CpG ODN loaded exosome nanovesicles: enhanced immunostimulatory activity

Abstract

Exosomes are naturally occurring, membranous nanovesicles known to function as intercellular communication vectors. To explore whether these naturally occuring bilayer vesicles could be exploited as a nucleic acid delivery system we intentionally loaded exosomes with TLR9 ligands and studied their immunostimulatory properties. METHODS: Exosomes were isolated from RAW264.7 and EG-7 cell supernatants by differential centrifugation, filtration and ultracentrifugation and were loaded with CpG ODNs via dehydration-rehydration protocol. Splenocytes were stimulated and analyzed by flow cytometry for cell surface marker upregulation, intracellular cytokine production and co-stimulatory molecule upregulation. Confocal microscopy analyses were done to assess internalization properties of these nanovesciles. Cell supernatants were studied by ELISA to assess Th1 biased cytokine/chemokine secretion.