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      Tangential Flow Filtration for Highly Efficient Concentration of Extracellular Vesicles from Large Volumes of Fluid

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          Abstract

          Concentration of extracellular vesicles (EVs) from biological fluids in a scalable and reproducible manner represents a major challenge. This study reports the use of tangential flow filtration (TFF) for the highly efficient isolation of EVs from large volumes of samples. When compared to ultracentrifugation (UC), which is the most widely used method to concentrate EVs, TFF is a more efficient, scalable, and gentler method. Comparative assessment of TFF and UC of conditioned cell culture media revealed that the former concentrates EVs of comparable physicochemical characteristics, but with higher yield, less single macromolecules and aggregates (<15 nm in size), and improved batch-to-batch consistency in half the processing time (1 h). The TFF protocol was then successfully implemented on fluids derived from patient lipoaspirate. EVs from adipose tissue are of high clinical relevance, as they are expected to mirror the regenerative properties of the parent cells.

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          Intercellular nanovesicle-mediated microRNA transfer: a mechanism of environmental modulation of hepatocellular cancer cell growth.

          Takayuki Kogure, Wen-Lang Lin, Irene K. Yan … (2011)
          Hepatocellular carcinoma (HCC) is characterized by a propensity for multifocality, growth by local spread, and dysregulation of multiple signaling pathways. These features may be determined by the tumoral microenvironment. The potential of tumor cells to modulate HCC growth and behavior by secreted proteins has been extensively studied. In contrast, the potential for genetic modulation is poorly understood. We investigated the role and involvement of tumor-derived nanovesicles capable of altering gene expression and characterized their ability to modulate cell signaling and biological effects in other cells. We show that HCC cells can produce nanovesicles and exosomes that differ in both RNA and protein content from their cells of origin. These can be taken up and internalized by other cells and can transmit a functional transgene. The microRNA (miRNA) content of these exosomes was examined, and a subset highly enriched within exosomes was identified. A combinatorial approach to identify potential targets identified transforming growth factor β activated kinase-1 (TAK1) as the most likely candidate pathway that could be modulated by these miRNAs. Loss of TAK1 has been implicated in hepatocarcinogenesis and is a biologically plausible target for intercellular modulation. We show that HCC cell-derived exosomes can modulate TAK1 expression and associated signaling and enhance transformed cell growth in recipient cells. Exosome-mediated miRNA transfer is an important mechanism of intercellular communication in HCC cells. These observations identify a unique intercellular mechanism that could potentially contribute to local spread, intrahepatic metastases, or multifocal growth in HCC. Copyright © 2011 American Association for the Study of Liver Diseases.
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            Extracellular vesicles: emerging targets for cancer therapy.

            Pieter Vader, Xandra O. Breakefield, Matthew Wood (2014)
            Extracellular vesicles (EVs), including exosomes, microvesicles, and apoptotic bodies, are released by almost all cell types, including tumour cells. Through transfer of their molecular contents, EVs are capable of altering the function of recipient cells. Increasing evidence suggests a key role for EV mediated intercellular communication in a variety of cellular processes involved in tumour development and progression, including immune suppression, angiogenesis, and metastasis. Aspects of EV biogenesis or function are therefore increasingly being considered as targets for anticancer therapy. Here, we summarise the current knowledge on the contributions of EVs to cancer pathogenesis and discuss novel therapeutic strategies to target EVs to prevent tumour growth and spread. Copyright © 2014 Elsevier Ltd. All rights reserved.
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              Platelet-derived growth factor regulates the secretion of extracellular vesicles by adipose mesenchymal stem cells and enhances their angiogenic potential

              Tatiana Lopatina, Stefania Bruno, Ciro Tetta … (2014)
              Background Several studies demonstrate the role of adipose mesenchymal stem cells (ASCs) in angiogenesis. The angiogenic mechanism has been ascribed to paracrine factors since these cells secrete a plenty of signal molecules and growth factors. Recently it has been suggested that besides soluble factors, extracellular vesicles (EVs) that include exosomes and microvesicles may play a major role in cell-to-cell communication. It has been shown that EVs are implicated in the angiogenic process. Results Herein we studied whether EVs released by ASCs may mediate the angiogenic activity of these cells. Our results demonstrated that ASC-derived EVs induced in vitro vessel-like structure formation by human microvascular endothelial cells (HMEC). EV-stimulated HMEC when injected subcutaneously within Matrigel in SCID mice formed vessels. Treatment of ASCs with platelet-derived growth factor (PDGF) stimulated the secretion of EVs, changed their protein composition and enhanced the angiogenic potential. At variance of EVs released in basal conditions, PDGF-EVs carried c-kit and SCF that played a role in angiogenesis as specific blocking antibodies inhibited in vitro vessel-like structure formation. The enhanced content of matrix metalloproteinases in PDGF-EVs may also account for their angiogenic activity. Conclusions Our findings indicate that EVs released by ASCs may contribute to the ASC-induced angiogenesis and suggest that PDGF may trigger the release of EVs with an enhanced angiogenic potential.
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                Author and article information

                Journal
                Journal ID (nlm-ta): Cells
                Journal ID (iso-abbrev): Cells
                Journal ID (publisher-id): cells
                Title: Cells
                Publisher: MDPI
                ISSN (Electronic): 2073-4409
                Publication date (Electronic): 16 December 2018
                Publication date Collection: December 2018
                Volume: 7
                Issue: 12
                Electronic Location Identifier: 273
                Affiliations
                [1 ]Department of Transplantation Medicine; Department of Physiology and Biomedical Engineering, Mayo Clinic, Jacksonville, FL 32224, USA; vilanilam.george@ 123456mayo.edu (G.V.); taylorticer33@ 123456gmail.com (T.T.)
                [2 ]Department of Molecular and Translational Medicine, University of Brescia, 25123 Brescia, Italy
                [3 ]Department of Neuroscience, Mayo Clinic, Jacksonville, FL 32224, USA; lin.wenlang@ 123456mayo.edu (W.-L.L.); dickson.dennis@ 123456mayo.edu (D.W.D.)
                [4 ]Department of Orthopedic Surgery, Mayo Clinic, Jacksonville, FL 32224, USA; shapiro.shane@ 123456mayo.edu
                [5 ]CSGI, Research Center for Colloids and Nanoscience, 50019 Florence, Italy
                [6 ]Department of Nanomedicine, Houston Methodist Research Institute, Houston, TX 77030, USA
                Author notes
                [* ]Correspondence: Busatto.sara@ 123456mayo.edu (S.B.); paolo.bergese@ 123456unibs.it (P.B.); wolfram.joy@ 123456mayo.edu (J.W.); Tel.: +1-(904)-953-2138 (S.B.); +39-(030)-371-7555 (P.B.); +1-(904)-953-0834 (J.W.)
                [†]

                These authors contributed equally to this work.

                Author information
                https://orcid.org/0000-0003-0845-670X
                https://orcid.org/0000-0002-4652-2168
                Article
                Publisher ID: cells-07-00273
                DOI: 10.3390/cells7120273
                PMC ID: 6315734
                PubMed ID: 30558352
                SO-VID: c57b7b65-9239-4640-ac5c-7efcc8af382f
                Copyright © © 2018 by the authors.
                License:

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

                History
                Date received : 11 October 2018
                Date accepted : 14 December 2018
                Categories
                Subject: Communication

                Keywords: exosome,extracellular vesicles,isolation,ultrafiltration,tangential flow filtration,lipoaspirate
                Data availability:
                Keywords: exosome, extracellular vesicles, isolation, ultrafiltration, tangential flow filtration, lipoaspirate

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