Cells across biological kingdoms release extracellular vesicles (EVs) as a means of communication with other cells, be their friends or foes. This is indeed true for the intracellular malaria parasite Plasmodium falciparum (Pf), which utilizes EVs to transport bioactive molecules to various human host systems. Yet, the study of this mode of communication in malaria research is currently constrained due to limitations in high-resolution tools and the absence of commercial antibodies. Here, we demonstrate the power of an advanced spectral flow cytometry approach to robustly detect secreted EVs, isolated from Pf-infected red blood cells. By labeling both EV membrane lipids and the DNA cargo within (non-antibody staining approach), we were able to detect a subpopulation of parasitic-derived EVs enriched in DNA. Furthermore, we could quantitatively measure the DNA-carrying EVs isolated from two distinct blood stages of the parasite: rings and trophozoites. Our findings showcase the potential of spectral flow cytometry to monitor dynamic changes in nucleic acid cargo within pathogenic EVs.

• Klíčová slova
• DNA, cargo, extracellular vesicles, flow cytometry, host pathogen, malaria, parasite,
• MeSH
• erytrocyty * parazitologie   metabolismus   MeSH
• extracelulární vezikuly * metabolismus   MeSH
• lidé MeSH
• Plasmodium falciparum * metabolismus   genetika   MeSH
• protozoální DNA * metabolismus   MeSH
• průtoková cytometrie * metody   MeSH
• tropická malárie * parazitologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• protozoální DNA * MeSH

There is growing interest in the role of extracellular vesicles (EVs) in neonatal pathology. This study aimed to characterise circulating EVs following preterm birth. This single-centre prospective observational study included cord and postnatal plasma from preterm (n = 101) and full-term infants (n = 66). EVs were analysed using nanoparticle tracking analysis, flow cytometry, proteomics and procoagulant activity assay. We found changes in the concentration, size, cellular origin and proteomic content of circulating EVs in preterm infants during perinatal adaptation. To understand if these changes were related to prematurity or normal adaptation to extrauterine life, they were also investigated in term infants. There was a dramatic increase in the concentration of small and large EVs on Day 3 in the preterm group; specific subsets of platelet (CD42b+ and CD62P+), endothelial (VEGFR2) and tissue factor EVs were elevated. Differentially expressed proteins relating to haemostasis, pulmonary physiology and immunity were identified between Day 1 and 3 in preterm infants. These changes have never previously been described in a large cohort of preterm infants and differ from healthy term infants. These findings have major implications for future neonatal EV studies, particularly the timing of sample collection. Further work is required to understand the clinical implications of this unique EV profile following preterm birth.

• Klíčová slova
• haemorrhage, neonatal, umbilical cord, vascular biology,
• MeSH
• extracelulární vezikuly * metabolismus   MeSH
• fyziologická adaptace * MeSH
• lidé MeSH
• novorozenec nedonošený * krev   MeSH
• novorozenec MeSH
• prospektivní studie MeSH
• proteomika metody   MeSH
• těhotenství MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• novorozenec MeSH
• těhotenství MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• pozorovací studie MeSH

Extracellular vesicles (EVs) are emerging as critical mediators of intercellular communication in the tumor microenvironment (TME), profoundly influencing cancer progression. These nano-sized vesicles, released by both tumor and stromal cells, carry a diverse cargo of proteins, nucleic acids, and lipids, reflecting the dynamic cellular landscape and mediating intricate interactions between cells. This review provides a comprehensive overview of the biogenesis, composition, and functional roles of EVs in cancer, highlighting their significance in both basic research and clinical applications. We discuss how cancer cells manipulate EV biogenesis pathways to produce vesicles enriched with pro-tumorigenic molecules, explore the specific contributions of EVs to key hallmarks of cancer, such as angiogenesis, metastasis, and immune evasion, emphasizing their role in shaping TME and driving therapeutic resistance. Concurrently, we submit recent knowledge on how the cargo of EVs can serve as a valuable source of biomarkers for minimally invasive liquid biopsies, and its therapeutic potential, particularly as targeted drug delivery vehicles and immunomodulatory agents, showcasing their promise for enhancing the efficacy and safety of cancer treatments. By deciphering the intricate messages carried by EVs, we can gain a deeper understanding of cancer biology and develop more effective strategies for early detection, targeted therapy, and immunotherapy, paving the way for a new era of personalized and precise cancer medicine with the potential to significantly improve patient outcomes.

• Klíčová slova
• Biomarkers, Cancer, Extracellular vesicles, Immune evasion, Liquid biopsy, Metastasis, Targeted therapy, Tumor microenvironment,
• MeSH
• extracelulární vezikuly * metabolismus   MeSH
• lidé MeSH
• mezibuněčná komunikace * MeSH
• nádorové biomarkery metabolismus   MeSH
• nádorové mikroprostředí MeSH
• nádory * metabolismus   patologie   etiologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH
• Názvy látek
• nádorové biomarkery MeSH

Exosomes, nanosized extracellular vesicles released by various cell types, are intensively studied for the diagnosis and treatment of cancer and neurodegenerative diseases, and they also display high usability in regenerative medicine. Emphasizing their diagnostic potential, exosomes serve as carriers of disease-specific biomarkers, enabling non-invasive early detection and personalized medicine. The cargo loading of exosomes with therapeutic agents presents an innovative strategy for targeted drug delivery, minimizing off-target effects and optimizing therapeutic interventions. In regenerative medicine, exosomes play a crucial role in intercellular communication, facilitating tissue regeneration through the transmission of bioactive molecules. While acknowledging existing challenges in standardization and scalability, ongoing research efforts aim to refine methodologies and address regulatory considerations. In summary, this review underscores the transformative potential of exosomes in reshaping the landscape of medical interventions, with a particular emphasis on cancer, neurodegenerative diseases, and regenerative medicine.

• Klíčová slova
• biomarker, cancer, exosomes, neurodegenerative diseases, regenerative medicine,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Ischemic stroke is a leading cause of mortality and long-term disability globally. One of its aspects is the breakdown of the blood-brain barrier (BBB). The disruption of BBB's integrity during stroke exacerbates neurological damage and hampers therapeutic intervention. Recent advances in regenerative medicine suggest that mesenchymal stem cells (MSCs) derived extracellular vesicles (EVs) show promise for restoring BBB integrity. This review explores the potential of MSC-derived EVs in mediating neuroprotective and reparative effects on the BBB after ischemic stroke. We highlight the molecular cargo of MSC-derived EVs, including miRNAs, and their role in enhancing angiogenesis, promoting the BBB and neural repair, and mitigating apoptosis. Furthermore, we discuss the challenges associated with the clinical translation of MSC-derived EV therapies and the possibilities of further enhancing EVs' innate protective qualities. Our findings underscore the need for further research to optimize the therapeutic potential of EVs and establish their efficacy and safety in clinical settings.

• Klíčová slova
• acute ischemic stroke, blood-brain barrier, blood-brain barrier integrity, exosome, extracellular vesicle, mesenchymal stem cell, miRNA, tight junction,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

BACKGROUND: Ovarian follicular fluid (FF) is a dynamic environment that changes with the seasons, affecting follicle development, ovulation, and oocyte quality. Cells in the follicles release tiny particles called extracellular vesicles (EVs) containing vital regulatory molecules, such as microRNAs (miRNAs). These miRNAs are pivotal in facilitating communication within the follicles through diverse signaling and information transfer forms. EV-coupled miRNA signaling is implicated to be associated with ovarian function, follicle and oocyte growth and response to various environmental insults. Herein, we investigated how seasonal variations directly influence the ovulatory and anovulatory states of ovarian follicles and how are they associated with follicular fluid EV-coupled miRNA dynamics in horses. RESULTS: Ultrasonographic monitoring and follicular fluid aspiration of preovulatory follicles in horses during the anovulatory (spring: non-breeding) and ovulatory (spring, summer, and fall: breeding) seasons and subsequent EV isolation and miRNA profiling identified significant variation in EV-miRNA cargo content. We identified 97 miRNAs with differential expression among the groups and specific clusters of miRNAs involved in the spring transition (miR-149, -200b, -206, -221, -328, and -615) and peak breeding period (including miR-143, -192, -451, -302b, -100, and let-7c). Bioinformatic analyses showed enrichments in various biological functions, e.g., transcription factor activity, transcription and transcription regulation, nucleic acid binding, sequence-specific DNA binding, p53 signaling, and post-translational modifications. Cluster analyses revealed distinct sets of significantly up- and down-regulated miRNAs associated with spring anovulatory (Cluster 1) and summer ovulation-the peak breeding season (Clusters 4 and 6). CONCLUSIONS: The findings from the current study shed light on the dynamics of FF-EV-coupled miRNAs in relation to equine ovulatory and anovulatory seasons, and their roles in understanding the mechanisms involved in seasonal shifts and ovulation during the breeding season warrant further investigation.

• Klíčová slova
• Extracellular vesicle, Follicle growth, Follicular fluid, Horse, Mare, Ovulation, Pre-ovulation, Seasonal breeding,
• Publikační typ
• časopisecké články MeSH

Spinal cord injury (SCI) is a devastating condition with a complex pathology that affects a significant portion of the population and causes long-term consequences. After primary injury, an inflammatory cascade of secondary injury occurs, followed by neuronal cell death and glial scar formation. Together with the limited regenerative capacity of the central nervous system, these are the main reasons for the poor prognosis after SCI. Despite recent advances, there is still no effective treatment. Promising therapeutic approaches include stem cells transplantation, which has demonstrated neuroprotective and immunomodulatory effects in SCI. This positive effect is thought to be mediated by small extracellular vesicles (sEVs); membrane-bound nanovesicles involved in intercellular communication through transport of functional proteins and RNA molecules. In this review, we summarize the current knowledge about sEVs and microRNA as their cargo as one of the most promising therapeutic approaches for the treatment of SCI. We provide a comprehensive overview of their role in SCI pathophysiology, neuroprotective potential and therapeutic effect.

• Klíčová slova
• miRNA, regeneration, small extracellular vesicles, spinal cord injury, stem cells,
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Colorectal cancer (CRC) is the second most prevalent cancer type worldwide, which highlights the urgent need for non-invasive biomarkers for its early detection and improved prognosis. We aimed to investigate the patterns of long non-coding RNAs (lncRNAs) in small extracellular vesicles (sEVs) collected from low-volume blood serum specimens of CRC patients, focusing on their potential as diagnostic biomarkers. Our research comprised two phases: an initial exploratory phase involving RNA sequencing of sEVs from 76 CRC patients and 29 healthy controls, and a subsequent validation phase with a larger cohort of 159 CRC patients and 138 healthy controls. Techniques such as dynamic light scattering, transmission electron microscopy, and Western blotting were utilized for sEV characterization. Optimized protocol for sEV purification, RNA isolation and preamplification was applied to successfully sequence the RNA content of sEVs and validate the results by RT-qPCR. We successfully isolated sEVs from blood serum and prepared sequencing libraries from a low amount of RNA. High-throughput sequencing identified differential levels of 460 transcripts between CRC patients and healthy controls, including mRNAs, lncRNAs, and pseudogenes, with approximately 20% being lncRNAs, highlighting several tumor-specific lncRNAs that have not been associated with CRC development and progression. The validation phase confirmed the upregulation of three lncRNAs (NALT1, AL096828, and LINC01637) in blood serum of CRC patients. This study not only identified lncRNA profiles in a population of sEVs from low-volume blood serum specimens of CRC patients but also highlights the value of innovative techniques in biomolecular research, particularly for the detection and analysis of low-abundance biomolecules in clinical samples. The identification of specific lncRNAs associated with CRC provides a foundation for future research into their functional roles in cancer development and potential clinical applications.

• Klíčová slova
• Biomarker, Colorectal cancer, EVs, lncRNAs,
• MeSH
• biologické markery MeSH
• extracelulární vezikuly * genetika   MeSH
• kolorektální nádory * diagnóza   genetika   MeSH
• lidé MeSH
• RNA dlouhá nekódující * genetika   MeSH
• sekundární malignity * MeSH
• sérum MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• biologické markery MeSH
• RNA dlouhá nekódující * MeSH

This review presents a comprehensive overview of labelling strategies for endogenous and exogenous extracellular vesicles, that can be utilised both in vitro and in vivo. It covers a broad spectrum of approaches, including fluorescent and bioluminescent labelling, and provides an analysis of their applications, strengths, and limitations. Furthermore, this article presents techniques that use radioactive tracers and contrast agents with the ability to track EVs both spatially and temporally. Emphasis is also placed on endogenous labelling mechanisms, represented by Cre-lox and CRISPR-Cas systems, which are powerful and flexible tools for real-time EV monitoring or tracking their fate in target cells. By summarizing the latest developments across these diverse labelling techniques, this review provides researchers with a reference to select the most appropriate labelling method for their EV based research.

• Klíčová slova
• Bioluminescent labelling, CRISPR-Cas, Cre-loxP, EV tracking, Exosomes, Extracellular vesicles, Fluorescent labelling, In vivo imaging, Magnetic resonance, Radiolabelling,
• MeSH
• extracelulární vezikuly * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH

High-grade serous carcinoma of the ovary, fallopian tube and peritoneum (HGSC), the most common type of ovarian cancer, ranks among the deadliest malignancies. Many HGSC patients have excess fluid in the peritoneum called ascites. Ascites is a tumour microenvironment (TME) containing various cells, proteins and extracellular vesicles (EVs). We isolated EVs from patients' ascites by orthogonal methods and analyzed them by mass spectrometry. We identified not only a set of 'core ascitic EV-associated proteins' but also defined their subset unique to HGSC ascites. Using single-cell RNA sequencing data, we mapped the origin of HGSC-specific EVs to different types of cells present in ascites. Surprisingly, EVs did not come predominantly from tumour cells but from non-malignant cell types such as macrophages and fibroblasts. Flow cytometry of ascitic cells in combination with analysis of EV protein composition in matched samples showed that analysis of cell type-specific EV markers in HGSC has more substantial prognostic potential than analysis of ascitic cells. To conclude, we provide evidence that proteomic analysis of EVs can define the cellular composition of HGSC TME. This finding opens numerous avenues both for a better understanding of EV's role in tumour promotion/prevention and for improved HGSC diagnostics.

• Klíčová slova
• ascites, extracellular vesicles (EV), fallopian tube and peritoneum (HGSC), high-grade serous carcinoma of the ovary, macrophage, ovarian cancer (OC), tandem mass spectrometry (MS/MS), tumour microenvironment (TME),
• MeSH
• ascites metabolismus   patologie   MeSH
• extracelulární vezikuly * metabolismus   MeSH
• lidé MeSH
• nádorové mikroprostředí MeSH
• nádory vaječníků * diagnóza   MeSH
• proteomika MeSH
• serózní cystadenokarcinom * diagnóza   genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH