Fig. 3: oTau seeding induces rapid accumulation of toxic tau and neurodegeneration in the AstTau model.
From: Single cell transcriptomic profiling of a neuron-astrocyte assembloid tauopathy model
a Representative images of tau hyperphosphorylation (pS202/205 (AT8), red), reactive astrogliosis (GFAP, green), and neurons (βIII tubulin (Tuj1), violet) at 14 DIV3D. Scale bars = 50 µm. b Quantification of tau hyperphosphorylation (AT8) normalized to MAP2 fluorescence intensity at 14 DIV3D. Data from five independent asteroids. Error bars = SEM. Two-way ANOVA with Tukey’s multiple-comparison test (MCT), ***P < 0.01. c Immunoblotting and quantification of hyperphosphorylated tau (AT8) normalized to internal control (GAPDH) band intensity. Data from three independent asteroid pools. Error bars = SEM. Unpaired t test, *P < 0.05. Source data are provided as a Source Data file. d Quantification of reactive astrogliosis (GFAP) normalized to DAPI fluorescence intensity at 14 DIV3D. Data from five independent asteroids. Error bars = SEM. Two-way ANOVA with Tukey’s MCT, ****P < 0.001. e Representative images of neurodegeneration (Fluoro Jade Blue (FJB)), neurons (Tuj1, violet), and astrocytes (S100β, red) at 21 DIV3D. Scale bars = 50 µm. f Quantification of astrocytic S100β normalized to DAPI fluorescence intensity. Data from five independent asteroids. Error bars = SEM. Two-way ANOVA with Tukey’s MCT, ***P < 0.01, ****P < 0.001, comparisons to timepoint vehicle control. g Quantification of FJB normalized to Tuj1 fluorescence intensity. Data from five independent asteroids. Error bars = SEM. Two-way ANOVA with Tukey’s MCT, ****P < 0.001, comparisons to timepoint vehicle control. h Immunoblotting of cell death activation by BAX labeling with internal control (GAPDH). i, j Quantification of BAX (i) and cleaved BAX (j) band intensity normalized to GAPDH from (h). Data from three independent asteroid pools. Error bars = SEM. Unpaired t test, **P < 0.01. k Representative image of mixed (NGN2_GFP + /oTau-, NGN2_GFP-/oTau+) cultures evidencing tau propagation (NGN2_GFP + /MC1 + , closed arrows). Misfolded tau (MC1, red), neurons (Tuj1, violet), select neurons NGN2_GFP + . Scale bars = 50 µm. l Representative images of propagated misfolded tau (MC1, red) in indicated conditions. Scale bars = 50 µm. m Quantification of MC1 normalized by DAPI fluorescence intensity from (l). Data from seven independent asteroids. Error bars = SEM. Two-way ANOVA with Tukey’s MCT, *P < 0.05. n Quantification of MC1 colocalization by Pearson’s Correlation to Tuj1 and NGN2-GFP in 21 DIV3D mixed cultures.
