Abnormal amino acid synthesis and glutathione metabolism may affect PCOS blastocyst development: an examination of in vitro mouse blastocysts model utilizing RNA-sequencing.

BACKGROUND: Extensive research has been conducted on embryonic developmental disorders linked to Polycystic Ovary Syndrome (PCOS), a pathological condition that affects 5-10% of women and is characterized by irregularities in the menstrual cycle and infertility. By employing RNA sequencing (RNA-seq), we performed an in-depth investigation of PCOS-related changes in gene expression patterns at the mouse blastocyst stage. METHODS: The zygotes of female B6D2 mice were obtained and then differentiated into blastocysts in K + Simplex Optimised Medium (KSOM) cultures containing exo-NC (negative control for exosomes) or exo-LIPE-AS1 (a novel exosomal marker of PCOS). Subsequently, blastocysts were collected for RNA-seq. The bioinformatics was performed to analyze and compare the differences of gene expression profile between blastocysts of control and PCOS group. RESULTS: There were 1150 differentially expressed genes (DEGs) between the two groups of mouse blastocysts; 243 genes were upregulated and 907 downregulated in the blastocysts of the exo-LIPE-AS1 group compared to those of the exo-NC group. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed that the genes involved in amino acid synthesis and glutathione metabolic pathways were down-regulated in exo-LIPE-AS1 group. CONCLUSION: This study has revealed that blastocyst developmental retardation may be associated with the downregulation of amino acid synthesis and glutathione metabolism, which may affect energy metabolism, biosynthesis, cellular osmotic pressure, antioxidant synthesis, ROS clearance or mitochondrial function, and ultimately cause blastocyst cell development abnormalities. Our research offers encouraging data on the mechanisms underlying aberrant embryonic development in patients with PCOS as well as potential treatment strategies.

Application of genome tagging technology in elucidating the function of sperm-specific protein 411 (Ssp411).

The genome tagging project (GTP) plays a pivotal role in addressing a critical gap in the understanding of protein functions. Within this framework, we successfully generated a human influenza hemagglutinin-tagged sperm-specific protein 411 (HA-tagged Ssp411) mouse model. This model is instrumental in probing the expression and function of Ssp411. Our research revealed that Ssp411 is expressed in the round spermatids, elongating spermatids, elongated spermatids, and epididymal spermatozoa. The comprehensive examination of the distribution of Ssp411 in these germ cells offers new perspectives on its involvement in spermiogenesis. Nevertheless, rigorous further inquiry is imperative to elucidate the precise mechanistic underpinnings of these functions. Ssp411 is not detectable in metaphase II (MII) oocytes, zygotes, or 2-cell stage embryos, highlighting its intricate role in early embryonic development. These findings not only advance our understanding of the role of Ssp411 in reproductive physiology but also significantly contribute to the overarching goals of the GTP, fostering groundbreaking advancements in the fields of spermiogenesis and reproductive biology.

Deficiency of MFSD6L, an acrosome membrane protein, causes oligoasthenoteratozoospermia in humans and mice.

Oligoasthenoteratozoospermia is an important factor affecting male fertility and has been found to be associated with genetic factors. However, there are still a proportion of oligoasthenoteratozoospermia cases that cannot be explained by known pathogenic genetic variants. Here, we perform genetic analyses and identify bi-allelic loss-of-function variants of MFSD6L from an oligoasthenoteratozoospermia-affected family. Mfsd6l knock-out male mice also present male subfertility with reduced sperm concentration, motility, and deformed acrosomes. Further mechanistic analyses reveal that MFSD6L, as an acrosome membrane protein, plays an important role in the formation of acrosome by interacting with the inner acrosomal membrane protein SPACA1. Moreover, poor embryonic development is consistently observed after intracytoplasmic sperm injection treatment using spermatozoa from the MFSD6L-deficient man and male mice. Collectively, our findings reveal that MFSD6L is required for the anchoring of sperm acrosome and head shaping. The deficiency of MFSD6L affects male fertility and causes oligoasthenoteratozoospermia in humans and mice.

Monitoring Blood Glucose in Mouse Offspring after Intracytoplasmic sperm injection - Embryo Transfer.

Human lifespan is considerably long, while mouse models can simulate the entire human lifespan in a relatively short period, with one year of mouse life roughly equivalent to 40 human years. Intracytoplasmic sperm injection (ICSI) is a commonly used assisted reproductive technology in clinical practice. However, given its relatively recent emergence about 30 years ago, the long-term effects of this technique on human development remain unclear. In this study, we established the ICSI combined with embryo transfer (ET) method using a mouse model. The results demonstrated that normal mouse sperm, after undergoing in vitro culture and subsequent ICSI, exhibited a fertilization rate of 89.57% and a two-cell rate of 87.38%. Following ET, the birth rate of offspring was approximately 42.50%. Furthermore, as the mice aged, fluctuations in glucose metabolism levels were observed, which may be associated with the application of the ICSI technique. These findings signify that the mouse ICSI-ET technique provides a valuable platform for evaluating the impact of sperm abnormalities on embryo development and their long-term effects on offspring health, particularly concerning glucose metabolism. This study provides important insights for further research on the potential effects of the ICSI technique on human development, emphasizing the necessity for in-depth investigation into the long-term implications of this technology.

Increased miR-3074-5p expression promotes M1 polarization and pyroptosis of macrophages via ERα/NLRP3 pathway and induces adverse pregnancy outcomes in mice.

Decidual macrophages (dMϕs) play critical roles in regulation of immune-microhomeostasis at maternal-fetal interface during pregnancy, but the underlying molecular mechanisms are still unclear. In this study, it was found that litter size and fetal weight were significantly reduced, whereas the rate of embryo resorption was increased in miR-3074-5p knock-in (3074-KI) pregnant mice, compared to that of wild-type (WT) pregnant mice. Plasma levels of pro-inflammatory cytokines in 3074-KI pregnant mice were also significantly elevated compared to WT pregnant mice at GD7.5. The quantity of M1-Mϕs in uterine tissues of 3074-KI pregnant mice was significantly increased compared to WT pregnant mice at GD13.5. Estrogen receptor-α (ERα) was validated to be a target of miR-3074-5p. Either miR-3074-5p overexpression or ERα knockdown promoted transcriptional activity of NF-κB/p65, induced M1-polarization and pyroptosis of THP1-derived Mϕs, accompanied with increased intracellular levels of cleaved Caspase-1, cleaved IL-1ß, NLRP3, cleaved GSDMD and ASC aggregation. Furthermore, ERα could not only bind to NLRP3 or ASC directly, but also inhibit the interaction between NLRP3 and ASC. The endometrial miR-3074-5p expression level at the middle secretory stage of repeated implantation failure (RIF) patients was significantly decreased compared to that of control fertile women. These data indicated that miR-3074-5p could promote M1 polarization and pyroptosis of Mϕs via activation of NLRP3 inflammasome by targeting ERα, and the dysregulation of miR-3074-5p expression in dMϕs might damage the embryo implantation and placentation by interfering with inflammatory microenvironment at the maternal-fetal interface during early pregnancy.

Loss of ACTL7A causes small head sperm by defective acrosome-acroplaxome-manchette complex.

BACKGROUND: Actin-like 7 A (ACTL7A) is essential for acrosome formation, fertilization and early embryo development. ACTL7A variants cause acrosome detachment responsible for male infertility and early embryonic arrest. In this study, we aim to explore the additional functions of ACTL7A beyond the process of acrosome biogenesis and investigate the possible underlying mechanisms. METHODS: Nuclear morphology analysis was used to observe the sperm head shape of ACTL7A-mutated patients. Actl7a knock-out (KO) mouse model was generated. Immunofluorescence and transmission electron microscopy (TEM) were performed to analyze the structure of spermatids during spermiogenesis. Tandem mass tags labeling quantitative proteomics strategy was employed to explore the underlying molecular mechanisms. The expression levels of key proteins in the pathway were analyzed by western blotting. Intracytoplasmic sperm injection (ICSI)-artificial oocyte activation (AOA) technology was utilized to overcome fertilization failure in male mice with a complete knockout of Actl7a. RESULTS: The new phenotype of small head sperm associated with loss of ACTL7A in patients was discovered, and further confirmed in Actl7a-KO mice. Immunofluorescence and TEM analyses revealed that the deletion of ACTL7A damaged the formation of acrosome-acroplaxome-manchette complex, leading to abnormalities in the shaping of sperm heads. Moreover, a proteomic analysis of testes from WT and Actl7a-KO mice revealed that differentially expressed genes were notably enriched in PI3K/AKT/mTOR signaling pathway which is strongly associated with autophagy. Inhibition of autophagy via PI3K/AKT/mTOR signaling pathway activation leading to PDLIM1 accumulation might elucidate the hindered development of manchette in Actl7a-KO mice. Remarkably, AOA successfully overcame fertilization failure and allowed for the successful production of healthy offspring from the Actl7a complete knockout male mice. CONCLUSIONS: Loss of ACTL7A causes small head sperm as a result of defective acrosome-acroplaxome-manchette complex via autophagy inhibition. ICSI-AOA is an effective technique to rescue male infertility resulting from ACTL7A deletion. These findings provide essential evidence for the diagnosis and treatment of patients suffering from infertility.

Escaping Behavior of Sperms on the Biomimetic Oviductal Surface.

Before fertilization, sperms adhere to oviductal epithelium cells, and only a restrictive number of winner sperms can escape to reach the egg. To study the sperm escape behavior from the oviductal surface, we developed a microfluidic chip to fabricate an adhesive surface and to create a gradient of progesterone (P4) for mimicking the oviduct microenvironment in vivo. We identified three sperm motion patterns in such a microenvironment─anchored spin, run-and-spin, and escaped mode. By using kinetic analysis, we verified the hypothesis that the responsive rotation energy anchored with the adhered sperm head determines whether the sperm is trapped or detaching, which is defined as the hammer flying strategy of successful escape after accumulating energy in the process of rotating. Intriguingly, this hammer-throw escaping is able to be triggered by the P4 biochemical stimulation. Our results revealed the tangled process of sperm escape before fertilization in the ingenious microfluidic system.

Click-iT ® Plus OPP Alexa Fluor ® Protein Synthesis Assay in Embryonic Cells.

This protocol describes a method to assess relative changes in the level of global protein synthesis in the preimplantation embryo using the Click-iT ® Plus OPP Protein Synthesis Assays. In this assay, O-propargyl-puromycin (OPP), an analog of puromycin, is efficiently incorporated into the nascent polypeptide of newly translated proteins in embryonic cells. OPP is fluorescently labeled with a photostable Alexa Fluor TM dye and detected with fluorescence microscopy. The intensity of the fluorescence is quantitatively analyzed. This is a fast, sensitive, and non-radioactive method for the detection of protein synthesis in early embryo development. It provides a tool for analyzing the temporal regulation of protein synthesis, as well as the effects of changes in the embryonic microenvironment, and pharmacological and genetic modulations of embryo development. Graphical abstract: Figure 1. Brief overview of the procedures of the Click-iT ® Plus OPP Alexa Fluor ® protein synthesis assay in embryonic cells. (A) Set up OPP treatments: (1) Set up microdrops containing 50 µL of OPP working solution and label different treatments on the back of culture dishes ( e.g. , T0, T1, T2, and T3); (2) The drops are overlain with 2-3 mm heavy paraffin oil and then equilibrated in incubator for 2 h; (3) Collect the embryos from female reproductive tracts or following in vitro culture in desired treatments; (4) Culture embryos in the equilibrated OPP working solution for 2-6 h. ( B ) Example of OPP detection procedures working with 60-well plates labeled as T0, T1, T2, T3, T4, and T5 for different treatments: (1) The first 60-well plate is used for the procedures of washing, fixation, permeabilization, and Click-iT ® OPP detection. (2) The second 60-well plate is for DNA staining and washing. (C) Slide preparation: (1) Label the required number of slides and set up vaseline coverslip supports; (2) Add mounting medium; (3) Transfer embryos into mounting medium; (4) Set coverslip; (5) Seal the coverslip with nail polish.

Transcriptional Changes in Pearl Millet Leaves under Heat Stress.

High-temperature stress negatively affects the growth and development of plants, and therefore threatens global agricultural safety. Cultivating stress-tolerant plants is the current objective of plant breeding programs. Pearl millet is a multi-purpose plant, commonly used as a forage but also an important food staple. This crop is very heat-resistant and has a higher net assimilation rate than corn under high-temperature stress. However, the response of heat resistant pearl millet has so far not been studied at the transcriptional level. In this study, transcriptome sequencing of pearl millet leaves exposed to different lengths of heat treatment (1 h, 48 h and 96 h) was conducted in order to investigate the molecular mechanisms of the heat stress response and to identify key genes related to heat stress. The results showed that the amount of heat stress-induced DEGs in leaves differs with the length of exposure to high temperatures. The highest value of DEGs (8286) was observed for the group exposed to heat stress for 96 h, while the other two treatments showed lower DEGs values of 4659 DEGs after 1 h exposure and 3981 DEGs after 48 h exposure to heat stress. The DEGs were mainly synthesized in protein folding pathways under high-temperature stress after 1 h exposure. Moreover, a large number of genes encoding ROS scavenging enzymes were activated under heat stress for 1 h and 48 h treatments. The flavonoid synthesis pathway of pearl millet was enriched after heat stress for 96 h. This study analyzed the transcription dynamics under short to long-term heat stress to provide a theoretical basis for the heat resistance response of pearl millet.

Regulation of the mammalian maternal-to-embryonic transition by eukaryotic translation initiation factor 4E.

Eukaryotic translation initiation factor 4E (eIF4E) mediates cap-dependent translation. Genetic and inhibitor studies show that eIF4E expression is required for the successful transition from maternal to embryonic control of mouse embryo development. eIF4E was present in the oocyte and in the cytoplasm soon after fertilization and during each stage of early development. Functional knockout (Eif4e-/-) by PiggyBac [Act-RFP] transposition resulted in peri-implantation embryonic lethality because of the failure of normal epiblast formation. Maternal stores of eIF4E supported development up to the two- to four-cell stage, after which new expression occurred from both maternal and paternal inherited alleles. Inhibition of the maternally acquired stores of eIF4E (using the inhibitor 4EGI-1) resulted in a block at the two-cell stage. eIF4E activity was required for new protein synthesis in the two-cell embryo and Eif4e-/- embryos had lower translational activity compared with wild-type embryos. eIF4E-binding protein 1 (4E-BP1) is a hypophosphorylation-dependent negative regulator of eIF4E. mTOR activity was required for 4E-BP1 phosphorylation and inhibiting mTOR retarded embryo development. Thus, this study shows that eIF4E activity is regulated at key embryonic transitions in the mammalian embryo and is essential for the successful transition from maternal to embryonic control of development.

Machine learning-based microstructure prediction during laser sintering of alumina.

Predicting material's microstructure under new processing conditions is essential in advanced manufacturing and materials science. This is because the material's microstructure hugely influences the material's properties. We demonstrate an elegant machine learning algorithm that faithfully predicts the microstructure under new conditions, without the need of knowing the governing laws. We name this algorithm, RCWGAN-GP, which is regression-based conditional generative adversarial networks with Wasserstein loss function and gradient penalty. This algorithm was trained with experimental SEM micrographs from laser-sintered alumina under various laser powers. The RCWGAN-GP realistically regenerates the SEM micrographs under the trained laser powers. Impressively, it also faithfully predicts the alumina's microstructure under unexplored laser powers. The predicted microstructure features, including the morphology of the sintered particles and the pores, match the experimental SEM micrographs very well. We further quantitatively examined the prediction accuracy of the RCWGAN-GP. We trained the algorithm with computer-created micrograph datasets of secondary-phase growth governed by the well-known Johnson-Mehl-Avrami (JMA) equation. The RCWGAN-GP accurately regenerates the micrographs at the trained time series, in terms of the grains' shapes, sizes, and spatial distributions. More importantly, the predicted secondary phase fraction accurately follows the JMA curve.

Microwave-photonic low-coherence interferometry for dark zone free distributed optical fiber sensing.

A microwave-photonic low-coherence interferometry (MPLCI) system is proposed for fully distributed optical fiber sensing. Assisted by an unbalanced Michelson interferometer, a low-coherence laser source is used to interrogate cascaded Fabry-Perot interferometers along with an optical fiber for a dark zone free (or spatially continuous) distributed measurement. By combining the advantages of microwaves and photonics, the MPLCI system can synergistically achieve high sensitivity and high spatial resolution. Our tests have confirmed a strain resolution of 95 nε at the spatial resolution of 10 cm.

Disruption in ACTL7A causes acrosomal ultrastructural defects in human and mouse sperm as a novel male factor inducing early embryonic arrest.

Early embryonic arrest is a challenge for in vitro fertilization (IVF). No genetic factors were previously revealed in the sperm-derived arrest of embryonic development. Here, we reported two infertile brothers presenting normal in conventional semen analysis, but both couples had no embryos for transfer after several IVF and intracytoplasmic sperm injection (ICSI). Whole-exome sequencing identified a homozygous missense mutation of ACTL7A in both brothers. This mutation is deleterious and causes sperm acrosomal ultrastructural defects. The Actl7a knock-in mouse model was generated, and male mutated mice showed sperm acrosomal defects, which were completely consistent with the observations in patients. Furthermore, the sperm from ACTL7A/Actl7a-mutated men and mice showed reduced expression and abnormal localization of PLCζ as a potential cause of embryonic arrest and failure of fertilization. Artificial oocyte activation could successfully overcome the Actl7a-mutated sperm-derived infertility, which is meaningful in the future practice of IVF/ICSI for the ACTL7A-associated male infertility.

Laser-assisted embedding of all-glass optical fiber sensors into bulk ceramics for high-temperature applications.

We develop a laser-assisted sensor embedding process to embed all-glass optical fiber sensors into bulk ceramics for high-temperature applications. A specially designed two-step microchannel was fabricated on an Al2O3 substrate for sensor embedment using a picosecond (ps) laser. An optical fiber Intrinsic Fabry-Perot Interferometer (IFPI) sensor was embedded at the bottom of the microchannel and covered by Al2O3 slurry which was subsequently sintered by a CO2 laser. The sensor spectrum was in-situ monitored during the laser sintering process to ensure the survival of the sensor and optimize the laser sintering parameters. By testing in furnace through high temperature, the embedded optical fiber shows improved stability after CO2 laser sealing, resulting in the linear temperature response of the embedded optical fiber IFPI sensor. To improve the embedded IFPI sensor for thermal strain measurement, a dummy fiber was co-embedded with the sensing fiber to improve the mechanical bonding between the sensing fiber and the ceramic substrate so that the thermal strain of the ceramic substrate can apply on the sensing fiber. The response sensitivity, measurement repeatability and high-temperature long-term stability of the embedded optical fiber IFPI sensor were evaluated in this work.

Information integrated glass module fabricated by integrated additive and subtractive manufacturing.

In this Letter, we report a novel integrated additive and subtractive manufacturing (IASM) method to fabricate an information integrated glass module. After a certain number of glass layers are 3D printed and sintered by direct ${{\rm CO}_2}$CO2 laser irradiation, a microchannel will be fabricated on top of the printed glass by integrated picosecond laser, for intrinsic Fabry-Perot interferometer (IFPI) optical fiber sensor embedment. Then, the glass 3D printing process continues for the realization of bonding between optical fiber and printed glass. Temperature sensing up to 1000°C was demonstrated using the fabricated information integrated module. In addition, the long-term stability of the glass module at 1000°C was conducted. Enhanced sensor structure robustness and harsh temperature sensing capability make this glass module attractive for harsh environment structural health monitoring.

Redox-Responsive Polymeric RNAi Based on Multivalent Conjugation of siRNA for Improved Intracellular Delivery.

Learning from the design concept of antibody-drug conjugates (ADCs), we attempted to construct siRNA conjugated polymer brush by attaching a multiple of siRNA to the units of poly(amino acids) [poly(lysine) derivatives] through an intracellular cleavable disulfide bond. Note that the disulfide linkage is stable at extracellular milieu yet subjected to cleavage into free thiol residues at the intracellular reducing compartments. Consequently, ready release of arrays of active siRNA was achieved selectively in the intracellular compartments. Furthermore, tumor-targeted cyclic Asp-Gly-Arg (RGD) was conjugated to the aforementioned polymer brush in view that the RGD receptors (αVß3 and αVß5 integrins) were overexpressed over a wide spectrum of cancerous cells. Our subsequent results have achieved potent gene silencing in cultured cancerous cells from our proposed siRNA delivery construct. To our best knowledge, our proposed conjugate should be the first example of using an ADC platform in successful intracellular transportation of larger macromolecular biological payloads rather than small molecular chemotherapeutic drugs. Hence, the proposed strategy may serve as a promising avenue for targeted delivery of macromolecular pharmaceutical payloads.

Multiple facilitated glucose transporters SLC2As are required for normal mouse preimplantation embryo development.

BACKGROUND: Glucose metabolism is an essential energy source for mammalian preimplantation embryonic development. Therefore, we aimed to analyze the expression of all 12 known glucose transporters (facilitated solute carrier family 2, Slc2a) during early mouse embryo development. METHODS: Gene and protein expression of Slc2a transporters in oocytes and embryos were assessed by the TaqMan gene expression assay and confocal immunofluorescence, respectively. RESULTS: Except for Slc2a2, the other 11 Slc2a transcripts were detected in oocytes. Transcripts of Slc2a1, Slc2a3, Slc2a4, and Slc2a8 were the most enriched and detected in preimplantation embryos. The transcription of other Slc2a isoforms was barely detectable or absent after fertilization; however, they were detected in blastocysts, except for Slc2a10 and Slc2a13. Embryo culture in the simple defined medium caused a reduction in transcription of Slc2a1, Slc2a3, Slc2a4, and Slc2a8 in blastocyst; yet, amino acids partially reversed this impaired transcription of Slc2a1 and Slc2a4. SLC2A1 and SLC2A4 proteins were detected at all embryonic stages with nuclear accumulation in the embryos at the early cleavage stage. SLC2A3 and SLC2A8 were not detected in embryos until the eight-cell stage. The cellular membrane localization of SLC2A1, SLC2A3, and SLC2A8 occurred after compaction and was characterized in blastocysts. SLC2A4 was evenly distributed in the cytoplasm and nuclei without its characteristic membrane localization. Indinavir sulfate (a SLC2A4 inhibitor) decreased the rate of development and prevented glucose utilization in embryos after compaction. These inhibitory activities were partially reversed by exogenous insulin. CONCLUSION: The results unveil distinct expression patterns of individual Slc2a glucose transporters during early embryo development. Taken together, they provide novel insights into the understanding and management of glucose metabolic infertility in assisted-reproductive technologies (ART).

The ovarian estrogen synthesis function was impaired in Y123F mouse and partly restored by exogenous FSH supplement.

BACKGROUND: LepR tyrosine site mutation mice (Y123F) exhibit decreased serum E2 levels, immature reproductive organs, infertility as well as metabolic abnormalities. Although the actions of leptin and lepR in the control of reproductive function are thought to be exerted mainly via the hypothalamic-pituitary-gonadal axis, relatively less is known regarding their local effects on the peripheral ovary, especially on steroid hormone synthesis. Meanwhile, whether the decreased fertility of Y123F mouse could be restored by gonadotropin has not been clear yet. METHODS: The serum levels of E2, P4, FSH, LH, T and leptin of Y123F and WT mice at the age of 12 weeks were measured by enzyme-linked immunosorbent assay. Immunohistochemistry was used to compare the distribution of hormone synthases (STAR, CYP11A1, CYP19A1, HSD17B7) and FSHR in adult mouse ovaries of two genotypes. Western blot and real-time PCR were used to detect the expression levels of four ovarian hormone synthases and JAK2-STAT3 / STAT5 signaling pathway in 4 and 12 weeks old mice, as well as the effects of exogenous hFSH stimulation on hormone synthases and FSHR. RESULTS: Compared with WT mice, the serum levels of FSH, LH and E2 in 12-week-old Y123F mice were significantly decreased; T and leptin levels were significantly increased; but there was no significant difference of serum P4 levels. STAR, CYP11A1, HSD17B7 expression levels and the phosphorylation levels of JAK2 and STAT3 were significantly decreased in adult Y123F mice, while the expression of CYP19A1 and phospho-STAT5 were significantly increased. No significant differences were found between 4-week-old Y123F and WT mice. After exogenous hFSH stimulation, E2 levels and expression of CYP19A1 and HSD17B7 were significantly higher than that in the non-stimulated state, but significant differences still existed between Y123F and WT genotype mice under the same condition. CONCLUSIONS: Abnormal sex hormone levels of Y123F mice were due to not only decreased gonadotropin levels in the central nervous system, but also ovarian hormone synthase abnormalities in the peripheral gonads. Both FSH signaling pathway and JAK2-STAT3/STAT5 signaling pathway were involved in regulation of ovarian hormone synthases expression. Exogenous FSH just partly improved the blood E2 levels and ovarian hormone synthase expression.

Disruption of Ssp411 causes impaired sperm head formation and male sterility in mice.

BACKGROUND: We previously cloned the Ssp411 gene. We found that the Ssp411 protein is predominantly expressed in elongated spermatids in the rat testis in a stage-dependent manner. Although our findings strongly suggested that Ssp411 might play an important role in mammalian spermatogenesis, this hypothesis has not been studied. METHODS: We first used real-time PCR, Western blotting and immunohistochemistry to confirm that the expression pattern of Ssp411 in several murine tissues is similar to its expression pattern in corresponding rat tissues. To better understand the roles of Ssp411 in male reproduction in vivo, we identified and characterized an Ssp411 expression-disrupted murine strain (Ssp411PB/PB) that was generated by piggyBac (PB) transposon insertion. We studied Ssp411-interacting proteins using proteome microarray, co-IP and GST pull-down assay. RESULTS: Both Ssp411 mRNA and protein were detected exclusively in spermatids after step 9 during spermiogenesis in testis. Phenotypic analysis suggested that only Ssp411PB/PB males are sterile. These males have smaller testes, reduced sperm counts, decreased sperm motility and deformed spermatozoa. Microscopy analysis indicated that the manchette, a structurally reshaped sperm head, is aberrant in Ssp411PB/PB spermatids. The results of proteome microarray analysis and GST pull-down assays suggested that Ssp411 participates the ubiquitin-proteasome system by interacting with PSMC3. This has been reported to be manchette-associated and important for the head shaping of spermatids. CONCLUSIONS: Our study suggested that Ssp411 is required for spermiogenesis. It seems to play a role in sperm head shaping. The lack of Ssp411 causes sperm deformation and results in male infertility. GENERAL SIGNIFICANCE: Ssp411PB/PB mouse strain is an animal model of idiopathic oligoasthenoteratozoospermia (iOAT), and the gene may represent a therapeutic target for iOAT patients.

Reproductive effects of cadmium on sperm function and early embryonic development in vitro.

Cadmium is a major environmental toxicant that is released into the atmosphere, water and soil in the form of cadmium oxide, cadmium chloride, or cadmium sulfide via industrial activities, such as the manufacturing of batteries and pigments, metal smelting and refining and municipal waste incineration. In the present study, we investigated the effects of cadmium exposure on sperm quality parameters, fertilization capacity and early embryonic development. Our study showed that in vitro incubation of human or mouse sperms with cadmium for a long time (up to 24 hours) could significantly decreased sperm motility in a concentration- and time-dependent manner. Exposure to cadmium in the environment for a short term (30 min) did not affect sperm motility but significantly reduced in vitro fertilization rate. We also evaluated the effects of cadmium at concentrations of 0.625 µg/ml, and 1.25 µg/ml on early embryonic development in vitro and observed that the blastocyst formation rate dramatically decreased with increasing cadmium concentration. This finding emphasizes the hazardous effects of cadmium on sperm quality as well as on natural embryo development and raises greater concerns regarding cadmium pollution.