Journal: Frontiers in Aging Neuroscience

Article Title: PU.1 dictates β-amyloid-induced TREM2 expression upregulation in microglia in a transgenic model of Alzheimer’s disease

doi: 10.3389/fnagi.2025.1537388

Figure Lengend Snippet: Elevated TREM2 protein and mRNA levels in activated microglia in the hippocampus of 5 × FAD mice during disease progression. (A) Analysis of the mRNA expression of TREM2 in the hippocampi of 1 month-old to 18–20 months-old 5 × FAD mice and aged-matched WT mice. The expression of TREM2 mRNA is presented as the mean ± S.E.M. and is shown as the relative expression level according to the 2 Δ Δ Ct method. The data were analyzed via one-way analysis of variance followed by Bonferroni post hoc correction. # p < 0.05, ## p < 0.01, ### p < 0.001, 5 × FAD young group (3–4 months old) vs. other 5 × FAD groups. * p < 0.05, ** p < 0.01, *** p × 0.001, WT vs. 5 × FAD. (B) Representative western blot of TREM2 in whole hippocampal lysate, expressed as the mean ± S.E.M. (C) Representative confocal photomicrographs of the whole hippocampus. Note the increased immunoreactivity of TREM2 (red) in the hippocampus, especially in the CA1 region, of 5 × FAD mice. The expression increased with age. Scale bar = 200 μm. (D) Dual-label immunofluorescence image showing TREM2 (red) with microglia (green). Nuclei are stained with DAPI (blue). Notably, the increased immunoreactivity of TREM2 (red) was mostly associated with activated microglia (Iba1, green) rather than with resting microglia. Scale bar = 10 μm. (E) Immunohistochemistry (IHC) showing Aβ deposits stained with 6E10 in WT mice and 5 × FAD mice. Scale bar = 100 μm. (F) Dual-label immunofluorescence showing that TREM2 (red) is expressed mainly in plaque-associated areas (6E10, green). Scale bar = 10 μm. ns, not significant; TREM2, triggering receptor expressed on myeloid cells-2; WT, wild type; Iba1, ionized calcium-binding adapter molecule 1.

Article Snippet: The sections were then subjected to a blocking solution comprising TBS supplemented with 0.3% Triton X-100, 0.25% bovine serum albumin (BSA), and 5% normal donkey serum (NDS) at room temperature for 2 h. The slices were then incubated with either Iba1 (1:1000; Wako Pure Chemical Industries) and 6E10 (1:5000; Covance), Iba1 and TREM2 (1:100, R&D Systems), or TREM2 and PU.1 (1:300, Cell Signaling) antibodies followed by species-specific Alexa Fluor secondary antibodies (1:1000; Life Technologies).

Techniques: Biomarker Discovery, Expressing, Western Blot, Immunofluorescence, Staining, Immunohistochemistry, Binding Assay