Extracellular vesicles production and proteomic cargo varies with incubation time and temperature

Rayamajhi, Sagar; Sulthana, Shoukath; Ferrel, Colin; Shrestha, Tej B.; Aryal, Santosh

doi:10.1016/j.yexcr.2022.113454

Cited by 10 publications

(7 citation statements)

References 34 publications

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“…We showed that ExoArc-isolated EVs from HAoEC exhibited higher consistency and less EV aggregations than UC. The temporal heterogeneity of EV secreted by inflamed HAoEC also highlights the potential inconsistency in manufactured EVs which may affect the therapeutic effects . With the automated peristaltic pump system, in-line EV monitoring can be established as a quality control measure for early-stage EV therapeutics and autologous EV manufacturing to optimize production time and enhance EV yield.…”

Section: Discussionmentioning

confidence: 99%

High-Throughput Microfluidic Extraction of Platelet-free Plasma for MicroRNA and Extracellular Vesicle Analysis

Leong,

Lok,

Goh

et al. 2024

ACS Nano

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Cell-free RNAs and extracellular vesicles (EVs) are valuable biomarkers in liquid biopsies, but they are prone to preanalytical variabilities such as nonstandardized centrifugation or ex vivo blood degradation. Herein, we report a high-throughput and labelfree inertial microfluidic device (ExoArc) for isolation of platelet-free plasma from blood for RNA and EV analysis. Unlike conventional inertial microfluidic devices widely used for cell sorting, a submicrometer size cutoff (500 nm) was achieved which completely removed all leukocytes, RBCs, platelets, and cellular debris based on differential lateral migration induced by Dean vortices. The single-step operation also reduced platelet-associated miRNAs (∼2-fold) compared to centrifugation. We clinically validated ExoArc for plasma miRNA profiling (39 samples) and identified a 7-miRNA panel that detects non-small cell lung cancer with ∼90% sensitivity. ExoArc was also coupled with size exclusion chromatography (SEC) to isolate EVs within 50 min with ∼10-fold higher yield than ultracentrifugation. As a proof-of-concept for EV-based transcriptomics analysis, we performed miRNA analysis in healthy and type 2 diabetes mellitus (T2DM) subjects (n = 3 per group) by coupling ExoArc and ExoArc+SEC with quantitative polymerase chain reaction (RT-qPCR) assay. Among 293 miRNAs detected, plasmas and EVs showed distinct differentially expressed miRNAs in T2DM subjects. We further demonstrated automated in-line EV sorting from low volume culture media for continuous EV monitoring. Overall, the developed ExoArc offers a convenient centrifugation-free workflow to automate plasma and EV isolation for point-of-care diagnostics and quality control in EV manufacturing.

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Section: Discussionmentioning

confidence: 99%

High-Throughput Microfluidic Extraction of Platelet-free Plasma for MicroRNA and Extracellular Vesicle Analysis

Leong,

Lok,

Goh

et al. 2024

ACS Nano

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“…For instance, a study by Rayamajhi et al highlighted that an increase in the incubation temperature of cells not only augments EV secretion but also results in the significant enrichment of Serpinb1, at least in EVs secreted by K7M2 mouse osteosarcoma cells. Rayamajhi et al further depicted that cellular stresses induced by heat and nutrient deprivation trigger the increased detection of anti-proliferative and pro-proliferative proteins in EVs, respectively [68]. Similarly, exercise-induced EVs are enriched in various myokines as wells as proteins, such as RAB1A, ANXA2, ITGB1, ITGB2, ITGB5, Alix and FLOT1 [57].…”

Section: Cargo Of Evsmentioning

confidence: 99%

“…In the recent years, the use of in vitro and in vivo assessments, including patient samples, has allowed for the expansion of knowledge concerning the various biomolecular profiles within EVs [63][64][65]. Studies further indicate the dynamic alterations in the functional cargo of EVs, which are often influenced by intrinsic and/or extrinsic factors such as temperature, hypoxia, exercise and diseases [57,[66][67][68]. For instance, a study by Rayamajhi et al highlighted that an increase in the incubation temperature of cells not only augments EV secretion but also results in the significant enrichment of Serpinb1, at least in EVs secreted by K7M2 mouse osteosarcoma cells.…”

Section: Cargo Of Evsmentioning

confidence: 99%

Extracellular Vesicles in Pathophysiology: A Prudent Target That Requires Careful Consideration

Shahi,

Kang,

Fonseka

2024

Cells

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Extracellular vesicles (EVs) are membrane-bound particles released by cells to perform multitudes of biological functions. Owing to their significant implications in diseases, the pathophysiological role of EVs continues to be extensively studied, leading research to neglect the need to explore their role in normal physiology. Despite this, many identified physiological functions of EVs, including, but not limited to, tissue repair, early development and aging, are attributed to their modulatory role in various signaling pathways via intercellular communication. EVs are widely perceived as a potential therapeutic strategy for better prognosis, primarily through utilization as a mode of delivery vehicle. Moreover, disease-associated EVs serve as candidates for the targeted inhibition by pharmacological or genetic means. However, these attempts are often accompanied by major challenges, such as off-target effects, which may result in adverse phenotypes. This renders the clinical efficacy of EVs elusive, indicating that further understanding of the specific role of EVs in physiology may enhance their utility. This review highlights the essential role of EVs in maintaining cellular homeostasis under different physiological settings, and also discusses the various aspects that may potentially hinder the robust utility of EV-based therapeutics.

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“…Extracellular Vesicles (EVs) are proteolipid bi-layered particles (15)(16)(17)(18)(19), secreted by eukaryotic cells (fungi (20), protozoa (21), plants (22) and mammals (23)); archaea (24); and bacteria. EVs are released under normal and pathological conditions by metabolically active cells (25).…”

Section: Introductionmentioning

confidence: 99%

Unveiling Oral Anaerobic Bacteria Outer Membrane Vesicles: A Comprehensive Systematic Review

Castro-Vargas,

Barloy-Hubler,

Acuña-Amador

2024

Odovtos - Int J Dent Sc

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Extracellular vesicles (EV) are spherical structures limited by membranes and shed by several cell types. Specifically, outer membrane vesicles (OMVs) are nanometric particles constitutively produced by Gram-negative bacteria (GNB) under different environmental conditions. OMVs are biologically active; they are loaded with selected lipids, polysaccharides, proteins, and even different types of nucleic acids. OMVs from pathogenic oral bacteria play key roles in pathogen-host interactions, constituting a possible link between oral health and systemic disease. OMVs participate in adhesion, invasion, and damage to cells, as well as in modulating the host's immune response, biofilm formation, and promotion of virulence. The objective of this systematic review was to collect, analyze and synthesize the knowledge available on literature reviews on OMVs of the most studied pathogenic oral anaerobic GNB. This information was classified into the following categories: induction of vesiculation and biogenesis, its liberation from the parental cell, content, internalization by another host cell, and the interaction with the host cell. It was found that the most studied OMVs are those of Porphyromonas gingivalis and Bacteroides spp. and, to a lesser extent, Aggregatibacter spp., and Treponema spp. This systematic review provides a synthesis of the current knowledge regarding OMVs, with emphasis on the information available for periodontopathogens.

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Extracellular vesicles production and proteomic cargo varies with incubation time and temperature

Cited by 10 publications

References 34 publications

High-Throughput Microfluidic Extraction of Platelet-free Plasma for MicroRNA and Extracellular Vesicle Analysis

High-Throughput Microfluidic Extraction of Platelet-free Plasma for MicroRNA and Extracellular Vesicle Analysis

Extracellular Vesicles in Pathophysiology: A Prudent Target That Requires Careful Consideration

Unveiling Oral Anaerobic Bacteria Outer Membrane Vesicles: A Comprehensive Systematic Review

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