Ultracentrifugation Versus Kit Exosome Isolation: NanoLC–MS and Other Tools Reveal Similar Performance Biomarkers, But Also Contaminations

Skottvoll, Frøydis Sved; Berg, Henriette Engen; Bjørseth, Kamilla; Lund, Kaja; Roos, Norbert; Bekhradnia, Sara; Thiede, Bernd; Sandberg, Cecilie; Vik-Mo, Einar O.; Røberg-Larsen, Hanne; Nyström, Bo; Lundanes, Elsa; Wilson, Steven Ray

doi:10.4155/fsoa-2018-0088

Cited by 29 publications

(22 citation statements)

References 67 publications

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“…It is demonstrated that sEVs obtained by a polymer-based commercial precipitation kit contained abundant contaminants, including aggregated proteins [24,25]. Basically, a principal of these kits is similar to the PEG method in the present study where the aqueous solubility of sEVs is reduced [15].…”

Section: Discussionmentioning

confidence: 93%

Optimal Isolation Method of Small Extracellular Vesicles from Rat Plasma

Otani

Fujioka

Okada

et al. 2019

IJMS

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Small extracellular vesicles (sEVs) mediate cell–to–cell communication. We recently reported that circulating sEVs regulate systolic blood pressure in an animal model of human systemic hypertension. However, the underlying mechanisms still remain to be elucidated. As the first step for detailed analyses, we sought to increase the yield and purity of sEVs isolated from rat plasma. We compared the concentration and size distribution of sEVs as well as protein expression of the sEV marker and contaminants among plasma sEVs isolated by the ultracentrifugation (UC) method, the precipitation with polyethylene-glycol and ultracentrifugation (PEG-UC) method, or the precipitation with polyethylene-glycol (PEG) method. Effects of anticoagulants were also examined. The total concentration of plasma sEVs isolated by the PEG or PEG-UC method was much higher than that of the UC method. In the plasma sEVs isolated by the PEG-UC method, contaminating proteins were lower, while the protein expression of certain sEV markers was higher than that of the PEG method. There was no significant difference in total concentration or protein expression of sEV markers in sEVs isolated from rat plasma treated with three different anticoagulants (heparin, ethylenediaminetetraacetic acid, or acid citrate dextrose buffer) by the PEG-UC method. We, for the first time, determined that the PEG-UC method was optimal for sEV isolation from rat plasma.

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Section: Discussionmentioning

confidence: 93%

Optimal Isolation Method of Small Extracellular Vesicles from Rat Plasma

Otani

Fujioka

Okada

et al. 2019

IJMS

View full text Add to dashboard Cite

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“…Thirdly, we isolated our exosomes by ultra-centrifugation and ultra-filtration using a 100 kDa MWCO ultrafiltration filter, which could allow the possibility of other extracellular vesicles and proteins larger than 100 kDa to also be isolated alongside the exosomes. Alternatively, we could have used a commercially available exosome isolation kit, however studies have shown that exosomes isolated via kits could potentially contain contamination from chemicals in the kits that could affect in-vivo applications and analysis 50,51 . Lastly, we presented here the proteomic and genomic composition of LSC-Sec and LSC-Exo, respectively, but it would be clinically and pharmaceutically beneficial to determine which proteins, small molecules, or RNAs are the functional components capable of achieving the therapeutic effects.…”

Section: Discussionmentioning

confidence: 99%

Inhalation of lung spheroid cell secretome and exosomes promotes lung repair in pulmonary fibrosis

et al. 2020

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Idiopathic pulmonary fibrosis (IPF) is a fatal and incurable form of interstitial lung disease in which persistent injury results in scar tissue formation. As fibrosis thickens, the lung tissue loses the ability to facilitate gas exchange and provide cells with needed oxygen. Currently, IPF has few treatment options and no effective therapies, aside from lung transplant. Here we present a series of studies utilizing lung spheroid cell-secretome (LSC-Sec) and exosomes (LSC-Exo) by inhalation to treat different models of lung injury and fibrosis. Analysis reveals that LSC-Sec and LSC-Exo treatments could attenuate and resolve bleomycin-and silica-induced fibrosis by reestablishing normal alveolar structure and decreasing both collagen accumulation and myofibroblast proliferation. Additionally, LSC-Sec and LSC-Exo exhibit superior therapeutic benefits than their counterparts derived from mesenchymal stem cells in some measures. We showed that an inhalation treatment of secretome and exosome exhibited therapeutic potential for lung regeneration in two experimental models of pulmonary fibrosis.

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“…They demonstrated that density gradient ultracentrifugation gave the purest exosome preparations, while the other three techniques also co-isolate contaminating factors. Similarly, Skottvoll et al (2019) evaluated the performance of different isolation methods based on differential ultracentrifugation and a commercial isolation kit (total exosome isolation reagent). These results shown that the two isolation methods had similar performance with only some differences based on the origin of the cell.…”

Section: Extracellular Vesicles: Origins and Functionsmentioning

confidence: 99%

Extracellular Vesicles: Delivery Vehicles of Myokines

2019

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Movement and regular physical activity are two important factors that help the human body prevent, reduce and treat different chronic diseases such as obesity, type 2 diabetes, heart diseases, hypertension, sarcopenia, cachexia and cancer. During exercise, several tissues release molecules into the blood stream, and are able to mediate beneficial effects throughout the whole body. In particular, contracting skeletal muscle cells have the capacity to communicate with other organs through the release of humoral factors that play an important role in the mechanisms of adaptation to physical exercise. These muscle-derived factors, today recognized as myokines, act as endocrine and paracrine hormones. Moreover, exercise may stimulate the release of small membranous vesicles into circulation, whose composition is influenced by the same exercise. Combining the two hypotheses, these molecules related to exercise, named exer-kines, might be secreted from muscle cells inside small vesicles (nanovesicles). These could act as messengers in tissue cross talk during physical exercise. Thanks to their ability to deliver useful molecules (such as proteins and miRNA) in both physiological and pathological conditions, extracellular vesicles can be thought of as promising candidates for potential therapeutic and diagnostic applications for several diseases.

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Ultracentrifugation Versus Kit Exosome Isolation: NanoLC–MS and Other Tools Reveal Similar Performance Biomarkers, But Also Contaminations

Cited by 29 publications

References 67 publications

Optimal Isolation Method of Small Extracellular Vesicles from Rat Plasma

Optimal Isolation Method of Small Extracellular Vesicles from Rat Plasma

Inhalation of lung spheroid cell secretome and exosomes promotes lung repair in pulmonary fibrosis

Extracellular Vesicles: Delivery Vehicles of Myokines

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