Isolation and Identification of Fusobacterium necrophorum OMVs at Different Growth Phase and its Effect on the Secretion of TNF-α and IL-8 in RAW264.7 Cells
To investigate the morphology,protein composition,and function of outer membrane vesicles(OMVs)in Fusobacterium necrophorum throughout the growth phase,F.necrophorum OMVs were isolated and purified using gradient centrifugation.A transmission electron microscope was used to analyze the morphology of OMVs(TEM).SDS-PAGE and Western blot were used to analyze the protein compositions.After stimulation with OMVs,ELISA was used to measure the levels of IL-8 and TNF-α in the cells.The results showed that after 2 hours of culture,F.necrophorum entered the logarithmic phase,peaked at 8 hours,and then entered the stationary phase and decline phase.The levels of 43K OMP and leukotoxin in OMVs produced by F.necrophorum cultured for 24 hours were significantly higher than those in other groups(P<0.01).OMVs produced by F.necrophorum cultured for 12 hours stimulated RAW264.7 cells secreted IL-8 and TNF-α level after 12 and 24 hours was the highest.As a result,OMVs produced by F.necrophorum stable phase could better stimulate RAW264.7 cells to produce inflammatory factors IL-8 and TNF-α.
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NSTL
万方数据
维普
