Xi bao yu fen zi mian yi xue za zhi = Chinese journal of cellular and molecular immunology · 2015
[NADPH oxidase-induced macrophage autophagy mediated by reactive oxygen species in Aspergillus fumigatus infection].
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Hu Suxia, Yang Xiaokang, He Jiang
Аннотация
OBJECTIVE: To investigate whether or not NADPH oxidase (NOX) participates in Aspergillus fumigatus induced macrophage autophagy in RAW264.7 cells and explore the possible mechanism.
METHODS: RAW264.7 cells in the exponential growth phase were randomized into three groups: Aspergillus fumigatus conidia (MOI 5:1) treated group, Aspergillus fumigatus conidia plus NOX inhibitor diphenyleneiodonium chloride (DPI) treated group; negative control group (untreated). The levels of intracellular reactive oxygen species (ROS) were measured with dichiorodihydrofluorescein diacetate (DCFH-DA) staining by fluorescence microscope. The expression of microtubule-associated protein light chain-3 (LC3) protein in RAW264.7 cells was detected by Western blotting. Then the expression and positioning of LC-3 were observed by immunofluorescence staining.
RESULTS: After stimulated with conidia of Aspergillus fumigatus, RAW264.7 cells showed significant increases of ROS and LC3BII. Interestingly, LC3 was recruited to phagosomes which contained Aspergillus fumigatus conidia. After the NOX inhibitor DPI blocked the ROS production, the expressions of ROS and LC3BII had no significant increases as compared with the negative control group, and the distribution of LC3 was diffuse in cytoplasm.
CONCLUSION: NOX induces macrophage autophagy by producing ROS in Aspergillus fumigatus infection.
METHODS: RAW264.7 cells in the exponential growth phase were randomized into three groups: Aspergillus fumigatus conidia (MOI 5:1) treated group, Aspergillus fumigatus conidia plus NOX inhibitor diphenyleneiodonium chloride (DPI) treated group; negative control group (untreated). The levels of intracellular reactive oxygen species (ROS) were measured with dichiorodihydrofluorescein diacetate (DCFH-DA) staining by fluorescence microscope. The expression of microtubule-associated protein light chain-3 (LC3) protein in RAW264.7 cells was detected by Western blotting. Then the expression and positioning of LC-3 were observed by immunofluorescence staining.
RESULTS: After stimulated with conidia of Aspergillus fumigatus, RAW264.7 cells showed significant increases of ROS and LC3BII. Interestingly, LC3 was recruited to phagosomes which contained Aspergillus fumigatus conidia. After the NOX inhibitor DPI blocked the ROS production, the expressions of ROS and LC3BII had no significant increases as compared with the negative control group, and the distribution of LC3 was diffuse in cytoplasm.
CONCLUSION: NOX induces macrophage autophagy by producing ROS in Aspergillus fumigatus infection.