Fig. 5. Reversed nuclear localization of NF-kB (P65) in microglia reduces microglia TNF-α expression and inhibits EC necroptosis in co-culture system under OGDR.

a, d Expression levels of p-IκBα/IκBα in cytoplasm (a, b) and NF-kB (p65) nuclear (c, d) from microglia subjected to 0, 2, 6, and 12 h OGDR. n = 6; *P < 0.05 vs. control group; **P < 0.01 vs. control group. e–f Expression levels of NF-κB (p65) in microglia pretreated with or without BAY-117082 under OGDR. n = 6; *P = 0.0069 vs. vehicle group. g ELISA analysis for TNF-α show that BAY-117082 reduced TNF-α secretion from microglia under OGDR. n = 6; *P < 0.001 vs. control group; **P < 0.0001 vs. control group; ^#P < 0.0001 vs. vehicle group. h–k Representative images and statistical results of PI (red)/Hoechst (blue) staining and PI/annexin V flow cytometry of ECs treated with microglia (pretreated with BAY-117082 or vehicle) in co-culture system under OGDR. n = 6; *P < 0.01 vs. vehicle group; Scale bar: 100 μm. l–n Expression levels of p-RIP1/RIP1 and p-MLKL/MLKL in ECs from different group. n = 3; *P < 0.001 vs. control group; ^#P < 0.01 vs. vehicle group