Flow virometry: recent advancements, best practices, and future frontiers

Journal of Virology, Год журнала: 2025, Номер unknown

Опубликована: Янв. 27, 2025

ABSTRACT The imperative for developing robust tools to detect, analyze, and characterize viruses has become increasingly evident as they continue threaten human health. In this review, we focus on recent advancements in studying with flow virometry (FV), an emerging technique that gained considerable momentum over the past 5 years. These include application of FV viral surface phenotyping, protein functionality, virus sorting, vaccine development, diagnostics. With examples illustrated using primary data from our studies, demonstrate is a powerful yet underutilized methodology that, when employed best practices experimental rigor, can be highly valuable individual virion heterogeneity, phenotypes, virus-antibody interactions. also address current challenges performing propose strategies overcome these obstacles, outline both new experienced researchers. Finally, discuss promising future prospects within broader context virology research.

Язык: Английский

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Reproductive physiology of the boar: What defines the potential fertility of an ejaculate?

Animal Reproduction Science, Год журнала: 2024, Номер 269, С. 107476 - 107476

Опубликована: Апрель 18, 2024

Despite decades of research and handling semen for use in artificial insemination (AI) other assisted reproductive technologies, 5-10% selected boar sires are still considered sub-fertile, escaping current assessment methods sperm quality resilience to preservation. As end-product, the ejaculate (emitted spermatozoa sequentially exposed composite seminal plasma, SP) ought define homeostasis testes, epididymis, accessory sexual glands. Yet, linking findings production biology fertility is suboptimal. The present essay critically reviews how a fertile can help us diagnose both health handling, focusing on -available under development- identify suitable biomarkers cryotolerance fertility. Bulk SP, proteins microRNAs (miRNAs) have, albeit linked function after AI, failed enhance outcomes at commercial level, perhaps just being components complex functional pathway. Hence, focus now interaction sperm-SP, comparing vivo with ex vivo, regarding nano-sized lipid bilayer extracellular vesicles (sEVs) as priority. sEVs transport fragile molecules (lipids, proteins, nucleic acids) which, shielded from degradation, mediate cell-to-cell communication female internal genital tract. Such modulates essential processes, immunological tolerance. be harvested, characterized, stored, manipulated, e.g. used andrological diagnosis, selection breeders, alternatively additives improve cryosurvival

Язык: Английский

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Small and Large Extracellular Vesicles of Porcine Seminal Plasma Differ in Lipid Profile

International Journal of Molecular Sciences, Год журнала: 2024, Номер 25(13), С. 7492 - 7492

Опубликована: Июль 8, 2024

Seminal plasma contains a heterogeneous population of extracellular vesicles (sEVs) that remains poorly characterized. This study aimed to characterize the lipidomic profile two subsets differently sized sEVs, small (S-) and large (L-), isolated from porcine seminal by size-exclusion chromatography characterized an orthogonal approach. High-performance liquid chromatography-high-resolution mass spectrometry was used for analysis. A total 157 lipid species 14 classes 4 major categories (sphingolipids, glycerophospholipids, glycerolipids, sterols) were identified. Qualitative differences limited cholesteryl ester present only in S-sEVs. L-sEVs had higher levels all quantified due their larger membrane surface area. The distribution pattern different, especially sphingomyelins (more S-sEVs) ceramides L-sEVs). In conclusion, this reveals suggesting they differ biogenesis functionality.

Язык: Английский

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Cryogenic electron microscopy reveals morphologically distinct subtypes of extracellular vesicles among porcine ejaculate fractions

Scientific Reports, Год журнала: 2024, Номер 14(1)

Опубликована: Июль 13, 2024

Seminal plasma (SP) is rich in extracellular vesicles (EVs), which are still poorly studied, especially livestock species. To better understand their functional role both spermatozoa and endometrial epithelial cells, proper characterization of EVs an essential step. The objective was to phenotypically characterize porcine seminal (sEVs) using cryogenic electron microscopy (cryo-EM), allows visualization native state. Porcine ejaculates released fractions, each containing SP from different source. This sEVs various male reproductive tissues. Two experiments were performed, the first with entire ejaculate (n:6) second three fractions (n:15): 10 mL sperm-rich fraction (SRF-P1) mainly epididymis, remainder SRF (SRF-P2) prostate, post-SRF vesicles. isolated by size exclusion chromatography 1840 cryo-EM sEV images acquired a Jeol-JEM-2200FS/CR-EM. size, density, complexity, peripheral corona layer measured ImageJ software. experiment showed that structurally morphologically heterogeneous, although most (83.1%) small (less than 200 nm), rounded, electrodense, some have coronal layer. There also larger (16.9%) irregularly shaped, more few common SRF-P1 SRF-P2, indicating they originated epididymis prostate. Large abundant post-SRF, heterogeneous. would be explained diversity organs origin.

Язык: Английский

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Multi-Omics Sequencing Dissects the Atlas of Seminal Plasma Exosomes from Semen Containing Low or High Rates of Sperm with Cytoplasmic Droplets

International Journal of Molecular Sciences, Год журнала: 2025, Номер 26(3), С. 1096 - 1096

Опубликована: Янв. 27, 2025

Sperm cytoplasmic droplets (CDs) are remnants of cytoplasm that can cause a number problems if it not shed from the sperm after ejaculation. Exosomes rapidly bind to sperm, but is clear whether exosomes affect migration and shedding CDs. We first extracted characterized seminal plasma boar semen containing with low or high rates Then, transcriptomic proteomic detection these were performed analyze differences between two groups exosomes. The results revealed 486 differentially expressed genes (DEGs), 40 proteins (DEPs), 503 lncRNAs (DElncRNAs) identified CD rate group group. Integrative multi-omics analysis showed exosome components may by influencing cytoskeletal regulation insulin signaling, including actin cytoskeleton, ECM–receptor interaction, axon guidance, secretion, signaling pathway. Overall, our study systematically DEGs, DEPs, DElncRNAs in semen, which will help broaden understanding complex molecular mechanisms involved

Язык: Английский

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Isolation and Characterization of Extracellular Vesicle Subsets in Donkey Seminal Plasma

Опубликована: Янв. 1, 2025

Язык: Английский

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Protective role of extracellular vesicles against oxidative DNA damage

Biological Research, Год журнала: 2025, Номер 58(1)

Опубликована: Март 13, 2025

Abstract Background Oxidative stress, a source of genotoxic damage, is often the underlying mechanism in many functional cell disorders. Extracellular vesicles (EVs) have been shown to be key regulators cellular processes and may involved maintaining redox balance. Herein, we aimed develop method assess effects EVs on DNA oxidation using porcine seminal plasma extracellular (sEVs). Results The technique was set cell-free plasmid avoid bias generated by uptake sEVs sperm cells, employing increasing concentrations hydrogen peroxide (H 2 O ) that generate single-strand breaks (SSBs). Because SSBs contain free 3’-OH end allow extension through quantitative PCR, such -and therefore SYBR intensity- showed proportional amount SSB. In next stage, H co-incubated with two size-differentiated subpopulations (small large) permeabilized non-permeabilized whether intravesicular content (IC) or surface protects from oxidative damage. obtained small reduced incidence ( P < 0.05), whereas large had no impact generation > 0.05). IC protective effect against Conclusions Our results suggest sEVs, including peripheral corona layer, exert function alterations are originated mechanisms. addition, our vitro study opens path detect, localize quantify derived different fluids, elucidating their physiopathological states.

Язык: Английский

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Milk Extracellular Vesicles: Natural Nanoparticles for Enhancing Oral Drug Delivery against Bacterial Infections

ACS Biomaterials Science & Engineering, Год журнала: 2024, Номер 10(4), С. 1988 - 2000

Опубликована: Март 26, 2024

Oral drug delivery is typically preferred as a therapeutic intervention due to the complexities and expenses associated with intravenous administration. However, some drugs are poorly absorbed orally, requiring administration bypass gastrointestinal tract deliver directly into bloodstream. Thus, there an urgent need develop novel platforms overcome challenges of oral low solubility, permeability, degradation, bioavailability. Advances in extracellular vesicles (EVs) natural carriers have provided emerging approaches improve potential applications. Milk not only contains traditional nutrients but also rich EVs. In this Review, we focus mainly on purification milk EVs (mEVs), their safety, advantages mEV-based combatting intestinal infections. Additionally, summarize several mEVs over conventional synthetic carriers, such immunogenicity, high biocompatibility, ability transfer bioactive molecules between cells. Considering unmet gaps clinical translation, it essential review cargo loading future perspectives for use delivery. This overview sheds light alternative treat infections pathogens development systems.

Язык: Английский

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Sperm functionality is differentially regulated by porcine oviductal extracellular vesicles from the distinct phases of the estrous cycle

Reproduction Fertility and Development, Год журнала: 2024, Номер 36(8)

Опубликована: Май 7, 2024

Context Extracellular vesicles (EVs) derived from the oviductal fluid (oEVs) play a critical role in various reproductive processes, including sperm capacitation, fertilisation, and early embryo development. Aims To characterise porcine oEVs (poEVs) different stages of estrous cycle (late follicular, LF; luteal, EL; mid ML; late LL) investigate their impact on functionality. Methods poEVs were isolated, characterised, labelled to assess binding boar spermatozoa. The effects motility, viability, acrosomal status, protein kinase A phosphorylation (pPKAs), tyrosine (Tyr-P), vitro fertility analysed. Key results observed as round or cup-shaped membrane-surrounded vesicles. Statistical analysis showed that did not significantly differ size, quantity, concentration among phases cycle. However, LF demonstrated higher affinity for sperm. Treatment with EL, ML, LL resulted decrease progressive motility total motility. Moreover, pPKA levels reduced presence LF, ML poEVs, while Tyr-P between groups. also penetration rate number spermatozoa per penetrated oocyte (P < 0.05). Conclusions modulatory functionality by interacting spermatozoa, affecting participating sperm–oocyte interaction. Implications differential suggest potential use additives IVF systems regulate

Язык: Английский

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Updating Research on Extracellular Vesicles of the Male Reproductive Tract in Farm Animals: A Systematic Review

Animals, Год журнала: 2024, Номер 14(21), С. 3135 - 3135

Опубликована: Окт. 31, 2024

This systematic review examined research studies on extracellular vesicles (EVs) of the male reproductive tract in livestock species to summarize topics and methodologies used, key findings, future directions. PubMed Scopus were searched for time ranges up 1 September 2024, 1383 articles identified. The application screening eligibility criteria resulted selection 79 focusing EVs livestock. Porcine bovine most studied. A variety EV isolation techniques with ultracentrifugation being common. Characterization was a weak point, only 24.05% characterizing according MISEV guidelines. Inadequate characterization compromises reliability results. results 19 that provided good showed from are phenotypically compositionally heterogeneous. These papers also these would be involved regulation sperm functionality. Research remains scarce, further is needed, which should include appropriate aim find efficient methods isolate them assess their involvement functionality spermatozoa cells female genital tract.

Язык: Английский

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