between bulls and humans, than in mice. In conclusion, we showed that H2B, H3.3, and H4 were detectable in bull spermatozoa and that sperm chromatin condensation status, changed by histone retention, is related to bull fertility.

Interrelationships between apoptosis and fertility in bull sperm.

Male fertility, the ability of sperm to fertilize and activate the egg and support early embryogenesis, is vital for mammalian reproduction. Despite producing adequate numbers of sperm with normal motility and morphology, some males suffer from low fertility whose molecular mechanisms are not known. The objective was to determine apoptosis in sperm from high and low fertility bulls and its relationship with male fertility. DNA damage, phosphatidylserine (PS) translocation, and expression of pro- and anti-apoptotic proteins (BAX and BCL-2) in the sperm were determined using TUNEL, Annexin V, and immunoblotting approaches, respectively. Amounts of apoptotic spermatozoa were 2.86 (± 1.31) and 3.00 (± 0.96) in high and low fertility bulls, respectively (P=0.548), and were not correlated with fertility. There was a negative correlation between early necrotic spermatozoa and viable spermatozoa (r = -0.99, P<0.0001). Fertility scores were correlated with live spermatozoa detected by eosin-nigrosin test and necrotic spermatozoa determined via flow cytometry (r = -0.49, P<0.006 and r = -0.266, P<0.0113, respectively). BAX level was higher in low fertile group than high fertile group; however, this difference was not statistically significant due to the variations of bull samples (Bull 1-3 vs. Bull 4-5) in low fertile group (P<0.283). BCL-2 was not detectable in any of the sperm samples. The results shed light onto molecular and cellular underpinnings of male fertility.

Androgen receptor signaling and pyrethroids: Potential male infertility consequences.

Infertility is a global health concern inflicting a considerable burden on the global economy and a severe socio-psychological impact. Approximately 15% of couples suffer from infertility globally, with a male factor contribution of approximately 50%. However, male infertility remains largely unexplored, as the burden of infertility is mostly assigned to female people. Endocrine-disrupting chemicals (EDCs) have been proposed as one of the factors causing male infertility. Pyrethroids represent an important class of EDCs, and numerous studies have associated pyrethroid exposure with impaired male reproductive function and development. Therefore, the present study investigated the potentially toxic effects of two common pyrethroids, cypermethrin and deltamethrin, on androgen receptor (AR) signaling. The structural binding characterization of cypermethrin and deltamethrin against the AR ligand-binding pocket was performed using Schrodinger's induced fit docking (IFD) approach. Various parameters were estimated, such as binding interactions, binding energy, docking score, and IFD score. Furthermore, the AR native ligand, testosterone, was subjected to similar experiments against the AR ligand-binding pocket. The results revealed commonality in the amino acid-binding interactions and overlap in other structural parameters between the AR native ligand, testosterone, and the ligands, cypermethrin and deltamethrin. The estimated binding energy values of cypermethrin and deltamethrin were very high and close to those calculated for AR native ligand, testosterone. Taken together, the results of this study suggested potential disruption of AR signaling by cypermethrin and deltamethrin, which may result in androgen dysfunction and subsequent male infertility.

Delivering value from sperm proteomics for fertility.

Fertilization of an egg by a spermatozoon sets the stage for mammalian development. Viable sperm are a prerequisite for successful fertilization and beyond. Spermatozoa have a unique cell structure where haploid genomic DNA is located in a tiny cytoplasmic space in the head, mitochondria in the midpiece and then the tail, all enclosed by several layers of membrane. Proteins in sperm play vital roles in motility, capacitation, fertilization, egg activation and embryo development. Molecular defects in these proteins are associated with low fertility or in some cases, infertility. This review will first summarize genesis, molecular anatomy and physiology of spermatozoa, fertilization, embryogenesis and then those proteins playing important roles in various aspects of sperm physiology.

Dynamics of microRNAs in bull spermatozoa.

BACKGROUND: MicroRNAs are small non-coding RNAs that regulate gene expression and thus play important roles in mammalian development. However, the comprehensive lists of microRNAs, as well as, molecular mechanisms by which microRNAs regulate gene expression during gamete and embryo development are poorly defined. The objectives of this study were to determine microRNAs in bull sperm and predict their functions. METHODS: To accomplish our objectives we isolated miRNAs from sperm of high and low fertility bulls, conducted microRNA microarray experiments and validated expression of a panel of microRNAs using real time RT-PCR. Bioinformatic approaches were carried out to identify regulated targets. RESULTS: We demonstrated that an abundance of microRNAs were present in bovine spermatozoa, however, only seven were differentially expressed; hsa-aga-3155, -8197, -6727, -11796, -14189, -6125, -13659. The abundance of miRNAs in the spermatozoa and the differential expression in sperm from high vs. low fertility bulls suggests that the miRNAs possibly play important functions in the regulating mechanisms of bovine spermatozoa. CONCLUSION: Identification of specific microRNAs expressed in spermatozoa of bulls with different fertility phenotypes will help better understand mammalian gametogenesis and early development.

Mycotoxin alpha-zearalenol impairs the quality of preimplantation porcine embryos.

Alpha-Zearalenol (α-ZEA) is one of derivatives from Zearalenone (ZEA) which impacts mammalian reproduction and development. Previous studies have shown that pigs are sensitive to the estradiol-like effects of α-ZEA. However, the effect of α-ZEA for the early embryonic development has not been fully studied. The objective of this study was to identify the direct toxicity of α-ZEA on porcine preimplantation embryonic development, embryo quality and expression of developmentally important genes. Presumptive zygotes were cultured in porcine zygote medium 3 (PZM-3) in the presence of α-ZEA (n=2,957) or 17ß-estradiol (E2) (n=1,333) dissolved in 0.1% Dimethyl Sulfoxide (DMSO) from 24 to 84 h post insemination followed by determination of apoptotic cell numbers and transcript levels of BAX, BCL2L1 and POU5F1 in blastocysts. Cleavage rates on day 2 were significantly decreased in 10, 30 and 60 µM α-ZEA groups; whereas blastocyst rates on day 6 were significantly decreased in the 30 and 60 µM of α-ZEA groups. Only the 100 µM E2 group significantly decreased cleavage and blastocyst rates. Total cell numbers (TCN) in blastocysts were significantly lower in the 10 µM α-ZEA group, but no differences in apoptotic cell rates were found. The expression levels of POU5F1 and BCL2L1 transcripts were similar; however, levels of BAX transcripts and the BAX/BCL2L1 ratio were increased in both α-ZEA groups. Since α-ZEA and E2 did not elicit similar effects, results suggest that α-ZEA might impact porcine preimplantation embryonic development through pathways other than estrogen receptor binding.