Deep dive on the proteome of salivary extracellular vesicles: comparison between ultracentrifugation and polymer-based precipitation isolation

Li, Meng; Lou, Doudou; Chen, Joyce; Shi, Ke‐Qing; Wang, Yong; Zhu, Qingfu; Liu, Fei; Zhang, Yating

doi:10.1007/s00216-020-03004-w

Cited by 22 publications

(23 citation statements)

References 34 publications

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“… 48 , 51–54 There are many isolation methods for sEV, and polyethylene glycol (PEG) precipitation is a rapid, convenient and high yield isolation method that widely used for the isolation of large amounts of sEV. 55 A recent study showed that the sEV obtained by PEG precipitation had the same high purity as that isolated by ultracentrifugation. 56 Therefore PEG precipitation was chosen to isolate sEV in our study.…”

Section: Discussionmentioning

confidence: 99%

Preservation of Small Extracellular Vesicle in Gelatin Methacryloyl Hydrogel Through Reduced Particles Aggregation for Therapeutic Applications

Wu¹,

He²,

Wu³

et al. 2021

IJN

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Purpose Small extracellular vesicles (sEV) play an irreplaceable role in cell–cell communication. However, sEV in solution aggregate with each other during preservation, leading to impairment of the structures, contents, and functions of sEV. Therefore, there is a need to develop an optimal preservation method that combines high recovery rate, low cost, convenience, and easy-transportation in one. In this study, a new preservation strategy different from the cryopreservation or lyophilization was developed by reducing sEV particles aggregation. Methods The sEV were encapsulated in thermoresponsive gelatin methacryloyl (GelMA) hydrogels at 4°C to reduce particles aggregation during the reversible cross-linking process. The sEV movement was visualized in different mediums and particles’ number, size, structure and protein of 28 days preserved sEV were compared to fresh sEV. Human umbilical vein endothelial cells (HUVEC) and rat adipose-derived stromal stem cells (rASC) were isolated and cultured with fresh and preserved sEV to test the cellular response. A mice subcutaneous model was adopted to detect controlled release and angiogenesis ability of preserved sEV. Results Through particles tracks visualization, GelMA hydrogels significantly decreased the sEV movement. After 28 days preservation in GelMA at 4°C, the particles number, size, structure and protein of sEV were similar to fresh sEV. In vitro, preserved sEV had the same ability to promote cell proliferation, migration and angiogenesis as fresh sEV. In vivo, preserved sEV-GelMA could artificially regulate the absorptivity of GelMA hydrogels and controlled released sEV for therapeutic application, and preserved sEV encapsulated in GelMA significantly promoted angiogenesis in mice. Conclusion Our results demonstrated that sEV encapsulated in GelMA could be a novel strategy for long-term preservation of sEV for therapeutic application.

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Section: Discussionmentioning

confidence: 99%

Preservation of Small Extracellular Vesicle in Gelatin Methacryloyl Hydrogel Through Reduced Particles Aggregation for Therapeutic Applications

Wu¹,

He²,

Wu³

et al. 2021

IJN

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“…Li et al. compared salivary EV proteins extracted from DUC and polyethylene glycol‐based precipitation, and the results indicated DUC method yield more EV‐related proteins [157]. Moreover, polymer‐based precipitation of plasma exosomes may face the challenge of insufficient efficiency and purity [158].…”

Section: Discussionmentioning

confidence: 99%

“…Notably, compared with DUC that is the gold standard, the application of polymer-based precipitation in EV isolation is still controversial. Li et al compared salivary EV proteins extracted from DUC and polyethylene glycol-based precipitation, and the results indicated DUC method yield more EV-related proteins[157]. Moreover, polymer-based precipitation of plasma exosomes may face the challenge of insufficient efficiency and purity[158].Besides, Subedi et al explored the effects of different lysis buffers on tumor-derived EV protein extraction.…”

mentioning

confidence: 99%

Proteomic profiling in extracellular vesicles for cancer detection and monitoring

et al. 2021

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Extracellular vesicles (EVs) are nanometer-size lipid vesicles released by cells, which play essential biological functions in intercellular communication. Increasing evidence indicates that EVs participate in cancer development, including invasion, migration, metastasis, and cancer immune modulation. One of the key mechanisms is that EVs affect different cells in the tumor microenvironment through surface-anchor proteins and protein cargos. Moreover, proteins specifically expressed in tumor-derived EVs can be applied in cancer diagnosis and monitoring. Besides, the EV proteome also helps to understand drug resistance in cancers and to guide clinical medication. With the development of mass spectrometry and array-based multi-protein detection, the research of EV proteomics has entered a new era. The high-throughput parallel proteomic profiling based on these new platforms allows us to study the impact of EV proteome on cancer progression more comprehensively and to describe the proteomic landscape in cancers with more details. In this article, we review the role and function of different types of EVs in cancer progression. More importantly, we summarize the proteomic profiling of EVs based on different methods and the application of EV proteome in cancer detection and monitoring.

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“…Based on their cumulative evidence in using SEC for isolating macromolecules, the authors concluded that use of precipitation reagents, such as ExoQuick, yielded increased quantity and quality of exosomal proteins and RNA, followed by SEC, then immunocapture; and the least effective method was ultracentrifugation [76]. However, as the science and technology of EV separation evolves, the precipitation method has been found to co-isolate EVs with other contaminants, and as such, is considered the least EV-specific method [77][78][79][80][81][82].…”

Section: Bloodmentioning

confidence: 99%

The Past, the Present, and the Future of the Size Exclusion Chromatography in Extracellular Vesicles Separation

Kaddour

Tranquille

Okeoma

2021

Viruses

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Extracellular vesicles (EVs) are cell-derived membranous particles secreted by all cell types (including virus infected and uninfected cells) into the extracellular milieu. EVs carry, protect, and transport a wide array of bioactive cargoes to recipient/target cells. EVs regulate physiological and pathophysiological processes in recipient cells and are important in therapeutics/drug delivery. Despite these great attributes of EVs, an efficient protocol for EV separation from biofluids is lacking. Numerous techniques have been adapted for the separation of EVs with size exclusion chromatography (SEC)-based methods being the most promising. Here, we review the SEC protocols used for EV separation, and discuss opportunities for significant improvements, such as the development of novel particle purification liquid chromatography (PPLC) system capable of tandem purification and characterization of biological and synthetic particles with near-single vesicle resolution. Finally, we identify future perspectives and current issues to make PPLC a tool capable of providing a unified, automated, adaptable, yet simple and affordable particle separation resource.

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Deep dive on the proteome of salivary extracellular vesicles: comparison between ultracentrifugation and polymer-based precipitation isolation

Cited by 22 publications

References 34 publications

Preservation of Small Extracellular Vesicle in Gelatin Methacryloyl Hydrogel Through Reduced Particles Aggregation for Therapeutic Applications

Preservation of Small Extracellular Vesicle in Gelatin Methacryloyl Hydrogel Through Reduced Particles Aggregation for Therapeutic Applications

Proteomic profiling in extracellular vesicles for cancer detection and monitoring

The Past, the Present, and the Future of the Size Exclusion Chromatography in Extracellular Vesicles Separation

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