[Different virulence Mycobacterium promotes RAW264.7 macrophage autophagy and phosphorylation of mTOR and AKT]

Objective To explore the change of autophagy levels of the macrophages infected by Mycobacterium with different virulence. Methods RAW264.7 cells were infected with Mycobacterium tuberculosis standard strain (H37Rv), Bacille Calmette Guerin (BCG) and Mycobacterium smegmatis (Ms). The cell autophagy was detected by flow cytometry and the colocalization of phagosomes and lysosomes was detected by confocal laser scanning microscopy. Meanwhile, the autophagy-associated protein LC3, mTOR, p-mTOR, AKT and p-AKT were detected by Western blotting. Results At 24 hours after RAW264.7 cells were infected by Mycobacterium with different virulence, flow cytometry showed that the level of autophagy was significantly up-regulated by H37Rv, BCG and Ms infection, and the highest level was in the Ms infection group. The level of LC3-II was significantly up-regulated after H37Rv, BCG and Ms infection, and the tendency was consistent with the result of flow cytometry. The colocalization rates of phagosomes and lysosomes after H37Rv, BCG and Ms infection were (11.33±0.88)%, (18.33±0.88)% and (48.67±0.66)%. The expression of mTOR and AKT had no significant changes after H37Rv, BCG and Ms infection, but the phosphorylation level significantly increased, which meant that PI3K-AKT-mTOR was activated. Conclusion Mycobacterium infection can induce the autophagy in macrophages and promote the phosphorylation of mTOR and AKT.