Focal χ-ray irradiation induces neuropathology in mouse brains. a Experimental timeline for focal-brain irradiation (25 Gy χ-rays) and subsequent analyses. Mice (6–8 weeks old) received a cumulative 25 Gy (5 Gy × 5 60Co χ-ray focal brain irradiation, followed by behavioral tests at 1, 4, 9, 12, and 16 wpi, with histological analyses at 16 wpi. b and c Immunofluorescence characterization of activated glial morphology in the cortex (b) and hippocampus (c) of mice at 2 wpi. Green fluorescence: microglial marker iba1; yellow fluorescence: astrocyte marker GFAP; red fluorescence: neuronal marker NeuN. d Y-maze behavioral profiling. Schematic of Y maze compartments: start arm (S), other arm (O), and novel arm (N). Heatmaps of exploration patterns at 1, 9, and 16 wpi. e Y-maze cognitive metrics. Number of arm entries and durations in the novel arm (n = 5 mice; one-way ANOVA). f Novel object recognition test. Schematic showing novel object A (cube). Heatmaps of exploration at 1, 9, and 16 wpi. g Recognition performance. Interaction frequency and distance traveled in relation to the novel object (n = 5 mice; one-way ANOVA). h Histopathological analysis at 16 wpi. HE staining (h1), Nissl staining of the hippocampal CA1 region (h2), phosphorylated Tau (p-Tau) immunohistochemical (DAB) staining (h3) and phosphorylated amyloid-beta (p-Aβ) immunohistochemical (DAB) staining (h4) were performed. i Quantification of degenerating neurons (n = 5 mouse sections; Student’s t test). j Quantification of Nissl bodies (n = 15 cells; Student’s t test). k Quantification of p-Tau-positive signals (n = 3 mouse sections; Student’s t test). l Quantification of DAB signals (n = 3 mouse sections; Student’s t test). All the statistical data are presented as the means ± SEMs. Scale bars: 20 μm, 5 μm (b, c), 100 μm (h1), 50 μm (h2), 20 μm (h3), and 20 μm (h4)
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