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MicroRNAs are minor constituents of extracellular vesicles that are rarely delivered to target cells

Manuel Albanese , Yen-Fu Adam Chen , Corinna Hüls, Kathrin Gärtner, Takanobu Tagawa, Ernesto Mejias-Perez, Oliver T. Keppler, Christine Göbel, Reinhard Zeidler, Mikhail Shein, Anne K. Schütz, Wolfgang Hammerschmidt

Mammalian cells release different types of vesicles, collectively termed extracellular vesicles (EVs). EVs contain cellular microRNAs (miRNAs) with an apparent potential to deliver their miRNA cargo to recipient cells to affect the stability of individual mRNAs and the cells’ transcriptome. The extent to which miRNAs are exported via the EV route and whether they contribute to cell-cell communication are controversial.

Cells release different types of extracellular vesicles (EVs) into the extracellular space. EVs have been reported to transfer proteins and RNA molecules from cell to cell and are thought to be important vehicles of intercellular communication [1]. They are released by a broad range of cell types and have been found in all body fluids, including blood, urine, and saliva [2–4]. A class of EVs, termed exosomes, can originate from cytoplasmic multivesicular bodies (MVB), which fuse with the cellular plasma membrane to release a burst of EVs.

Materials and methods:

Cell lines and cell culture
EBV-infected primary human B cells, the resulting lymphoblastoid cell lines (LCLs), peripheral blood mononuclear cell (PBMCs), the monocytic cell lines THP-1, the EBV-positive Burkitt lymphoma cell line Raji and the HEK293-based EBV producer cell lines were maintained in RPMI medium 1640 (Life Technologies). HEK293T cells were maintained in DMEM (Life Technologies).

Separation of human primary lymphocytes
Human primary B cells were prepared from adenoidal mononuclear cells by Ficoll Hypaque (PAN Biotech) gradient centrifugation as described in [66]. B cells were isolated using CD19 MicroBeads and MACS separation columns (Miltenyi Biotec), according to the manufacturer’s instruction.

Isolation of extracellular vesicles (EVs)
We submitted all relevant data of our experiments to EV-TRACK knowledgebase (EV-TRACK ID: EV200039) (EV-TRACK et al., 2017). EVs were isolated from the supernatant of EBV-infected B cells incubated for 72 hours with medium containing EV-free FCS.

Many groups have reported that miRNAs contained in EVs are involved in a wide range of paracrine and endocrine biological activities and fulfil important functions in different types of target cells [16,17,36–39] (for a small selection of papers). Similarly, viral miRNAs encoded by EBV were reported to be delivered via exosomal transfer from latently infected B cells to recipient cells where viral miRNAs repress their cognate cellular mRNA targets.

Citation: Albanese M, Chen Y-FA, Hüls C, Gärtner K, Tagawa T, Mejias-Perez E, et al. (2021) MicroRNAs are minor constituents of extracellular vesicles that are rarely delivered to target cells. PLoS Genet 17(12): e1009951.

Editor: Lin He, University of California Berkeley, UNITED STATES

Received: June 24, 2021; Accepted: November 16, 2021; Published: December 6, 2021.

Copyright: © 2021 Albanese et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Data Availability: All relevant data are within the manuscript and its Supporting Information files.

Funding: Funding for this research was provided by Deutsche Forschungsgemeinschaft grant nr. SFB1064/TP A13 to WH, Deutsche Forschungsgemeinschaft grant nr. SFB-TR36/TP A04 to WH, Deutsche Krebshilfe, grant nr. 70112875 to WH, and by the National Cancer Institute (grant nr. CA70723). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Competing interests: The authors have declared that no competing interests exist.

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