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Spatial exosome analysis using cellulose nanofiber sheets reveals the location heterogeneity of extracellular vesicles.

ABSTRACT: Extracellular vesicles (EVs), including exosomes, are recognized as promising functional targets involved in disease mechanisms. However, the intravital heterogeneity of EVs remains unclear, and the general limitation for analyzing EVs is the need for a certain volume of biofluids. Here, we present cellulose nanofiber (CNF) sheets to resolve these issues. We show that CNF sheets capture and preserve EVs from ~10 μL of biofluid and enable the analysis of bioactive molecules inside EVs. By attaching CNF sheets to moistened organs, we collect EVs in trace amounts of ascites, which is sufficient to perform small RNA sequence analyses. In an ovarian cancer mouse model, we demonstrate that CNF sheets enable the detection of cancer-associated miRNAs from the very early phase when mice did not have apparent ascites, and that EVs from different locations have unique miRNA profiles. By performing CNF sheet analyses in patients, we identify further location-based differences in EV miRNA profiles, with profiles reflecting disease conditions. We conduct spatial exosome analyses using CNF sheets to reveal that ascites EVs from cancer patients exhibit location-dependent heterogeneity. This technique could provide insights into EV biology and suggests a clinical strategy contributing to cancer diagnosis, staging evaluation, and therapy planning.

SUBMITTER: Yokoi A

PROVIDER: S-EPMC10632339 | biostudies-literature | 2023 Nov

REPOSITORIES: biostudies-literature

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Spatial exosome analysis using cellulose nanofiber sheets reveals the location heterogeneity of extracellular vesicles.

Yokoi Akira A Yoshida Kosuke K Koga Hirotaka H Kitagawa Masami M Nagao Yukari Y Iida Mikiko M Kawaguchi Shota S Zhang Min M Nakayama Jun J Yamamoto Yusuke Y Baba Yoshinobu Y Kajiyama Hiroaki H Yasui Takao T

Nature communications 20231108 1

Extracellular vesicles (EVs), including exosomes, are recognized as promising functional targets involved in disease mechanisms. However, the intravital heterogeneity of EVs remains unclear, and the general limitation for analyzing EVs is the need for a certain volume of biofluids. Here, we present cellulose nanofiber (CNF) sheets to resolve these issues. We show that CNF sheets capture and preserve EVs from ~10 μL of biofluid and enable the analysis of bioactive molecules inside EVs. By attachi ...[more]

PMID: 37938557

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Project description:IntroductionOptical techniques are routinely used to size and count extracellular vesicles (EV). For comparison of data from different methods and laboratories, suitable calibrators are essential. A suitable calibrator must have a refractive index (RI) as close to that of EV as possible but the RI of EV is currently unknown. To measure EV, RI requires accurate knowledge of size and light scattering. These are difficult to measure as most EVs cannot be resolved by light microscopy and their diameter is smaller than the wavelength of visible light. However, nanoparticle tracking analysis (NTA) provides both size and relative light scattering intensity (rLSI) values. We therefore sought to determine whether it was possible to use NTA to measure the RI of individual EVs.MethodsNTA was used to measure the rLSI and size of polystyrene and silica microspheres of known size and RI (1.470 and 1.633, respectively) and of EV isolated from a wide range of cells. We developed software, based on Mie scattering code, to calculate particle RI from the rLSI data. This modelled theoretical scattering intensities for polystyrene and silica microspheres of known size (100 and 200 nm) and RI. The model was verified using data from the polystyrene and silica microspheres. Size and rLSI data for each vesicle were processed by the software to generate RI values.ResultsThe following modal RI measurements were obtained: fresh urinary EV 1.374, lyophilised urinary EV 1.367, neuroblastoma EV 1.393, blood EV 1.398, EV from activated platelets 1.390, small placental EV 1.364-1.375 and 1.398-1.414 for large placental EV (>200 nm). Large placental EV had a significantly higher RI than small placental EV (p<0.0001). The spread of RI values was narrower for small EV than for the more heterogeneous large EV.DiscussionUsing NTA and Mie scattering theory, we have demonstrated that it is possible to estimate the RI of sub-micron EV using NTA data. EV typically had a modal RI of 1.37-1.39, whereas values of >1.40 were observed for some large (>200 nm) microvesicles.ConclusionThis method for measuring EV RI will be useful for developing appropriate calibrators for EV measurement.

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Spatial exosome analysis using cellulose nanofiber sheets reveals the location heterogeneity of extracellular vesicles.

Publications

Spatial exosome analysis using cellulose nanofiber sheets reveals the location heterogeneity of extracellular vesicles.

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