OBJECTIVE: To explore the mechanism underlying the molecular immune response of macrophages stimulated with exosome [(+)exosome] from macrophages after Mycobacterium avium (M. avium) infection and analyze the differential protein component of the exosome. METHODS: The culture supernatants of M.avium-infected macrophages and uninfected ones were collected and exosome was harvested from the supernatants by frozen ultra centrifugation. The concentrations of IFN-γ, TNF-α in supernatants were detected by enzyme-linked immunosorbent assay (ELISA), and CD80, CD86 expressions on macrophages were analyzed by flow cytometry after macrophages were stimulated with exosome. Meanwhile, 2-DE MALDI TOF/TOF MS was used to identify differentially expressed proteins of exosome between M. avium infected group and uninfected group. RESULTS: IFN-γ, TNF-α concentrations were increased in the supernatant after stimulated with (+)exosome and CD80, CD86 were raised on macrophage surface by stimulation with (+)exosome. With 2-DE MALDI TOF/TOF MS analysis, we obtained 18 differentially expressed proteins and 12 proteins were identified successfully. CONCLUSION: (+)exosome induces TNF-α, IFN-γ secretion from macrophages and result in the promotion of inflammatory response. In addition, (+)exosome enhances CD80 and CD86 protein expressions. The function of the differentially expressed proteins we identified is closely related to cytoskeleton, protein synthesis and processing, inflammatory response.
OBJECTIVE: To explore the mechanism underlying the molecular immune response of macrophages stimulated with exosome [(+)exosome] from macrophages after Mycobacterium avium (M. avium) infection and analyze the differential protein component of the exosome. METHODS: The culture supernatants of M.avium-infected macrophages and uninfected ones were collected and exosome was harvested from the supernatants by frozen ultra centrifugation. The concentrations of IFN-γ, TNF-α in supernatants were detected by enzyme-linked immunosorbent assay (ELISA), and CD80, CD86 expressions on macrophages were analyzed by flow cytometry after macrophages were stimulated with exosome. Meanwhile, 2-DE MALDI TOF/TOF MS was used to identify differentially expressed proteins of exosome between M. avium infected group and uninfected group. RESULTS: IFN-γ, TNF-α concentrations were increased in the supernatant after stimulated with (+)exosome and CD80, CD86 were raised on macrophage surface by stimulation with (+)exosome. With 2-DE MALDI TOF/TOF MS analysis, we obtained 18 differentially expressed proteins and 12 proteins were identified successfully. CONCLUSION: (+)exosome induces TNF-α, IFN-γ secretion from macrophages and result in the promotion of inflammatory response. In addition, (+)exosome enhances CD80 and CD86 protein expressions. The function of the differentially expressed proteins we identified is closely related to cytoskeleton, protein synthesis and processing, inflammatory response.