Figure 2.

Quantitative analysis of expression of miR-124 in activated microglia. (a) Analysis of expression of activation markers CD45 and MHC class II in populations of gated F4/80⁺CD11b⁺GFP⁻ microglia (upper row) and F4/80⁺CD11b⁺GFP⁺ peripheral macrophages (lower row) isolated from CNS of healthy chimeric mice (no disease) or chimeric mice with EAE at the disease onset (day 14), peak (day 21) or recovery (day 40). Four to five mice per group were used. The isotype controls are shown in upper left quadrants of contour plots. Percentages of CD45⁺ cells (right quadrants) and MHC class II⁺ cells (upper quadrants) are shown. (b) Comparison of miR-124 expression in populations of resting CD45^lowMHC class II⁻GFP⁻ and of activated CD45^int–hiMHC class II⁺GFP⁻ microglia, as well as populations of activated CD45^hiMHC class II⁺GFP⁺ and deactivated CD45^int–hiMHC class II⁻GFP⁺ peripheral macrophages. The cells were sorted by FACS from the CNS of chimeric mice with EAE at day 14, and miR-124 expression was assessed by real-time qRT-PCR. Representative results from two independent experiments are shown, with mean ± s.d. of quadruplicate wells plotted; **P < 0.01. (c) Analysis of expression of miR-124 in microglia activated in vitro by GM-CSF and IFN-γ or LPS and IFN-γ. Microglia cells were isolated from healthy mice and incubated in medium alone, with GM-CSF and IFN-γ, or with LPS and IFN-γ for 6 h, and miR-124 expression was assessed by real-time qRT-PCR. Mean ± s.d. of triplicate experiments is shown; *P < 0.05 compared to ex vivo–isolated microglia.