Original ArticleSize-dependent cellular uptake of exosomes
Graphical Abstract
Exosomes from glioma-associated stem cells (GASC) were purified using either ExoQuick (EQ) or ultracentrifugation (UC). The two purification methods generated different particle size distributions and atomic force microscope images showed that EQ preparations were richer in smaller vesicles compared to UC. Also, cellular uptake from A172 glioblastoma cells showed that small EQ exosomes were preferentially internalized compared to UC and this increased the motility of EQ-exposed cells compared to UC and a negative control (NC) not exposed to GASC exosomes. We concluded that smaller exosomes are preferentially internalized and they elicit a cellular response more effectively.
Section snippets
Cell cultures
Human glioma samples were collected at the Department of Neuroscience, Santa Maria della Misericordia University Hospital in Udine, after a written informed consent from the patient was obtained, in accordance with the declaration of Helsinki, and with approval by the independent ethics committee of the hospital. Glioma associated stem cells (GASC) and A172 human glioblastoma cells were cultured as described in26, 27 and detailed in the Supplementary methods.
Exosome purification, size and density measurements
A172 and GASC supernatants were
Effect of the extraction method on the size distribution of exosomal fractions
EQ and UC isolated exosomes from three GASC cultures supernatants derived from the same patient were adsorbed on mica coverslips and imaged using AFM to estimate the size distribution of the particles in the exosome fractions. Figure 1, A and B are representative 5 × 5 μm images of EQ and UC purified exosomes respectively, obtained from the same supernatant. Although both preparations produced individually separated particles with a near-spherical morphology, the EQ method generated particles
Discussion
In this work, we emphasized the importance of looking at the physical properties of exosomal preparations rather than limiting the characterization to molecular attributes. Our rationale is that exosomes play an important role in cellular communication and, regardless of the specific molecules they carry, the nature of the “envelope” might be of great importance for the delivery of the message. For example, in the case of nanoengineered nanoparticles it has been shown that size per se can
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Funding: Timea Palmai-Pallag was funded by the BBSRC grant BB/K019597/1.
Conflict of interest: No competing interests are present.