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Engineered MSC-derived extracellular vesicles maintain proteomic characteristics comparable with naïve MSC-derived extracellular vesicles

ABSTRACT: Extracellular vesicles reflect the contents and characteristics of their donor cells. When donor cells were engineered to express specific protein, their characteristic may change, potentially altering the composition of EVs. This study aims to evaluate the degree of similarity in protein contents between engineered EVs and naïve EVs to investigate the impact of cellular engineering on EV composition.

INSTRUMENT(S): Orbitrap Fusion

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Stem Cell

SUBMITTER: Yoon Kyoung Kim

LAB HEAD: Gi-Hoon Nam

PROVIDER: PXD060047 | Pride | 2025-01-31

REPOSITORIES: pride

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			Action	DRS
	20220506_uniprot-proteome_UP000005640_add_sirp_rec.fasta	Fasta
	B10001Shiftbio-Exosome-RA20240416.mzML	Mzml
	B10001Shiftbio-Exosome-RA20240416.raw	Raw
	B10002Shiftbio-Exosome-RA20240416.mzML	Mzml
	B10002Shiftbio-Exosome-RA20240416.raw	Raw

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Project description:Background: Acute coronary syndromes (ACS) are associated with aberrant gene expression and epigenetic mechanisms. In particular, de novo DNA methylation is typically linked to gene silencing, but its role in heart disease remains not fully understood. Extracellular vesicles (EVs) are active components in cellular communication for their ability to carry a plethora of signalling biomolecules, thus representing a promising new diagnostic/therapeutic approach in cardiovascular diseases (CVDs). Indeed, there is the need of novel biomarkers for ACS prediction and timely detection. Purpose: We hypothesized that specific epigenetic signals can be carried by EVs. In this regard, we isolated and characterized circulating EVs from ACS patients and evaluated their potential role to influence DNA methylation in target cells. Methods: Circulating EVs were recovered, by ultracentrifugation, from plasma samples of 19 ACS patients and 50 healthy subjects (HS). Nanoparticle tracking analysis (NTA) and western blot (WB) were used to confirm the EVs integrity and purity. Peripheral blood mononuclear cells (PBMCs) of volunteer donors were treated with both ACS and HS derived EVs and genomic DNA was extracted to perform epigenome wide analysis through Reduced Representation Bisulfite Sequencing. ShinyGO, PANTHER, and STRING tools were interrogated to perform GO and PPI network analyses. Flow Cytometry, qRT-PCR, and WB analysis were also performed to evaluate and validate both intra-vesicular and intra-cellular signals. Results: Plasma ACS-derived EVs showed a significant up-regulation of DNA methyltransferases (DNMTs) gene expression levels as compared to HS (P<0.001). Specifically, de novo methylation transcripts, as DNMT3A and DNMT3B were significantly increased in plasma ACS-EVs. DNA methylation analysis of PBMCs from volunteer donors treated with HS- and ACS-derived EVs showed that RNF166 and CCND3 genes resulted the most hyper- and hypo-methylated, respectively, after by ACS-EV treatment. In addition, PPI network analysis specifically evidenced the subnetwork with SRC, as a hub gene, connecting it to NOTCH1, FOXO3, CDC42, IKBKG, RXRA, DGKG, known as important genes already involved in the onset of CVDs. Surprising, other novel genes, BAIAP2, SYP, CHL1, and SHB, which were hypomethylated, were found significantly overexpressed in PBMCs (P<0.005), underlying the fundamental modulating properties of EV cargo in atherosclerosis. Conclusions: These findings support the significant role of ACS plasma-derived EVs, inducing de novo DNA methylation signals, and modulating specific signaling pathways in target cells.

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Engineered MSC-derived extracellular vesicles maintain proteomic characteristics comparable with naïve MSC-derived extracellular vesicles

Dataset's files

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