Potentials and capabilities of the Extracellular Vesicle (EV) Array

Jørgensen, Maléne Møller; Bæk, Rikke; Varming, Kim

doi:10.3402/jev.v4.26048

Cited by 71 publications

(64 citation statements)

References 12 publications

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“…Additionally, the EV array might be an interesting alternative for future studies. 40 Although we have shown many important advances in EVs' labeling and imaging, many questions are still needed to be answered in future studies. We have shown that labeling of hBM-MSCs with PKH26 or Molday ION is an effective way to isolate labeled EVs from them.…”

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confidence: 99%

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Imaging of extracellular vesicles derived from human bone marrow mesenchymal stem cells using fluorescent and magnetic labels

Dąbrowska

Fattore

Karnas

et al. 2018

IJN

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BackgroundMesenchymal stem cells have been shown therapeutic in various neurological disorders. Recent studies support the notion that the predominant mechanism by which MSCs act is through the release of extracellular vesicles (EVs). EVs seem to have similar therapeutic activity as their cellular counterparts and may represent an interesting alternative standalone therapy for various diseases. The aim of the study was to optimize the method of EV imaging to better understand therapeutic effects mediated by EVs.MethodsThe fluorescent lipophilic stain PKH26 and superparamagnetic iron oxide nanoparticles conjugated with rhodamine (Molday ION Rhodamine B™) were used for the labeling of vesicles in human bone marrow MSCs (hBM-MSCs). The entire cycle from intracellular vesicles to EVs followed by their uptake by hBM-MSCs has been studied. The identity of vesicles has been proven by antibodies against: anti-CD9, -CD63, and -CD81 (tetraspanins). NanoSight particle tracking analysis (NTA), high-resolution flow cytometric analysis, transmission electron microscopy (TEM), ELYRA PS.1 super-resolution microscopy, and magnetic resonance imaging (MRI) were used for the characterization of vesicles.ResultsThe PKH26 and Molday ION were exclusively localized in intracellular vesicles positively stained for EV markers: CD9, CD63, and CD81. The isolated EVs represent heterogeneous population of various sizes as confirmed by NTA. The TEM and MRI were capable to show successful labeling of EVs using ION. Co-culture of EVs with hBM-MSCs revealed their uptake by cells in vitro, as visualized by the co-localization of PKH26 or Molday ION with tetraspanins inside hBM-MSCs.ConclusionPKH26 and Molday ION seem to be biocompatible with EVs, and the labeling did not interfere with the capability of EVs to re-enter hBM-MSCs during co-culture in vitro. Magnetic properties of IONs provide an additional advantage for the imaging of EV using TEM and MRI.

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confidence: 99%

“…Interestingly, Molday ION was uniformly distributed inside the EVs in our study, while the nonfluorescent USPIO nanoparticles reported by others looked rather clumped. 40 Further studies to investigate if such difference in intravesicular distribution affects their therapeutic potential are warranted.…”

mentioning

confidence: 99%

Imaging of extracellular vesicles derived from human bone marrow mesenchymal stem cells using fluorescent and magnetic labels

Dąbrowska

Fattore

Karnas

et al. 2018

IJN

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“…After the detection of p53 in exosomes that were secreted at least by one cell line, we decided to test whether it is possible to detect this protein in exosomes of human body fluids, e.g., to confirm its presence in exosomes of blood plasma (Jorgensen et al, 2015). Immunoblotting has not given a positive result: no p53 was observed in exosomes of blood plasma.…”

Section: Resultsmentioning

confidence: 99%

“…Some authors believe that p53 can enhance the secretion of exosomes by activating the transcription of several genes that are involved in their production (Yu et al, 2006;Lespagnol et al, 2008;Naghibalhossaini et al, 2011;Lim et al, 2012). However, to date, p53 has not been found in exosomes that are secreted by cell lines (Kalra et al, 2012;Keerthikumar et al, 2015), with only one study demonstrating its presence in exosomes of human blood plasma (Jorgensen et al, 2015). Thus, this problem requires a more detailed study.…”

Section: Introductionmentioning

confidence: 99%

Exosomes Transfer p53 between Cells and Can Suppress Growth and Proliferation of p53-Negative Cells

et al. 2018

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Exosomes are nanosized vesicles that are secreted by many types of cells. We have found that exosomes secreted by HEK293 and HT-1080 can suppress growth and proliferation of p53-deficient cells. Upon overexpression of exogenous p53-GFP in HEK293 cells, we observed p53 protein in exosomes that were secreted by these cells. We also found endogenous p53 in exosomes that were secreted by HT-1080 cells with a higher level of p53 expression. We were able to detect endogenous p53 protein in exosomes that originated from human plasma and were transferred to p53-deficient cells. Our findings indicate that p53 protein can be transferred between cells and may play an important physiological role.

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“…A recent example applied to EV is the "EV array" [21,22]. The EV array utilizes slides printed with an array of antibodies to capture EV and then determines the amount of captured EV by labeling with a cocktail of biotinylated CD9, CD63 and CD81 and the use of streptavidin-Cy5 as a reporter molecule.…”

Section: C3 Fluorescent Immunosorbent Assay (Flisa)mentioning

confidence: 99%

Bulk immunoassays for analysis of extracellular vesicles

2017

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There is increasing clinical interest in extracellular vesicles (EV) for diagnostic and treatment purposes. This review provides an overview of bulk immunoassays to analyse EV. Western blot and enzyme-linked immunosorbent assay are still the two predominant bulk immunoassays. Recently, new assays have become available that can detect exposure to EV concentrations that are up to 10,000-fold lower. This is advantageous for applications that detect rare EV. Other important parameters are the detectable concentration range, the required sample volume, whether simultaneous presence of different antigens on a single EV can be detected, size selectivity of each assay and practical considerations. In this review, we will explain the working principles of the traditional and novel assays together with their performance parameters. The most sensitive assays are micro-nuclear magnetic resonance, surface plasmon resonance, and time-resolved fluorescent immunoassay.

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Potentials and capabilities of the Extracellular Vesicle (EV) Array

Cited by 71 publications

References 12 publications

Imaging of extracellular vesicles derived from human bone marrow mesenchymal stem cells using fluorescent and magnetic labels

Imaging of extracellular vesicles derived from human bone marrow mesenchymal stem cells using fluorescent and magnetic labels

Exosomes Transfer p53 between Cells and Can Suppress Growth and Proliferation of p53-Negative Cells

Bulk immunoassays for analysis of extracellular vesicles

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