Figure 1.

SIRT2 regulates the brain response to LPS. (A) SIRT2 expression by microglial cells. A representative image of co-localization of SIRT2 with Iba-1 was obtained from brain sections stained for Iba-1 (red) and SIRT2 (green). (B) SIRT2 protein levels in mice injected intracortically with PBS or LPS (0.2 μg). Cortical extracts from mice injected with PBS or LPS were analysed by western blotting and stained for SIRT2. Quantification was done by densitometry analysis of the protein bands and normalized to β-actin levels (mean±s.d.; n=3, PBS; n=5, LPS; t-test, *P<0.05). (C) Microglial activation induced by intracortical injection of LPS. Representative images of brain sections of mice injected with PBS and LPS at day 2 and 7 post injection and stained for Iba-1. (D) Microglial activation in wild-type versus SIRT2^−/− mice. SIRT2^−/−mice show increased microglial activation induced by LPS injection on bright-field and confocal (below) microscopy images of mice brain sections stained for Iba-1. Confocal images reveal thicker microglial processes in SIRT2^−/−mice. (E) mRNA levels of IL-1, IL-6, Mpa2l, TNF, and Ip-10 inflammatory genes were determined by qPCR, normalized to β-actin expression levels. IL-6, Mpa2I, TNF, and lp-10 expression is increased after 6 h in LPS-injected SIRT2^−/−mice compared to LPS-injected WT mice (mean±s.d., n=4; two-way ANOVA, ***P<0.001). (F) Flow cytometry analysis of cortical cells isolated from injected mice and stained for CD11b, CD45 and IL-6 (mean±s.d.; n=3, PBS; n=4, LPS; two-way ANOVA, test, ***P<0.001). (G) Induction of free radicals by LPS. Representative images of wt and SIRT2^−/− mouse brain sections stained for nitro tyrosine 5 days post injection with PBS or LPS (5 μg). (H) Neuronal cell loss induced by LPS. Mice brain sections were stained for NeuN and the number of positive cells counted and normalized to the numbers of cells in the cortice of wt mice 5 days after PBS or LPS (5 μg) injection (mean±s.d., n=3; two-way ANOVA, *P<0.05).