Fig. 3

ROS activates macrophages M1 polarization through MAPK-NFκB P65 signaling pathway. A and B Raw264.7 cells were treated with H₂O₂ or H₂O₂ + DPI. M1 macrophages surface expression (CD86) was detected using flow cytometry. H2O2 could promote M1 macrophage polarization, but DPI had no effect on M1 macrophage polarization induced by H2O2. C and D The ERK1/2 and p-ERK1/2 protein expression in Raw264.7 cells was either investigated. H2O2 could increase the P-ERK/ERK level and that DPI had no influence on the phosphorylation of ERK in cells cultured with H2O2. E–G P65 protein expression are determined by immunofluorescences and flow cytometry. P65 expression was also increased after H2O2 treatment for 24 h, and DPI could not suppress the P65 fluorescence increase caused by H2O2. H and I Raw264.7 cells were treated with H₂O₂, H₂O₂ + BAY 11–7082 or H₂O₂ + PD98059. J and K Raw264.7 cells were treated with LPS, LPS + BAY 11–7082 or LPS + PD98059. L–N P65 protein expression are determined by immunofluorescences and flow cytometry. M1 macrophages surface expression (CD86) was detected using using flow cytometry. macrophage M1 polarization caused by both LPS and H2O2 could be suppressed by the P65 inhibitor. PD98059, an inhibitor of MEK, could also inhibit P65 expression and that M1 macrophage polarization was increased by LPS or H2O2. *p < 0.05, ***p < 0.01