Figure 1.

A1 KO in microglia caused an increase in Aβ1–42 clearance. A, Microglia were isolated from neonatal wild-type (WT) or Acat1^−/− knock-out (A1 KO) mice. ACAT1 protein and activity were analyzed by Western blot and by ACAT activity assay, respectively. In ACAT activity assay, cells were preincubated with or without 1 μm of the ACAT1-specific inhibitor K604 for 24 h. Data are mean ± SEM of two experiments. ***p < 0.001. n.s., Not significant. B, C, G, WT or A1 KO microglia were incubated with 0.5 μm Aβ1–42 for the indicated time. B, The remaining Aβ1–42 in the media was separated by tricine-SDS-PAGE and analyzed by Western blot. Representative blot is shown. C, Oligomeric Aβ1–42 levels (monomer + dimer + trimer + tetramer) in the media were quantified with ImageJ software. Data are mean ± SEM of four experiments. **p < 0.01. ***p < 0.001. D, Aβ1–42 was conjugated to Cy3 dye as described in Materials and Methods. The Cy3-labeled Aβ1–42 (Cy3-Aβ1–42) and unlabeled Aβ1–42 were separated by tricine-SDS-PAGE and analyzed by Western blot. E, WT or A1 microglia were incubated with 0.5 μm Cy3-Aβ1–42 for the indicated time. Cells were washed several times, and intracellular Aβ1–42 levels were analyzed by flow cytometry. Data are mean ± SEM of five experiments. *p < 0.05. F, Expression levels of ABCA7, CD36, and SRA were examined in WT and A1 KO microglia by quantitative PCR (qPCR). Data are mean ± SEM of three experiments. n.s., Not significant. G, WT or A1 microglia were pretreated with 400 μg/ml of fucoidan for 1 h and then incubated with 0.5 μm of Cy3-Aβ1–42 for 3 h in the presence of the same inhibitor. Cells were washed several times, and intracellular Cy3-Aβ1–42 levels were analyzed by flow cytometry. Data are mean ± SEM of four experiments. **p < 0.01. H, Intracellular Aβ1–42 levels in WT and A1 KO microglia at the indicated time point were analyzed by ELISA. Data are mean ± SEM of three experiments. *p < 0.05. **p < 0.01.