scholarly journals Molecular Characterization of Temozolomide-Treated and Non Temozolomide-Treated Glioblastoma Cells Released Extracellular Vesicles and Their Role in the Macrophage Response

2020 ◽  
Vol 21 (21) ◽  
pp. 8353
Author(s):  
Elisa Panzarini ◽  
Stefano Tacconi ◽  
Elisabetta Carata ◽  
Stefania Mariano ◽  
Ada Maria Tata ◽  
...  
Keyword(s):  
Molecular Markers ◽  
Extracellular Vesicles ◽  
Cell Activation ◽  
Cell Communication ◽  
Molecular Profile ◽  
Surface Markers ◽  
Macrophage Response ◽  
A Cell ◽  
Potential Biomarkers

Extracellular vesicles (EVs) are widely investigated in glioblastoma multiforme (GBM) for their involvement in regulating GBM pathobiology as well as for their use as potential biomarkers. EVs, through cell-to-cell communication, can deliver proteins, nucleic acids, and lipids that are able to reprogram tumor-associated macrophages (TAMs). This research is aimed to concentrate, characterize, and identify molecular markers of EVs subtypes released by temozolomide (TMZ)-treated and non TMZ-treated four diverse GBM cells. Morphology, size distribution, and quantity of small (sEVs) and large (lEVs) vesicles were analyzed by cryo-TEM. Quality and quantity of EVs surface markers were evaluated, having been obtained by Western blotting. GBM cells shed a large amount of EVs, showing a cell line dependent molecular profile A comparative analysis distinguished sEVs and lEVs released by temozolomide (TMZ)-treated and non TMZ-treated GBM cells on the basis of quantity, size and markers expression. Finally, the GBM-derived sEVs and lEVs, irrespective of TMZ treatment, when challenged with macrophages, modulated cell activation toward a tendentially M2b-like phenotype.

scholarly journals The Biological Function of Extracellular Vesicles during Fertilization, Early Embryo—Maternal Crosstalk and Their Involvement in Reproduction: Review and Overview

Biomolecules ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. 1510
Author(s):  
Emanuele Capra ◽  
Anna Lange-Consiglio
Keyword(s):  
Extracellular Vesicles ◽  
Spatial Interaction ◽  
Cell Communication ◽  
Early Embryo ◽  
Molecular Profile ◽  
Cell Of Origin ◽  
Non Invasive ◽  
Temporal And Spatial ◽  
Mediate Cell

Secretory extracellular vesicles (EVs) are membrane-enclosed microparticles that mediate cell to cell communication in proximity to, or distant from, the cell of origin. Cells release a heterogeneous spectrum of EVs depending on their physiologic and metabolic state. Extracellular vesicles are generally classified as either exosomes or microvesicles depending on their size and biogenesis. Extracellular vesicles mediate temporal and spatial interaction during many events in sexual reproduction and supporting embryo-maternal dialogue. Although many omic technologies provide detailed understanding of the molecular cargo of EVs, the difficulty in obtaining populations of homogeneous EVs makes difficult to interpret the molecular profile of the molecules derived from a miscellaneous EV population. Notwithstanding, molecular characterization of EVs isolated in physiological and pathological conditions may increase our understanding of reproductive and obstetric diseases and assist the search for potential non-invasive biomarkers. Moreover, a more precise vision of the cocktail of biomolecules inside the EVs mediating communication between the embryo and mother could provide new insights to optimize the therapeutic action and safety of EV use.

scholarly journals Analysis of the RNA content of the exosomes derived from blood serum and urine and its potential as biomarkers

2014 ◽  
Vol 369 (1652) ◽  
pp. 20130502 ◽  
Author(s):  
Mu Li ◽  
Emily Zeringer ◽  
Timothy Barta ◽  
Jeoffrey Schageman ◽  
Angie Cheng ◽  
...  
Keyword(s):  
Blood Serum ◽  
Cell Communication ◽  
Cell Culture Model ◽  
Culture Model ◽  
Wide Range ◽  
Abnormal Cells ◽  
A Cell ◽  
Invasive Diagnostics ◽  
Serum And Urine

Exosomes are tiny vesicles (30–150 nm) constantly secreted by all healthy and abnormal cells, and found in abundance in all body fluids. These vesicles, loaded with unique RNA and protein cargo, have a wide range of biological functions, including cell-to-cell communication and signalling. As such, exosomes hold tremendous potential as biomarkers and could lead to the development of minimally invasive diagnostics and next generation therapies within the next few years. Here, we describe the strategies for isolation of exosomes from human blood serum and urine, characterization of their RNA cargo by sequencing, and present the initial data on exosome labelling and uptake tracing in a cell culture model. The value of exosomes for clinical applications is discussed with an emphasis on their potential for diagnosing and treating neurodegenerative diseases and brain cancer.

Microglia extracellular vesicles: focus on molecular composition and biological function

2021 ◽  
Vol 49 (4) ◽  
pp. 1779-1790 ◽  
Author(s):  
Lorenzo Ceccarelli ◽  
Chiara Giacomelli ◽  
Laura Marchetti ◽  
Claudia Martini
Keyword(s):  
Extracellular Vesicles ◽  
Molecular Mechanisms ◽  
Biological Function ◽  
Biological Effects ◽  
Genetic Material ◽  
Cell Communication ◽  
Molecular Composition ◽  
Cargo Proteins ◽  
A Cell ◽  
The Central Nervous System

Extracellular vesicles (EVs) are a heterogeneous family of cell-derived lipid bounded vesicles comprising exosomes and microvesicles. They are potentially produced by all types of cells and are used as a cell-to-cell communication method that allows protein, lipid, and genetic material exchange. Microglia cells produce a large number of EVs both in resting and activated conditions, in the latter case changing their production and related biological effects. Several actions of microglia in the central nervous system are ascribed to EVs, but the molecular mechanisms by which each effect occurs are still largely unknown. Conflicting functions have been ascribed to microglia-derived EVs starting from the neuronal support and ending with the propagation of inflammation and neurodegeneration, confirming the crucial role of these organelles in tuning brain homeostasis. Despite the increasing number of studies reported on microglia-EVs, there is also a lot of fragmentation in the knowledge on the mechanism at the basis of their production and modification of their cargo. In this review, a collection of literature data about the surface and cargo proteins and lipids as well as the miRNA content of EVs produced by microglial cells has been reported. A special highlight was given to the works in which the EV molecular composition is linked to a precise biological function.

scholarly journals Multiplexed Profiling of Single-cell Extracellular Vesicles Secretion

2018 ◽  
Author(s):  
Yahui Ji ◽  
Dongyuan Qi ◽  
Linmei Li ◽  
Haoran Su ◽  
Xiaojie Li ◽  
...  
Keyword(s):  
Single Cell ◽  
Cell Lines ◽  
Extracellular Vesicles ◽  
Single Cells ◽  
Cell Communication ◽  
Deep Understanding ◽  
Surface Markers ◽  
Cell Heterogeneity ◽  
Health And Disease ◽  
Cytokines Secretion

AbstractExtracellular vesicles (EVs) are important intercellular mediators regulating health and disease. Conventional EVs surface marker profiling, which was based on population measurements, masked the cell-to-cell heterogeneity in the quantity and phenotypes of EVs secretion. Herein, by using spatially patterned antibodies barcode, we realized multiplexed profiling of single-cell EVs secretion from more than 1000 single cells simultaneously. Applying this platform to profile human oral squamous cell carcinoma (OSCC) cell lines led to deep understanding of previously undifferentiated single cell heterogeneity underlying EVs secretion. Notably, we observed the decrement of certain EV phenotypes (e.g. CD63+EVs) were associated with the invasive feature of both OSCC cell lines and primary OSCC cells. We also realized multiplexed detection of EVs secretion and cytokines secretion simultaneously from the same single cells to investigate multidimensional spectrum of intercellular communications, from which we resolved three functional subgroups with distinct secretion profiles by visualized clustering. In particular, we found EVs secretion and cytokines secretion were generally dominated by different cell subgroups. The technology introduced here enables comprehensive evaluation of EVs secretion heterogeneity at single cell level, which may become an indispensable tool to complement current single cell analysis and EV research.SignificanceExtracellular vesicles (EVs) are cell derived nano-sized particles medicating cell-cell communication and transferring biology information molecules like nucleic acids to regulate human health and disease. Conventional methods for EV surface markers profiling can’t tell the differences in the quantity and phenotypes of EVs secretion between cells. To address this need, we developed a platform for profiling an array of surface markers on EVs from large numbers of single cells, enabling more comprehensive monitoring of cellular communications. Single cell EVs secretion assay led to previously unobserved cell heterogeneity underlying EVs secretion, which might open up new avenues for studying cell communication and cell microenvironment in both basic and clinical research.

The more the better – determining the optimal range when performing single-vesicle phenotyping

2021 ◽  
Vol 1 (1) ◽  
pp. 26-33
Author(s):  
María Gómez-Serrano ◽  
Christian Preußer ◽  
Kathrin Stelter ◽  
Elke Pogge von Strandmann
Keyword(s):  
Extracellular Vesicles ◽  
Particle Number ◽  
Experiment Design ◽  
Surface Markers ◽  
Optimal Range ◽  
Design And Optimization ◽  
Highly Sensitive ◽  
Single Vesicle

The characterization of extracellular vesicles (EVs) has evolved rapidly in recent years due to advances in straightforward technologies. Based on these more sensitive methods, it is now possible to describe EV populations in their entirety more precisely. However, these applications require an equivalently delicate experiment design and optimization steps to draw valid conclusions in the end. One of these methods is represented by the highly sensitive nanoflow cytometry (nFCM), by which particles can be analyzed not only on their size (< 40 nm) and concentration but also concerning surface markers. In this work, we addressed some of the potential caveats of this method, especially when characterizing particles with fluorescently labelled antibodies. In particular, we show, when using low particle concentrations, which are inevitably encountered when working with EVs, the characterization of surface markers is prone to significantly varying. We hypothesized that these technical limitations could respond to the stickiness of EVs and should be properly counteracted. As a reference, we strongly recommend performing particle number-based comparisons with at least 109 particles as staining input in nFCM analyses. Moreover, we provided representative particle-number based immunoblotting results, underlying the significance of this parameter as a normalizer in future EV research.

scholarly journals Human Tumor-Infiltrating Dendritic Cells: From in Situ Visualization to High-Dimensional Analyses

Cancers ◽  
2019 ◽  
Vol 11 (8) ◽  
pp. 1082 ◽  
Author(s):  
Hubert ◽  
Gobbini ◽  
Bendriss-Vermare ◽  
Caux ◽  
Valladeau-Guilemond
Keyword(s):  
Dendritic Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Human Tumor ◽  
High Dimensional ◽  
Prognostic Impact ◽  
In Situ Visualization ◽  
A Cell

The interaction between tumor cells and the immune system is considered to be a dynamic process. Dendritic cells (DCs) play a pivotal role in anti-tumor immunity owing to their outstanding T cell activation ability. Their functions and activities are broad ranged, triggering different mechanisms and responses to the DC subset. Several studies identified in situ human tumor-infiltrating DCs by immunostaining using a limited number of markers. However, considering the heterogeneity of DC subsets, the identification of each subtype present in the immune infiltrate is essential. To achieve this, studies initially relied on flow cytometry analyses to provide a precise characterization of tumor-associated DC subsets based on a combination of multiple markers. The concomitant development of advanced technologies, such as mass cytometry or complete transcriptome sequencing of a cell population or at a single cell level, has provided further details on previously identified populations, has unveiled previously unknown populations, and has finally led to the standardization of the DCs classification across tissues and species. Here, we review the evolution of tumor-associated DC description, from in situ visualization to their characterization with high-dimensional technologies, and the clinical use of these findings specifically focusing on the prognostic impact of DCs in cancers.

scholarly journals Quantification and Phenotypic Characterization of Extracellular Vesicles from Patients with Acute Myeloid and B-Cell Lymphoblastic Leukemia

Cancers ◽  
2021 ◽  
Vol 14 (1) ◽  
pp. 56
Author(s):  
Marijana Miljkovic-Licina ◽  
Nicolas Arraud ◽  
Aicha Dorra Zahra ◽  
Patricia Ropraz ◽  
Thomas Matthes
Keyword(s):  
Extracellular Vesicles ◽  
Lymphoblastic Leukemia ◽  
Cell Communication ◽  
Phenotypic Characterization ◽  
Cryo Electron Microscopy ◽  
Small Cohort ◽  
Leukemia Development ◽  
Free Plasma

Extracellular vesicles (EVs) act in cell-to-cell communication, delivering cargo from donor to recipient cells and modulating their physiological condition. EVs secreted by leukemic blasts in patients with leukemia have been shown to influence the fate of recipient cells in the bone marrow microenvironment. Methods to quantify and to characterize them phenotypically are therefore urgently needed to study their functional role in leukemia development and to evaluate their potential as targets for therapy. We have used cryo-electron microscopy to study morphology and size of leukemic EVs, and nanoparticle tracking analysis and fluorescence triggering flow cytometry to quantify EVs in platelet-free plasma from a small cohort of leukemia patients and healthy blood donors. Additional studies with a capture bead-based assay allowed us to establish phenotypic signatures of leukemic EVs from 17 AML and 3 B-ALL patients by evaluating the expression of 37 surface antigens. In addition to tetraspanins and lineage-specific markers we found several adhesion molecules (CD29, and CD146) to be highly expressed by EVs from B-ALL and several leukemic stem cell antigens (CD44, CD105, CD133, and SSEA-4) to be expressed by EVs from AML patients. Further improvements in analytical methods to study EVs are needed before potentially using them as biomarkers for leukemia prognosis and follow-up.

scholarly journals Population Analysis of Extracellular Vesicles in Microvolumes of Biofluids

2020 ◽  
Author(s):  
Joana Maia ◽  
Silvia Batista ◽  
Nuno Couto ◽  
Ana C. Gregório ◽  
Cristian Bodo ◽  
...  
Keyword(s):  
Flow Cytometry ◽  
Extracellular Vesicles ◽  
Population Analysis ◽  
Cell Communication ◽  
Membrane Vesicles ◽  
Sample Volume ◽  
Clinical Samples ◽  
Liquid Biopsies ◽  
Clinical Biomarkers

AbstractExtracellular Vesicles (EVs), membrane vesicles released by all cells, are emerging mediators of cell-cell communication. By carrying biomolecules from tissues to biofluids, EVs have attracted attention as non-invasive sources of clinical biomarkers in liquid biopsies. Although frequently employed for content characterization of EVs, the study of bulk preparations lacks information on sub-populations and the intrinsic heterogeneity of vesicles. Importantly, these strategies also difficult the characterization of EVs from small quantities of samples. We here present a Flow Cytometry strategy that enables detailed population analysis of EVs, at the same time decreasing sample volume requirements and accelerating the overall processing time. We show its unique application for quality control of isolates of EVs by comparing the proportion of vesicular and non-vesicular particles in samples prepared by different protocols. In addition, we demonstrate its suitability for the study of populations of EVs from samples characterized by challenging small volumes. To illustrate that, we perform longitudinal non-lethal analysis of EVs in mouse plasma and in single-animal collections of murine vitreous humor. By allowing for the analysis of EVs from minimal amounts of sample, our Flow Cytometry strategy has an unexplored potential in the study of EVs in clinical samples with intrinsically limited volumes. When compared to conventional methods, it also multiplies by several times the number of different analytes that can be studied from a single collection of biofluid.

scholarly journals Effects of Cocaine on Human Glial-Derived Extracellular Vesicles

2021 ◽  
Vol 8 ◽  
Author(s):  
Sanjay Kumar ◽  
Qiana L. Matthews ◽  
Brian Sims
Keyword(s):  
Extracellular Vesicles ◽  
Cell Activation ◽  
Microglial Cell ◽  
Drugs Of Abuse ◽  
Inflammatory Responses ◽  
Cell Communication ◽  
Brain Parenchyma ◽  
Cocaine Exposure ◽  
Apoptotic Marker ◽  
The Central Nervous System

BackgroundMicroglia are important myeloid cells present in the brain parenchyma that serve a surveillance function in the central nervous system. Microglial cell activation results in neuroinflammation that, when prolonged, can disrupt immune homeostasis and neurogenesis. Activated microglia-derived extracellular vesicles (EVs) may be involved in the propagation of inflammatory responses and modulation of cell-to-cell communication. However, a complete understanding of how EVs are regulated by drugs of abuse, such as cocaine, is still lacking.FindingsCocaine exposure reduced human microglial cell (HMC3) viability, decreased expression of CD63 and dectin-1 in HMC3-derived EVs, and increased expression of the apoptotic marker histone H2A.x in HMC3-derived EVs.ConclusionCocaine impacts HMC3 cell viability and specific EV protein expression, which could disrupt cellular signaling and cell-to-cell communication.

Sign in / Sign up

Export Citation Format

Share Document