Figure 1.

Germ-free 3xTg mice display reduced AD pathologies and improved cognitive functions compared with SPF 3xTg mice.

(A) Immunofluorescent staining of Aβ (red) and ThS (green) in frontal cortex region of brains, AT8 (green) and T22 (red) in hippocampus CA1 region of brains from germ-free 3xTg mice and SPF mice. Scale bar: 20 μm (B) quantitative analysis of Aβ positive cells, ThS positive cells, AT8 positive cells and T22 positive cells, respectively. The density of Aβ, ThS, AT8 and T22 positive cells were significantly increased in spf mice brain. (n=5 in each group, data are shown as mean±SEM. **P<0.01, ****p<0.0001 compared with control, unpaired t-tests.). (C) Aβ40 and Aβ42 concentrations in the cortex of germ-free 3xTg mice and SPF 3xTg mice were measured using human aβ40 and Aβ42 ELISA kit. the concentration of Aβ42 not aβ40 was significantly increased in spf 3xTg mice cortex compared with germ-free 3xTg mice cortex. (n=5 in each group, data are shown as mean±SEM ***p<0.001 compared with control, multiple unpaired T tests). (D, E) Y-maze behavioural tests. Spontaneous alternation (%) (D), number of arms entered (E); n=8 in each group, data are shown as mean±SEM. *P<0.05 compared with control, unpaired T tests. (F) Representative images of immunofluorescent staining of Iba-1 (red) in cortex (upper panel) and 3D reconstruction of Iba-1-stained microglia (lower panel) residing in the cortex of germ-free 3xTg mice and SPF 3xTg mice. (G–I) Quantitative analysis of diameter, number of branch points, and total branch length of microglia residing in the cortex. Data represent the mean±SEM; representative data of 12 samples; *p<0.05, ****p<0.0001 compared with control, unpaired t-tests. Aβ, amyloid-β; GF, germ-free; SPF, specific-pathogen-free.