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Fig. 4 | Journal of Neuroinflammation

Fig. 4

From: Microglia prevent peripheral immune cell invasion and promote an anti-inflammatory environment in the brain of APP-PS1 transgenic mice

Fig. 4

Microglia ablation did not alter amyloid-beta phagocytosis and revealed high numbers of macrophages located at sites of plaques resistant to CSF1R inhibition. a Representative image of Iba1+ cell at site of plaque with incorporated ThioflavinS+ particle in the hippocampus of APP-PS1 mice. The total number of Iba1+/ThioflavinS+ cells was significantly reduced upon PLX5622 treatment in the hippocampus (b) and the cortex (d). However, the percentage (%) of Iba1+/ThioflavinS+ cells from total Iba1+ cell counts remained the same in both brain regions of APP-PS1 animals treated with PLX5622 (c, e). Colocalization analysis with ThioflavinS was performed to analyze the plaque uptake by ThioflavinS+/Iba1+/TMEM119 cells. There was no observed increase in the amount of engulfed plaque material, represented by the percentage of dataset colocalized with ThioflavinS; however, in the cortex, the percentage of dataset colocalized was significantly reduced in microglia-ablated APP-PS1 mice (f). This was not seen in the hippocampus (g). Mechanically isolated CD11b+ cells were further characterized by their CD45 expression via flow cytometric analysis (h) to distinguish microglia (CD11b+/CD45low, green) from macrophage populations (CD11b+/CD45high, red). Besides strongly reduced CD11b+/CD45low microglia numbers in WT and APP-PS1 animals treated with PLX5622, APP-PS1 mice revealed significantly increased numbers of CD11b+/CD45low microglia compared to WT animals (i). Analysis of CD11b+/CD45high macrophages revealed significantly increased numbers of CD11b+/CD45high cells in APP-PS1 compared to WT and treatment with PLX5622 reduced this cell population in WT and APP-PS1 mice (j). Using the newly identified microglia marker TMEM119 for detailed immunohistochemical analysis in the hippocampus revealed strong co-localization of Iba1+ (white) cells with TMEM119 (red) in WT and WT animals treated with PLX5622; however, Iba1+ cells at sites of plaques (green) in APP-PS1 animals and APP-PS1 treated with PLX5622 did not express TMEM119 (k, arrow). Quantitative analysis of Iba1+/TMEM119+ revealed a significant reduction upon PLX5622 treatment in WT and APP-PS1 mice (l). Surprisingly, APP-PS1 animals had increased numbers of Iba1+/TMEM119 cells that were more resistant to PLX5622 treatment than in WT animals (m). (n) Calculation of the percentage of Iba1+/TMEM119+ and Iba1+/TMEM119 cells from the total Iba1+ cell population. ThioflavinS was used to stain amyloid plaques (green), and Dapi (blue) was used as nucleus stain. Unpaired Student’s t test (b, c, d n = 6/group; f, g n = 3/group; j comparing WT with WT + PLX5622 n = 8–9/group;) with Welch’s correction (e n = 6/group; i comparing WT with WT + PLX5622 n = 8–9/group) and one-way ANOVA with Tukey’s multiple comparison test (i, j n = 8–9/group; l, m, n n = 6/group) was performed. Scale: 5 μm (a), 20 μm (k)

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