Fig. 3: NF-κB promotes tau processing in primary microglia.
From: Microglial NF-κB drives tau spreading and toxicity in a mouse model of tauopathy
a, b Representative images and quantification of 24 h fluorescent tau fibrils accumulation in primary Ikbkb−/− (a) and IkbkbCA (b) microglia compared with their littermate Ikbkb+/+ and IkbkbWT control, respectively. Scale bar, 25 μm. Values are mean ± SD, relative to littermate control. Total N = 9 wells/group from three independent experiments. P-values were from multilevel mixed-effect model with experiment as hierarchical level, ***p < 0.001. c Schematic diagram of pulse-chase assay to quantify tau clearance in microglia. d–g Representative images and quantifications of pulse-chase assay comparing Ikbkb−/− with Ikbkb+/+ microglia (d, e 25,000 cells/well) and comparing IkbkbCA with IkbkbWT microglia (f, g 20,000 cells/well). Scale bar, 25 μm. Values are mean ± SD, relative to 0 h. Total N = 16 wells (6 h, Ikbkb+/+); 20 wells(6 h, Ikbkb−/−); 14 wells/group of other conditions from three independent experiments. P-values were from multilevel mixed-effect model with experiment as hierarchical level, ***p < 0.001. h Schematic diagram illustrating the isolation procedures of human AD-tau. i Schematic diagram of quantifying release of AD-tau from microglia. j Detection of intracellular pTau by AT8 (green), cell membranes by Wheat Germ Agglutinin (WGA) (red), and nuclei by DAPI (blue). A representative image from three independent experiments. Scale bar: 10 μm. k Quantification of secreted and the intracellular pTau by AT8 ELISA and shown as the percentage of total pTau loaded in microglia after 2 h incubation. N = 4 wells. A representative dataset from two independent experiments. Values are mean ± SEM. l Representative images of tau aggregation in HEK biosensor cells induced by CM from microglia treated with AD-tau. Insert: High magnification confocal image of tau aggregation. Scale bar: 50 μm, insert: 15 μm. m Quantification of tau aggregation positive cells. N = 4 (control) and 10 (AD-tau) fields from three independent experiments, values are mean ± SEM, two-tailed Mann–Whitney test. n, o TPCA-1-treated microglia were incubated 2 h with AD-tau followed by 24 h chase. The pTau released to medium (n) and intracellular pTau (o) were determined by AT8 ELISA. A representative dataset from two independent experiments. N = 4 wells, values are mean ± SEM, relative to DMSO, two-tailed t-test. Source data are provided as a Source data file.
