The role of reproductive tract extracellular vesicles on boar sperm function.

Extracellular vesicles (EVs) are abundant in reproductive tract fluids and serve as important mediators of paracrine communication, influencing the function of gametes. Sperm undergo development in the male reproductive tract and exert their function within the female reproductive tract, engaging in interactions with various types of EVs present throughout the reproductive system. Previous studies have demonstrated that both male and female reproductive tract EVs can impact sperm function by transferring regulatory cargoes to them. Nevertheless, inconsistencies of previous research regarding the effects of EVs on sperm function, coupled with a lack of investigation into the influence of female reproductive tract EVs on sperm fertilization, have left the true role and underlying mechanisms of reproductive tract EVs on sperm function largely unexplored. Given that pigs represent significant economic livestock and serve as an ideal biomedical model for human diseases, this review aims to provide a comprehensive summary of the current knowledge regarding reproductive tract EVs and their influence on boar sperm function, while highlighting their potential roles. We anticipate that this review will facilitate future research on reproductive tract EVs and their impact on sperm function, contributing to improved animal reproductive efficiency and advancements in the treatment of male infertility.

Can policy advantages be transformed into industrial advantages? Evidences from digital economy industry development in Zhejiang Province, China.

The digital economy is a new impetus to promote high-quality economic development. We use the policies of Zhejiang Information Economy Development Demonstration Base (IEDD) and Zhejiang Software and Information Service Industry Base (SISI) established between 2015 and 2017 to design a quasi-natural experiment. By using a panel data from 2005 to 2020 in Zhejiang and the difference-in-differences model, we test the impacts of IEDD and SISI policies on digital economy development. We find that there are significant spatial differences for digital economy in Zhejiang. IEDD and SISI policies improve the digital economy development, that is, the policy advantages can indeed be transformed into industrial advantages. The IEDD policy can promote the digital economy industry development by enhancing the digital infrastructure and financial development; SISI policy can promote the development of the digital economy industry by promoting financial development. The results of quantile regression show that the promotion effect of IEDD and SISI policies increases with the improvement of the industrial basis of regional digital economy. The results of group regression show that the IEDD policy promotes the digital economy development in counties and county-level cities of Zhejiang, and the SISI policy plays a significant role in municipal districts.

Glycyrrhiza polysaccharides inhibits PRRSV replication.

Glycyrrhiza polysaccharide (GCP) is a natural plant active polysaccharide extracted from traditional Chinese medicine licorice. In this research, we studied the antiviral activity of glycyrrhiza polysaccharide against porcine reproductive and respiratory syndrome virus (PRRSV), a virus of the Arteriviridae family, with a high rate of variation and has caused huge economic losses to the pig industry in various countries since its discovery. Our results show that GCP can inhibit PRRSV replication in a dose-dependent manner. Furthermore, GCP could inhibit the mRNA expression of receptor genes CD163 and NF-κB p65 and promote the mRNA expression of the SLA-7 gene. Because of these results, GCP can be used as a candidate drug to prevent and treat PRRS.

Synthesis and characterization of low surface energy thermoplastic polyurethane elastomers based on polydimethylsiloxane.

Organosilicon modified polyurethane elastomers (Si-MTPUs) were synthesized in order to improve the anti-graffiti property of thermoplastic polyurethane elastomers (TPUs). Si-MTPUs were prepared from polydimethylsiloxane (PDMS) and polytetramethylene glycol (PTMG) as mixed soft segment, 1,4-butanediol (BDO) and imidazole salt ionic liquid N-glyceryl-N-methyl imidazolium chloride ([MIMl,g]Cl) used as chain extender, and 4,4'-dicyclohexylmethane diisocyanate (HMDI). The structure, thermal stability, mechanical properties and physical crosslinking density of Si-MTPUs were characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetry analysis (TGA), mechanical test and low field nuclear magnetic resonance. Surface energy and water absorption were characterized by static contact angle test and water resistance test, and anti-graffiti and self-cleaning properties were characterized with water, milk, ink, lipstick, oily markers and spray paint. It was found that the mechanical properties of Si-MTPU-10 with the content of PDMS 10 wt% were optimized, with a maximum tensile strength of 32.3 MPa and elongation at break of 656%. Surface energy reached the minimum value of 23.1 mN m-1 with the best anti-graffiti performance, which no longer decreased with the increase of PDMS contents. This work provides novel idea and strategy for the preparation of low surface energy TPUs.

Uterine luminal-derived extracellular vesicles: potential nanomaterials to improve embryo implantation.

Most pregnancy losses worldwide are caused by implantation failure for which there is a lack of effective therapeutics. Extracellular vesicles are considered potential endogenous nanomedicines because of their unique biological functions. However, the limited supply of ULF-EVs prevents their development and application in infertility diseases such as implantation failure. In this study, pigs were used as a human biomedical model, and ULF-EVs were isolated from the uterine luminal. We comprehensively characterized the proteins enriched in ULF-EVs and revealed their biological functions in promoting embryo implantation. By exogenously supplying ULF-EVs, we demonstrated that ULF-EVs improve embryo implantation, suggesting that ULF-EVs are a potential nanomaterial to treat implantation failure. Furthermore, we identified that MEP1B is important in improving embryo implantation by promoting trophoblast cell proliferation and migration. These results indicated that ULF-EVs can be a potential nanomaterial to improve embryo implantation.

Role of genetic factors in different swine breeds exhibiting varying levels of resistance/susceptibility to PRRSV.

Porcine reproductive and respiratory syndrome (PRRS), caused by the PRRS virus (PRRSV), is an economically significant contagious disease. Traditional approaches based on vaccines or medicines were challenging to control PRRSV due to the diversity of viruses. Different breeds of pigs infected with PRRSV have been reported to have different immune responses. However, due to the complexity of interaction mechanism between host and PRRSV, the genetic mechanism leading to PRRSV susceptibility/resistance in various pig breeds is still unclear. Herein, the role of host genetic components in PRRSV susceptibility is systematically described, and the molecular mechanisms by which host genetic factors such as SNPs, cytokines, receptor molecules, intestinal flora, and non-coding RNAs regulate PRRSV susceptibility/resistance. Therefore, improving the resistance to disease of individual animals through disease-resistance breeding technology is of profound significance for uplifting the sustainable and healthy development of the pig industry.

Extracellular vesicle-encapsulated miR-21-5p in seminal plasma prevents sperm capacitation via Vinculin inhibition.

To penetrate the zona pellucida before sperm-egg binding, sperm must undergo highly time-controlled capacitation and acrosome reaction in the female reproductive tract. Our previous study demonstrated that miR-21-5p is the most abundant miRNA in boar seminal plasma (SP)-derived extracellular vesicles (EVs) and can target Vinculin (VCL) gene, which may participate in boar sperm capacitation. Thus, this study aims to explore the potential role of miR-21-5p from SP-derived EVs in preventing sperm capacitation and its underlying mechanism. We observed that sperm could incorporate miR-21-5p from SP-derived EVs. The roles of SP-derived EVs miR-21-5p in sperm capacitation were then determined using gain- and loss-of-function analyses. In addition, the expression levels of miR-21-5p, VCL, and VCL protein in liquid-preserved boar sperm following transfection were determined using RT-qPCR and Western blotting. Our results revealed that miR-21-5p overexpression inhibited sperm capacitation and acrosome reaction. Similarly, miR-21-5p expression was significantly lower (P < 0.05) in capacitated sperm than un-capacitated sperm. However, the protein level of VCL was also significantly lower (P < 0.05) in capacitated sperm than un-capacitated sperm. Furthermore, immunofluorescence analysis showed that VCL protein mainly located in sperm head and sperm capacitation was inhibited after treating with VCL protein inhibitor (Chrysin). In conclusion, our study provides reasonable evidence that miR-21-5p expression in SP-derived EVs could prevent sperm capacitation via VCL inhibition.

LncRNA-mediated effects of vitrification temperatures and cryoprotectant concentrations on bovine oocyte development following vitrification at the GV stage.

We evaluated the effects of different vitrification temperatures (VTs) and cryoprotective agent concentrations (CPAs) on the viability and expressions of long non-coding RNA (lncRNA) in bovine oocytes following vitrification at the germinal vesicle (GV) stage. Our findings provide a theoretical support for improvement of the cryopreservation technology of bovine immature oocytes (BIOs). Bovine cumulus oocyte complexes (COCs) were collected and randomized into five groups: fresh oocytes (control), oocytes vitrified in liquid helium (LHe; -269 °C) with 5.6 M CPAs (LHe 5.6 M), oocytes vitrified in LHe with 6.6 M CPAs (LHe 6.6 M), oocytes vitrified in liquid nitrogen (LN; -196 °C) with 5.6 M CPAs (LN 5.6 M), and oocytes vitrified in LN with 6.6 M CPAs (LN 6.6 M). Of the four vitrification groups, the LHe 5.6 M group exhibited the highest blastocyst rate (13.22%), followed by the LHe 6.6 M group (10.19%) and LN 6.6 M group (9.77%), while the LN 5.6 M group had the lowest blastocyst rate (1.87%). Then, lncRNA expressions in the five groups were profiled. A total of 18,271 lncRNAs were identified, of which 2,158 were differentially expressed lncRNAs (DELs) in the vitrified groups, compared to the fresh group (P < 0.05; fold-change > 2). Co-location (cis) and co-expression (trans) prediction revealed 14 differentially expressed target genes (DETGs), which corresponded to 17 DELs. Based on grouping data and expression profiles of the DELs, we demonstrated that different VTs (-269 °C vs. -196 °C) can affect the expressions of MSTRG.12295.5, MSTRG.37123.1, MSTRG.37930.2, MSTRG.40464.9, MSTRG.8869.3 and MSTRG.26680.6. Expressions of these lncRNAs were affected by CPAs only in the condition of vitrification with LHe (-269 °C). Expressions of MSTRG.35129.6 were associated with exposures to both VTs and CPAs; while expressions of MSTRG.3578.3, MSTRG.40576.3, MSTRG.6723.5, MSTRG.32862.4, MSTRG.1184.4, MSTRG.33110.3, MSTRG.40454.2, MSTRG.41073.2, MSTRG.44732.4 and MSTRG.6729.3 might be related to vitrification. Co-expression analysis showed that MSTRG.12295.5, MSTRG.37930.2, MSTRG.40454.2, MSTRG.8869.3 and MSTRG.6723.5 expressions affect oocyte development after vitrification by regulating target gene expressions. Taken together, improvement of the developmental ability of BIOs after LHe vitrification maybe attributed to changes in expressions of some lncRNAs. Our findings elucidate on the molecular mechanisms underlying the development of BIOs under different VTs and CPAs.

Corrigendum: Global Transcriptomic Analyses Reveal Genes Involved in Conceptus Development During the Implantation Stages in Pigs.

[This corrects the article DOI: 10.3389/fgene.2021.584995.].

Integrated Insight into the Molecular Mechanisms of Spontaneous Abortion during Early Pregnancy in Pigs.

Due to the high rate of spontaneous abortion (SAB) in porcine pregnancy, there is a major interest and concern on commercial pig farming worldwide. Whereas the perturbed immune response at the maternal-fetal interface is an important mechanism associated with the spontaneous embryo loss in the early stages of implantation in porcine, data on the specific regulatory mechanism of the SAB at the end stage of the implantation remains scant. Therefore, we used high-throughput sequencing and bioinformatics tools to analyze the healthy and arresting endometrium on day 28 of pregnancy. We identified 639 differentially expressed lncRNAs (DELs) and 2357 differentially expressed genes (DEGs) at the end stage of implantation, and qRT-PCR was used to verify the sequencing data. Gene set variation analysis (GSVA), gene set enrichment analysis (GSEA), and immunohistochemistry analysis demonstrated weaker immune response activities in the arresting endometrium compared to the healthy one. Using the lasso regression analysis, we screened the DELs and constructed an immunological competitive endogenous RNA (ceRNA) network related to SAB, including 4 lncRNAs, 11 miRNAs, and 13 genes. In addition, Blast analysis showed the applicability of the constructed ceRNA network in different species, and subsequently determined HOXA-AS2 in pigs. Our study, for the first time, demonstrated that the SAB events at the end stages of implantation is associated with the regulation of immunobiological processes, and a specific molecular regulatory network was obtained. These novel findings may provide new insight into the possibility of increasing the litter size of sows, making pig breeding better and thus improving the efficiency of animal husbandry production.

SIMULATION AND MEASUREMENT OF EXTERNAL ELECTROMAGNETIC ENVIRONMENT OF TOKAMAK DEVICE.

Magnetic confinement nuclear fusion is an important way to realize controllable nuclear fusion. Due to the large current and complicated coil arrangement, there is a complicated electromagnetic environment around the fusion device. In this paper, the B-dot sensor is used to measure the magnetic field, the D-dot sensor is used to measure the electric field, the MAXWELL electromagnetic simulation software is used to simulate the electromagnetic field strength; the simulation and measurement of the spherical Tokamak SUNIST device and the measurement of MARX generator are carried out, then we give corresponding electromagnetic protection suggestions.

Catalytic amplification based on hierarchical heterogeneity bimetal-organic nanostructures with peroxidase-like activity.

In this paper, integrating heterometallic units and nanostructures into metal-organic frameworks (MOFs) were applied to improve the sensitivity of detecting hydrogen peroxide (H2O2) in neutral solution. The bimetal-MOFs (CuCo-BDC) and GO composite (CuCo-BDC/GO) were first synthesized via an ordinary one-step solvothermal synthesis. The CuCo-BDC/GO with admirable peroxidase-like catalytic activity could be applied to detect H2O2. The results have low detection limit of 69 nM (S/N = 3) and a wide linear detection range, from 100 nM to 3.5 mM. This is superior to recently published biosensors based on noble metal nanomaterials, which confirms CuCo-BDC/GO as the MOF electrocatalysts with high performance. The remarkable electroanalytical performance of CuCo-BDC/GO is due to the presence of numerous open metal active sites, the synergistic effect of Cu2+ and Co2+, hierarchical structure with high-specific surface areas and the marvelous electrochemical properties of GO. Therefore, CuCo-BDC/GO is a powerful candidate for detecting H2O2 in electrochemical biosensing fields. Moreover, H2O2 detection in real samples can be done with the CuCo-BDC/GO, including human serum samples. Therefore, the novel CuCo-BDC/GO is a promising catalyst that can be applied in biotechnological and environmental applications.

Global Transcriptomic Analyses Reveal Genes Involved in Conceptus Development During the Implantation Stages in Pigs.

Prenatal mortality remains a significant concern to the pig farming industry around the world. Spontaneous fetal loss ranging from 20 to 45% by term occur after fertilization, with most of the loss happening during the implantation period. Since the factors regulating the high mortality rates of early conceptus during implantation phases are poorly understood, we sought to analyze the overall gene expression changes during this period, and identify the molecular mechanisms involved in conceptus development. This work employed Illumina's next-generation sequencing (RNA-Seq) and quantitative real-time PCR to analyze differentially expressed genes (DEGs). Soft clustering was subsequently used for the cluster analysis of gene expression. We identified 8236 DEGs in porcine conceptus at day 9, 12, and 15 of pregnancy. Annotation analysis of these genes revealed rRNA processing (GO:0006364), cell adhesion (GO:1904874), and heart development (GO:0007507), as the most significantly enriched biological processes at day 9, 12, and 15 of pregnancy, respectively. In addition, we found various genes, such as T-complex 1, RuvB-like AAA ATPase 2, connective tissue growth factor, integrins, interferon gamma, SLA-1, chemokine ligand 9, PAG-2, transforming growth factor beta receptor 1, and Annexin A2, that play essential roles in conceptus morphological development and implantation in pigs. Furthermore, we investigated the function of PAG-2 in vitro and found that PAG-2 can inhibit trophoblast cell proliferation and migration. Our analysis provides a valuable resource for understanding the mechanisms of conceptus development and implantation in pigs.

Expression Pattern of Seminal Plasma Extracellular Vesicle Small RNAs in Boar Semen.

Extracellular vesicles (EVs) regulate multiple physiological processes. Seminal plasma contains numerous EVs that may deliver functional molecules such as small RNAs (sRNAs) to the sperm. However, the RNA profiles in the boar seminal plasma extracellular vesicles (SP-EVs) and its function have not been characterized. The aim of this study was to characterize the functions and sRNA profiles in the boar SP-EVs using deep sequencing technology. Briefly, boar SP-EVs were isolated by differential ultracentrifugation and confirmed with a transmission electron microscope (TEM), nanoparticle tracking analysis (NTA), and Western blot. The isolated boar SP-EVs contained numerous and diverse sRNA families, including microRNAs (miRNAs, 9.45% of the total reads), PIWI-interacting RNAs (piRNAs, 15.25% of the total reads), messenger RNA fragments (mRNA, 25.30% of the total reads), and tRNA-derived small RNAs (tsRNA, 0.01% of the total reads). A total of 288 known miRNAs, 37 novel miRNA, and 19,749 piRNAs were identified in boar SP-EVs. The identified ssc-miR-21-5p may confer negative effects on sperm fertility based on a dual-luciferase reporter experiment. This study therefore provides an effective method to isolate SP-EVs and characterizes the sRNA profile.

Density Functional Theory-Assisted Electrochemical Assay Manipulated by a Donor-Acceptor Structure toward Pharmaceutical Diagnostic.

Oxidative stress is a state of stress injury, which leads to the pathogenesis of most neurodegenerative diseases. Moreover, this is also one of the main reasons for the loss of dopaminergic neurons and the abnormal content of dopamine (DA). In the past decades, a number of studies have found that acetaminophen (AP) is metabolized and distributed in the brain when it is used as a neuroprotective compound. In this context, we proposed an electrochemical sensor based on 9-(4-(10-phenylanthracen-9-yl)phenyl)-9H-carbazole with the goal of diagnosing these two drugs in the body. Carbazole groups can easily be formed into large π-conjugated systems by electropolymerization. The introduction of anthracene exactly combined the carbazole group to establish an efficient electron donor-acceptor pattern, which enhanced π-π interaction with the electrode surface and charge transporting ability. The diagnostic platform showed good sensing activity toward the oxidation of DA and AP. The detection range for DA and AP is from 0.2 to 300 µM and from 0.2 to 400 µM, respectively. The simultaneous detection range is from 0.5 to 250 µM, which is wider than most reports. After a series of electrochemical assessments were determined, the sensor was finally developed to the analysis of pharmaceutical and human serum, displaying a meaningful potential in clinical evaluation.

Sex Manipulation Technologies Progress in Livestock: A Review.

Sex manipulation technologies allow predetermination of the sex of animal offspring by altering the normal reproductive process. In livestock production, the difference in type and gender can translate into significant economic benefits, including alleviation of severe food shortages. In livestock, however, the commercial application of sex manipulation technologies is currently available for cattle only. In this review, we described the brief history of sex manipulation, and the research progresses of common methods used in sex manipulation thus far. Information presented in this review can inform future studies on expanding the scope and use of sex manipulation technologies in livestock.

Deep-Sequencing Identification of MicroRNA Biomarkers in Serum Exosomes for Early Pig Pregnancy.

Early pregnancy diagnosis in sows can significantly improve the efficiency of pig industry. Exosomes are membrane-covered nanovesicles that can transport microRNAs (miRNAs) and other molecular signals between cells. In other species, serum exosome-derived miRNAs can serve as good biomarkers of diseases and different physiological states, including pregnancy status. We hypothesized that circulating exosome-derived miRNAs might be used to differentiate the pregnancy status as early as several days after insemination in pigs. To test this hypothesis, we randomly assigned pigs for artificial insemination with fertile or dead semen (control group). Serum samples were obtained from pregnant pigs on days 9, 12, and 15 after insemination and from non-pregnant pigs on days 0, 9, 12, and 15 after insemination. Exosomes were isolated for RNA extraction. The exosomal RNA samples from pigs on day 9 of the estrus cycle and pregnancy were used for small-RNA sequencing. A total 321 miRNAs were identified in all samples. Twenty eight differentially abundant miRNAs were identified between the pregnant and control groups. miRNAs with | log2 (fold change)| > 2 from sequencing results were selected for validation by quantitative reverse-transcription-polymerase chain reaction (RT-qPCR) in larger samples. Finally two upregulated miRNAs (miR-92b-3p and miR-17-5p) in the pregnant groups (on days 9, 12, and 15 of pregnancy) were confirmed by RT-qPCR. In summary, we have successfully identified circulating exosomal miRNA profiles in the serum of pigs in early pregnancy. miR-92b-3p and miR-17-5p could be used as potential circulating biomarkers for early pregnancy diagnosis.

Efficient deletion of LoxP-flanked selectable marker genes from the genome of transgenic pigs by an engineered Cre recombinase.

Genetically modified (GM) pigs hold great promises for pig genetic improvement, human health and life science. When GM pigs are produced, selectable marker genes (SMGs) are usually introduced into their genomes for host cell or animal recognition. However, the SMGs that remain in GM pigs might have multiple side effects. To avoid the possible side effects caused by the SMGs, they should be removed from the genome of GM pigs before their commercialization. The Cre recombinase is commonly used to delete the LoxP sites-flanked SMGs from the genome of GM animals. Although SMG-free GM pigs have been generated by Cre-mediated recombination, more efficient and cost-effective approaches are essential for the commercialization of SMG-free GM pigs. In this article we describe the production of a recombinant Cre protein containing a cell-penetrating and a nuclear localization signal peptide in one construct. This engineered Cre enzyme can efficiently excise the LoxP-flanked SMGs in cultured fibroblasts isolated from a transgenic pig, which then can be used as nuclear donor cells to generate live SMG-free GM pigs harboring a desired transgene by somatic cell nuclear transfer. This study describes an efficient and far-less costly method for production of SMG-free GM pigs.

An electrochemical thrombin aptasensor based on the use of graphite-like C3N4 modified with silver nanoparticles.

An electrochemical aptasensor for thrombin is introduced that makes use of a nanohybrid composed of silver nanoparticles and graphite-like carbon nitride (Ag-g-C3N4). The material has a large surface and good biocompatibility. AgNPs are modified directly on the surface of g-C3N4 via chemical reduction. A glass carbon electrode (GCE) modified with Ag-g-C3N4 can immobilize a large number of amino-terminated thrombin binding aptamers (NH2-TBA) through strong Ag-N bonds. The electrochemical impedance signal of the aptasensor increases in the presence of thrombin. Under the optimal conditions and by using [Fe(CN)6]3-/4- as an electrochemical probe, the aptasensor shows a wide linear range of 100 fM - 20 nM with a lower detection limit of 38 fM. The method was applied to the determination of thrombin in spiked human plasma and the recoveries fluctuated from 97.2% to 103%. Graphical abstractSchematic representation of an electrochemical aptasensor using graphite-like carbon nitride (C3N4) modified with silver nanoparticles as electrode substrate for thrombin (TB) detection.

The synergistic effect of Au-COF nanosheets and artificial peroxidase Au@ZIF-8(NiPd) rhombic dodecahedra for signal amplification for biomarker detection.

Here, a new type of signal amplification strategy is proposed employing Au nanoparticle (AuNP)-functionalized covalent organic framework (Au-COF) nanosheets and AuNP functionalized ZIF-8(NiPd) (Au@ZIF-8(NiPd)) rhombic dodecahedra nanocomposites for sandwich electrochemical sensor construction. The peroxidase mimics Au@ZIF-8(NiPd) took the place of natural enzymes in enzyme-assisted amplification strategies, both acting as catalysts for H2O2 reduction for signal amplification, and serving as ideal nanocarriers for signal probe anchoring. The cancer biomarker thrombin (TB) was selected as the target. Thrombin binding aptamers (TBA 2) were fixed on Au@ZIF-8(NiPd), and the obtained TBA 2-Au@ZIF-8(NiPd) bioconjugates were employed as tracer labels, and TB was sandwiched between the tracer labels and capture probe TBA 1 which were immobilized on the Au-COF nanosheet modified electrode. Au-COFs with a high specific area, super electroconductivity, and uniformly distributed AuNPs were utilized as the electrode substrate to fix TBA 1. Exploiting the sandwich method, the proposed TB aptasensor exhibited a wide linear range of 0.1 pM to 20 nM with a low detection limit of 15 fM (S/N = 3). The ingenious sensing strategy enriched the application diversity of the artificial enzyme and showed promise in research and development of point-of-care diagnostics.