Extracellular vesicles from microalgae Pavlova in non-heated condition potently enhance innate immune response as a...

In vaccine therapy, there is a growing interest in novel immunostimulatory adjuvants derived from food products or dietary supplements with safe profiles. Pavlova is a microalga commonly used as a dietary supplement. Owing to its immune-activating potency and safety profiles, Pavlova offers novel possibilities as an immunostimulatory adjuvant. However, precise control of their proliferation profile and subsequent immune activation when administered to the host is difficult, limiting their practicality as adjuvants. Extracellular vesicles (EVs) are cell-derived nanoparticles containing nucleic acids and proteins. EVs exhibit biological activity without proliferating, indicating their potential in medical applications as novel cell-free materials. However, information on EVs secreted by microalgae, including Pavlova, is limited. In this study, we characterized the EVs from Pavlova as an immunostimulatory adjuvant. Heated Pavlova-derived EVs (hP-EVs) and non-heated Pavlova-derived EVs (n-hP-EVs) were purified from their culture supernatants, with heated Pavlova secreting larger amounts of EVs. These EVs exhibited a particle size of approximately 130-170 nm and a negatively-charged zeta potential. RAW264.7 macrophage cells more actively took up n-hP-EVs than hP-EVs via endocytosis. As an adjuvant activity, n-hP-EVs potently increased the production of the inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), which are indicators of enhanced innate immune response. Moreover, the effect of the storage temperature on the particle size of n-hP-EVs was small, maintaining their ability to activate innate immune responses. Thus, n-hP-EVs demonstrate attractive characteristics for use as novel cell-derived immunostimulatory adjuvants.