Fig. 6.

Anxa2 knockdown inhibited NF-κB signaling activation in OGD/R-treated BV2 microglial cells.A–D, representative blot images and quantitative analysis of NF-κB signaling components including phosphorylated p65 (p-p65), total p65, phosphorylated IKBa, and total IKBa in shCtrl and shAnxa2 BV2 cells after OGD/R detected by Western blot. GAPDH was an internal normalized control. E, relative luciferase activity of NF-κB after Anxa2 knockdown in OGD/R-induced BV2 cells. F and G, representative blot images and quantitative analysis of the subcellular accumulated NF-κB p65 in the cytoplasmic and nuclear fractions, respectively, in shCtrl and shAnxa2 BV2 cells after OGD/R detected by Western blot. As a protein loading control, the data were normalized to GAPDH (cytoplasmic marker) or Histone H3 (nuclear marker). H and I, representative images (H) and quantitative analysis (I) of the positive nucleus localization of NF-kB p65 subunit accessed by immunofluorescence staining in shCtrl and shAnxa2 BV2 cells after OGD/R. Scale bar represents 20 μm. Data are shown as mean ± SEM. Statistical significance was determined by two-way ANOVA for multiple comparisons, ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.001, ∗∗∗∗p < 0.0001. All experiments were repeated at least three times. Anxa2, annexin A2; NF-κB, nuclear factor-kappa B; OGD/R, oxygen-glucose deprivation and reoxygenation.