Original ArticleExosome-mediated delivery of functionally active miRNA-155 inhibitor to macrophages
Graphical Abstract
We used B cell derived exosomes as drug delivery vehicles to deliver miRNA inhibitor and miRNA mimic. Exosome loaded with a miRNA-155 inhibitor has delivered the miRNA-155 inhibitor into RAW (264.7) macrophages and decreased TNFα production in vitro. Exosomes loaded with a miRNA-155 mimic have successfully delivered miRNA-155 mimic to hepatocytes in vitro as well as to the liver and isolated hepatocytes of miRNA-155 knockout mice in vivo.
Section snippets
Cell culture and exosome isolation
Murine B cells (M12.4) were cultured in RPMI medium plus 10% exosome-depleted FBS (Exo-FBS™) (Mountain View, CA, USA), and 1% penicillin/streptomycin (Gibco®, NY, USA). After 12 hours, the cells were exposed to CD40 (5 μg/ml) (PeproTech. Rocky Hill, New Jersey, USA) and IL-4 (50 ng/ml) (PeproTech. Rocky Hill, New Jersey, USA). Three days later, the culture media were harvested and exosomes were isolated. RAW 264.7 macrophages were cultured in Dulbecco's modified medium (Invitrogen) containing 10%
Characterization of B cell derived exosomes
Exosomes derived from B (M12.4) and macrophages (RAW 264.7) cell lines were isolated as described in the method section. The average size of exosomes for stimulated B cell line (with IL-4/CD40), non-stimulated B cell line, and RAW macrophages was 98 nm, 108 nm and 102 nm, respectively, as measured by Nanoparticle Tracking Analysis (NTA) (NanoSight), which were all in the range of exosome size described earlier.1 Figure 1, A shows the size versus concentration (particles/ml) of exosomes derived
Discussion
In this report, exogenous miRNA-155 mimic and miRNA-155 inhibitor were successfully introduced into murine B cell-derived exosomes (membranous nanovesicles) and it was shown that loaded exosomes can deliver miRNA-155 mimic and miRNA-155 inhibitor to primary mouse hepatocytes and RAW 264.7 macrophages, respectively. B cell exosomes effectively delivered miRNA-155 inhibitor into the RAW 264.7 cells, causing inhibition of miRNA-155, and functionally led to a statistically significant decrease of
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This work was supported by National Institutes of Health PHS grants AA020744 and AA021907 to G.S.
Conflict of interest: The authors have declared that no conflict of interest exists.
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These authors contributed equally to this work.